Bioconvection phenomena for MHD Williamson nanofluid flow over an extending sheet of irregular thickness are investigated theoretically, and non-uniform viscosity and thermal conductivity depending on temperature are taken into account. The magnetic field of uniform strength creates a magnetohydrodynamics effect. The basic formulation of the model developed in partial differential equations which are later transmuted into ordinary differential equations by employing similarity variables. To elucidate the influences of controlling parameters on dependent quantities of physical significance, a computational procedure based on the Runge–Kutta method along shooting technique is coded in MATLAB platform. This is a widely used procedure for the solution of such problems because it is efficient with fifth-order accuracy and cost-effectiveness. The enumeration of the results reveals that Williamson fluid parameter  Nanofluids have a lot of thermophysical attributes like improved heat conductivity, heat diffusivity, and viscosity as opposed to their common base liquids, such as oil or water. Nanofluids applications embrace mass and thermal transportation in engineering and industrial appliances, coolant in automotive electronics, such as microscale, microchips, etc. . The ideIn a wide range of technical, automobile, and regular housekeeping machinery, non-Newtonian nanofluids are used. Non-Newtonian fluids are those fluids that do not obey Newton’s viscosity law, i.e., continuous viscosity independent of stress. Under the impact of external forces, the viscosity of non-Newtonian fluids can change to either being more fluid or more solid . MathemaBioconvection is the study of structure formation built up due to motion of the swimming microorganism. Usually, the gyrotactic microorganism form a dense layer on the upper part of liquid and create density variation. The heavy layer breaks, and the microorganism come down to create a current of upward and downward motion. This phenomena is called bioconvection. In the modern era of biotechnology along with nanofluids are employed in the diagnosis of various dangerous diseases. Firstly, in 1961 Platt  studied Thermal radiation impact in heat transfer has a wide range of applications in thermal engineering, such as gas turbines, nuclear power plants, and numerous propulsion devices for space vehicles, satellites, missiles, and aircraft. Moreover, thermal radiation impact is certain for space applications where few devices are sketched to move levels to attain high thermal efficiency at high-temperature. For this reason, the radiation impact is important while determining thermal impacts in the processes having high-temperature . Pop et How do the transport behaviors change for Newtonian fluid as compared with non-Newtonian (Williamson fluid)?What is the impact of nanofluid slip parameters (Brownian and thermophoresis) on temperature?How do the bioconvection parameters influence the flow of fluids?A glance at the existing literature convinces that heat and mass transfer across an irregular geometry is rarely discussed with bioconvection for non-Newtonian Williamson nanofluids. The flow, temperature and concentrations distributions are investigated owing to a slender stretching sheet of varying thickness. Temperature dependent viscosity, thermal conductivity and radiations develop significant aspects of this work. The central idea pertains to the improvement in thermal transport for heat exchangers of compact heat density. The heat transfer is limited to conventional modes with less thermal conductivity of base fluid. Present work is perceived to enhance thermal conductivity of bulk fluid with homogeneous mixture of dilute nano particles. Bioconvection of gyrotactic self motive microorganisms is also useful aspect of this communication to avoid possible settling of nano entities. The following queries are sought:It is revealed that velocity for non-Newtonian (Williamson nanofluid) is solute as compare to Newtonian fluid, whereas the temperature behaves oppositely. Additionally, the biconvection parameter a, b and c are constants and used for the utterance of n delegates the velocity power parameter. In this problem, the sheet is adequately thin in contrast to its width We consider a steady magnetohydrodynamic two-dimensional laminar flow of an incompressible Williamson nanofluid over an extending surface issuing from a slit at the origin through which the sheet is drawn through the fluid medium. Thermal distribution through Williamson nanofluid transportation owing to the non-linear stretch in horizontally, lying slender sheet is taken into consideration. It is considered that the sheet is not flat. Magnetic field Equation of continuityBy consideration, a linear temperature dependent for thermal physical quantities The Rosseland approximation for radiation is (7)qr=−4The dimensional boundary conditions are (10)u^–(11) involve partial differentiation. It is very difficult to solve in this form. Therefore the following similarity transforms  are utilIn view of the above appropriate relations Equation  is satisIn order to yield simplification, we suppose,These transformations modify the Equations (13)–(16) as belowk signifies the thermal conductivity of the nanofluid, and This segment describes the attributes of important physical quantities of engineering interest. The local skin friction Skin friction coefficient in non-dimensional form isBy using Equations (10), (12) and (18), we get the following expressionsThe non-linearity characteristics of the fluid model described finally as boundary value formulation in Equations (19)–(22) with initial and boundary constraints (23) can not be solved analytically. In order to seek physical insight of the problem boundary value problem is required to yield a viable solution. The characteristics for heat and mass transportation as influence by the leading parameters help to understand the very nature of the problem. A reliable numerical procedure is sought for this purpose. Several investigators ,45 emploalong with the boundary conditionsM, n, Exploration for bioconvection of Williamson’s nanofluid over a horizontal extending surface with variable thickness is enumerated in presence of magnetic field and thermal radiation. The numerical findings as achieved from the above code are exhibited and elaborated in this segment. The data enlisted in wtonian) .M is caused due to the resistive force (Lorentz). This reactive force comes to play its role when magnetic field interacts with electric field. This slowing of flow helps the conservation for loss of kinetic energy to heat energy and, hence, the enhancement in temperature n (non-linear index) provide boosting to In n and mixed convection parameter M, The flow velocity enhances with higher inputs of the power index parameter M, n, The temperature field rises directly with The microorganism density is depreciated when the parameters M, n, The skin friction is intensified with Nusselt number is enhanced in direct relation to The implications of temperature-dependent viscosity and thermal conductivity for heat and mass transport of Williamson nanofluids across a non-linear stretching sheet of irregular thickness are examined when bioconvection of microorganisms is incorporated. Because of the inherent non-linear characteristics of the formulation developed for the problem, a numerical code based on the Runge–Kutta method is run on the Matlab platform. A reasonable agreement is achieved among the past and current studies to validate the results. The notable outcomes are presented below:


Patients. This clinical report describes the detailed prosthodontic management of a 23-year-old male patient suffering from soft tissue complication following the placement of a 3d-printed titanium patient-specific implant. This implant was implemented simultaneously with the resection of a calcifying cystic odontogenic tumor related to the maxillary arch. Later, soft tissue dehiscence and implant exposure were encountered with subsequent food impaction, infection, and pus discharge. The treatment plan was to fabricate removable partial denture. The prosthesis was planned to be retained by bar and clip attachment on the patient-specific implant side, while on the other side, the removable prosthesis was allowed to engage two abutments with an embrasure clasp assembly in addition to covering the palatal tissues to offer protection for the soft tissue dehiscence against food impaction. Discussion. Soft tissue dehiscence and implant exposure are among the frequently reported complications associated with the patient-specific implant. The resulting infection complicates the prognosis of the implemented implant and necessitates, in some occasions, its removal. The selection of the removable prosthesis to cover soft tissue dehiscence was a conservative alternative to the implant removal as it protects the exposed titanium surface from food impaction while maintaining the implant functionality. Conclusion. Three-year follow-up showed complete resolution of the patient's complaints while fulfilling the patient's aesthetic and functional demands and indicates that the use of detachable overlay prosthesis could be one of the proposed treatment options. For the past decade, facially anchored osseointegrated implants  and freeThanks to the advances in imaging modalities and manufacturing techniques, patient-specific implants (PSIs) have been proposed to overcome the pitfalls associated with the previously mentioned treatment options . InitialCiocca and his colleagues in 2012  were theDespite all attempts to enhance bone and soft tissue healing around PSI restorations, soft tissue dehiscence and recurrent infection have been reported in multiple studies , 16, 17.A 23-year-old male patient was referred to F.G. Maxillofacial Prosthodontics Unit, Faculty of Dentistry, Cairo University, after undergoing surgical installation of patient-specific implant (PSI) following surgical resection of calcifying cystic odontogenic tumor CCOT related to the left maxillary arch extending to the orbital floor. The patient was complaining of hygiene-related complications as a result of extensive soft tissue exposure of PSI.Upon reviewing the patient's records, it was revealed that the patient was scheduled for total maxillary resection with simultaneous reconstruction using patient-specific implant prosthesis. With the aid of digital software , the lesion was virtually resected, and the patient-specific implant was designed with maximum extensions engaging the whole maxillary surfaces superiorly, medially, and laterally. Finally, the implant was printed with an EBM machine  into medical grade 23 titanium Ti–6Al–4V, after then it was subjected to several procedures of surface treatment and sterilization.At the time of tumor ablation surgery, the implant was installed and stabilized using 2 mm grade 5 titanium fixation screws on the facial bones extending from the orbital floor to the palate. The palatal soft tissue on the tumor side was preserved and used as a soft tissue drape to cover the implant surface. The patient did not receive any adjunctive treatment before or after the surgery. Three months after the surgery, the patient received a five-unit fixed partial denture restoration cemented on the extruding metal posts.Six months later, the patient presented with soft tissue dehiscence and complained of frequent food impaction and persistent foul odor. The patient was instructed to rinse daily with antiseptic mouthwash till the condition subsides in addition to strict routine oral hygiene measures. By the age of 23, implant surface exposure has become more extensive, and the patient suffered further hygiene-related complications. Accordingly, the surgical team decided to remove the PSI. However, based on the patient's desire not to get involved in further surgical interventions, he was referred to our team for possible prosthetic management.Data collected from patient interviewing, clinical examination, and radiographic imaging revealed that PSI was stable, functioning, and pain-free. Titanium surface was exposed and extending along the buccal and palatal surfaces of PSI .Diagnostic impressions were made for both arches by using irreversible hydrocolloid impression material . A customized maxillary tray was fabricated by using cold cure acrylic resin  over which the definitive impression was made by using a two-step impression technique with rubber base impression material . Inlay pattern resin  supported with metal wires was used to pour the metal posts portion of the impression while the rest was completed with a hard stone. Modeling wax  occlusion rims were adjusted intraorally over cold-cured acrylic resin record bases, and the interocclusal record was made at maximum intercuspation. Casts were mounted on a semiadjustable articulator with a face bow transfer. Wax-up of acrylic teeth was done for diagnostic and treatment planning purposes.A condensation silicon putty index was adapted on the buccal aspect of the diagnostic wax-up to determine the prosthetic space available occlusocervically and buccolingually, the size of the artificial teeth, and its relation to the metal posts. A 20 mm interocclusal distance was revealed with sufficient space between the metal posts and the buccal aspect of the future teeth. Surveying of the cast was carried out to determine the amount of natural teeth undercuts, guiding planes, and the potential path of insertion.Accordingly, the plan was to exclude the middle post and fabricate a bar and clip attachment connecting the two distant metal posts to support a partial overdenture. Embrasure clasp assembly was planned on the maxillary right first molar and maxillary right second premolar to aid for retention, support, and bracing in addition to a palatal plate extending over the palatal surfaces of the maxillary right first premolar, canine, lateral incisor, and central incisor and maxillary left central incisor to the areas of soft tissue dehiscence at the patient-specific implant. Before the execution of the planned prosthetic management, the patient was instructed to follow comprehensive oral hygiene measures. These instructions included rinsing with chlorhexidine mouthwash twice daily and brushing the PSI with soft toothbrush and chlorhexidine gel. Besides that, he was encouraged to use dental floss between the remaining teeth and tooth brushing by soft toothbrush twice daily. Empirical antibiotics were prescribed to the patient to manage any possible infection at the exposure site.The planned mouth preparation was executed as follows: cutting the middle post by using a transmetal bur under copious irrigation, preparation of a mesial occlusal rest seat on the maxillary right first molar, and preparation of a distal occlusal rest seat on the maxillary right second premolar. Additionally, there is a mesial guiding plane on the proximal surface of the maxillary left central incisor. The definitive impression and mounting procedures were performed with the same techniques mentioned earlier. OT Bar multiuse  was waxed, cast into Co-Cr, and then cemented on the metal posts . Metal fThe trial setting of artificial acrylic teeth guided with the diagnostic wax-up was done and tried intraorally. The approved wax trial tooth arrangement was flasked and packed by using heat-cured acrylic resin . At the time of bar and clip attachment pickup, the undercut beneath the bar was blocked out using a putty consistency rubber base to avoid the unnecessary flow of the pickup material. After insulating the polished surface of the prosthesis, a doughy mix of cold-cure acrylic resin was added to the intaglio surface, and a medium retention clip was picked up under maximum intercuspation.The prosthesis was finished and polished, and the patient received instructions regarding prosthesis insertion, removal, and maintenance . AdditioThe patient was recalled monthly for the first year and every three months during the following years. During this period, the patient received professional intraoral cleaning and debridement every 3 months. After 3 years of follow-up, no signs or symptoms of hygiene-related complications were observed clinically or radiographically . AdditioSurgical reconstruction is nowadays considered a well-accepted treatment modality for the rehabilitation of oral defects, and it is achieved either by using free grafts, zygomatic implants, or patient-specific implants. Advances in volumetric imaging and 3d planning before the ablation surgery allow the surgeons to make more accurate decisions regarding the procedure and give the chance to manufacture personalized implants with the help of additive manufacturing technology to fit more readily during surgery.In this patient, the implant was 3d printed from Ti6Al4V. The surfaces of PSI facing the bone were subjected to sandblasting and chemical treatment procedures in an attempt to enhance osseointegration, while the surfaces facing the soft tissue were left unpolished after the 3d printing procedure. Although it was hypothetically claimed that the unpolished surface facing the soft tissue would decrease the possibility of its dehiscence , our patDespite the several attempts to manage the condition by medication, debridement, and daily oral hygiene measures, the soft tissue complications were persistent and bothersome to the patient. Custom implant removal was the first line of treatment proposed by the surgical team. Accordingly, an immediate restoration of the resultant surgical defect was addressed either using immediate surgical obturator or surgical reconstruction. Several authors  have sugOn the intact side, the removable prosthesis was allowed to engage two abutments with an embrasure clasp assembly in addition to covering the palatal tissues to offer cross arch stabilization. On the implant side, the presence of enough prosthetic space enabled the use of a bar and clip attachment, which fulfilled the high retention demands in comparison to other attachment systems .The main limitation associated with the chosen treatment option is the lack to provide a permanent soft tissue closure but rather preventing food impaction which is highly dependent on the patient compliance. The patient was instructed to follow strict oral hygiene measures; however, it was not anticipated to experience significant food impactions beneath this rigid design unlike soft tissue-supported ones. Another addressed concern is the need for continuous follow-up clinically and radiographically to ensure the patient's compliance and to detect early signs of possible complications. The possibility of discarding the prosthesis and the removal of the PSI was also discussed with the patient in case of the development of persistent infection, which could be life threatening.Patient-specific implants may be regarded successful in terms of fixation and stability; nevertheless, soft tissue dehiscence is a serious complication that should be anticipated and managed early during the treatment. The use of detachable overlay prosthesis can be considered a promising solution to conservatively overcome the hygiene-related complications while fulfilling the patient's aesthetic and functional demands.


Resin/reinforcement wetting is a key parameter in the manufacturing of carbon nanotube (CNT)-based composite materials. Determining the contact angle between combinations of liquid resin and reinforcement surfaces is a common method for quantifying wettability. As experimental measurement of contact angle can be difficult when screening multiple high-performance resins with CNT materials such as CNT bundles or yarns, computational approaches are necessary to facilitate CNT composite material design. A molecular dynamics simulation method is developed to predict the contact angle of high-performance polymer resins on CNT surfaces dominated by aromatic carbon, aliphatic carbon, or a mixture thereof (amorphous carbon). Several resin systems are simulated and compared. The results indicate that the monomer chain length, chemical groups on the monomer, and simulation temperature have a significant impact on the predicted contact angle values on the CNT surface. Difunctional epoxy and cyanate ester resins show the overall highest levels of wettability, regardless of the aromatic/aliphatic nature of the CNT material surface. Tetrafunctional epoxy demonstrates excellent wettability on aliphatic-dominated surfaces at elevated temperatures. Bismaleimide and benzoxazine resins show intermediate levels of wetting, while typical molecular weights of polyether ether ketone demonstrate poor wetting on the CNT surfaces. Polymer matrix composite (PMC) materials are widely used as structural materials in aerospace and aeronautical vehicles. A large number of high-performance polymers have been developed for use as the matrix in PMCs with various carbon reinforcements such as carbon fiber, carbon nanotubes (CNTs), and graphene nano-sheets. Recently, CNTs have shown exceptional mechanical properties and are being considered as a reinforcement material for the next generation of ultralight high-strength composite materials ,2. Many Wettability of resin onto the reinforcement surface is a key parameter in the manufacturing of PMCs. Determining the contact angle between specific combinations of liquid resin droplets and reinforcement surfaces is a common method to quantify wettability. However, the experimental measurement of contact angle can be difficult or cost prohibitive when screening multiple high-performance resins with CNT-based materials such as CNT bundles or yarns. Furthermore, the interpretation of experimental wetting results for some CNT materials is complicated by the presence of regions of amorphous carbon  on the aromatic CNT surface. Efficient computational prediction of contact angle could significantly facilitate the resin selection process for the design and development of new CNT-based PMCs.Molecular Dynamics (MD) simulation has been previously used to study the wetting properties of water and polymer resin systems on surfaces such as graphite and metals ,20,21,22The objective of this study is to use MD simulation to predict the relative levels of wettability of various high-performance resins onto CNT surfaces. All-atom simulations are used to determine the contact angle with aromatic and aliphatic carbon surfaces for a range of temperatures to provide guidelines for resin selection and process design. In the following sections, the details of the materials and the modeling procedures are outlined. The results of the simulations are subsequently presented for a comprehensive understanding of resin wettability with CNT surfaces. The results indicate that resin and temperature selection can have a significant impact on resin wettability. Bismaleimide (BMI): This is a two-part system consisting of 4,4′-bismaleimidodiphenylmethane (BMPM) and O, O’-diallyl bisphenol A (DABPA), shown in Benzoxazine: The Bisphenol-A Benzoxazine monomer is shown in Difunctional epoxy: This is a two-part system consisting of diglycidyl ether bisphenol F (DGEBF) and diethyltoluenediamine (DETDA) monomers, as shown in Tetrafunctional epoxy: This is a two-part system consisting of tetraglycidyl methylene dianiline (TGMDA) and diaminodiphenyl sulfone (DDS) monomers, as shown in Fluorinated cyanate ester: The hexafluorobisphenol A cyanate ester monomer is shown in Non-Fluorinated cyanate ester: The phenol novolac cyanate ester monomer is shown in PEEK monomer: The polyether ether ketone monomer is shown in PEEK dimer: The polyether ether ketone dimer is shown in The wetting behavior of eight polymer resin systems was simulated in this study:The selection of these monomers involves several contrasts that provide insight into the wetting behavior of polymer resins with CNT surfaces. The list of monomers includes a thermoplastic (PEEK) and six thermosets. For the PEEK thermoplastic, the monomer and dimer versions were modeled. The purpose of this comparison is to understand the role of molecular weight in wetting. The two cyanate ester systems were chosen to determine the role of fluorine on the wetting behavior. The two epoxy systems were chosen to establish the role of the epoxide group density on the wetting. The inclusion of one-part thermosets  and two-part thermosets (epoxies and BMI) provide insight into the effect of monomer mixtures on the wetting behavior. Finally, it is important to note that all of these resins represent aerospace-grade systems that can be used for a wide range of CNT-based composite applications.2 carbon bonding;Aromatic surface with pure sp3 carbon bonding near the surface and hydrogen terminations at the surface.Aliphatic carbon surface with spTwo carbon-based surfaces were simulated in this study: 2 carbon (graphite), sp3 carbon (diamond), and hydrocarbons [A wide range of amorphous carbon structures exist, which contain various levels of three principal carbon forms: spocarbons . As it iThe Large scale Atomic/Molecular Massively Parallel Simulator (LAMMPS) software package was used to perform the MD simulations in this study. The Interface Force Field (IFF) developed by Heinz et al.  was utilx- and y-directions. The positions of the carbon atoms on the graphene planes were obtained by generating an array of atoms in a graphene lattice structure in LAMMPS. A lattice constant of 2.46 Å was used with the primitive lattice vectors given by Gray et al. [Previous studies have shown that pseudo-2D simulations for wetting are efficient and capable of generating accurate results ,15,21. Sy et al. . Virtualy et al. ,27. A dey et al. . The aroA diamond lattice structure was used to create the aliphatic carbon surface model . The thiThe MD wetting simulations consisted of two steps: monomer droplet formation and monomer–surface contact simulation. The modeling steps are schematically represented in For the first step Step I, , the resijr is the distance between atoms i and j, ijσ is the equilibrium distance between atoms i and j, ϵij is the well depth parameter which is calculated as the geometric mean of individual atom parameters for atoms i and j, and cr is the specified cutoff distance. Columbic interactions were calculated using [ijr is the distance between atoms i and j; C is the energy unit conversion constant; iq and jq represents the atomic charges of atoms i and j, respectively; cr is the specified cutoff distance.In the second step of the simulation process Step II, , the resed using (2)E = CThese simulations were performed for 2 ns and are henceforth referred as the ‘contact simulations.’ The contact simulations were performed for a simulation time of 2 ns in the NVT ensemble at a simulation temperature selected from the temperature range of 87–207 °C with a time step size of 1 fs. The contact angle was predicted for each resin system, surface type, and temperature. The method used here to determine the contact angle value is similar to the method developed by Sumith et al. . An arraThe interaction energies (IE) between the droplet and the surface were determined in LAMMPS using the “compute group/group” command to gain physical insight on the contact angle values for the aromatic and aliphatic surfaces. In LAMMPS, IE is calculated byTo calculate the IE between the monomers and surface, monomer droplet atoms and surface atoms were assigned to two different groups in LAMMPS. Calculated IE values were averaged over the final 200 ps of the contact simulation. In addition, the atoms of each functional group present in the monomer were assigned to a respective subgroup. IE values between these subgroups and the surface were calculated using Equation (3), where A was assigned as the group of surface atoms and B was assigned as the atoms in a specific functional group in the system. The calculated IE values represent the individual contributions of the surface interaction of specific monomer functional groups towards the total IE values. Higher negative magnitudes of IE indicate higher levels of interaction between the monomers/group and carbon surface.It is helpful to interpret the predicted wetting behavior as a competition between adhesion and cohesion. That is, the monomers are attracted to both the carbon surface and to each other. It is the relative strength of these two interactions that dictates how the resin wets onto the carbon surface. Specifically, if the monomer’s interaction with the surface (adhesion) is in some way diminished, then monomer–monomer cohesion becomes dominant and will cause the droplet to form a tighter sphere, increasing the contact angle. If the adhesion is in some way enhanced, then the monomer–monomer cohesion plays a more minor role and the droplet will no longer hold together in a tight sphere, thus decreasing the contact angle.The decrease in contact angle with increasing temperature shown in To investigate improved wettability of tetrafunctional epoxy on the aliphatic surface at elevated temperatures, the IE of each functional group with the surface was determined. The fluorinated cyanate ester monomers include trifluoromethyl groups that may provide a steric hinderance that restricts mobility, thus providing resistance to the spread of the monomer droplet over the aromatic carbon surface in this temperature range. The contact angles for the two systems on the aromatic surface are nearly the same for temperatures above about 130 °C. For the aliphatic surface, the curve fit line of the non-fluorinated system consistently shows a lower contact angle than that of the fluorinated system, however, examination of the overlap of the error bars indicates that there is no statistical difference in the contact angle of the two systems. Furthermore, considering the overlap of the error bars, there is no consistent difference in the contact angles of either system when comparing the two carbon surfaces. The detailed IE contribution per functional group analysis for both cyanate esters is provided in the SI.Other important information can be determined from the data in Other important conclusions can be discerned from An MD framework was developed to predict the contact angle values of polymer resins on CNT surfaces dominated by aromatic carbon, aliphatic carbon, or a mixture thereof (amorphous carbon). The monomer chain length, temperature, and the functional groups in the monomers have a significant impact on the contact angle values on the CNT surface. An overall trend of decreasing contact angle (improvements in wetting) with increasing temperature was observed for all the resins studied. Considering the results from this study, the difunctional epoxy demonstrates the best wettability of the thermoset resins considered herein, regardless of the CNT material surface structure. However, for applications that require relatively high temperatures, the difunctional epoxy may be unsuitable due to its lower glass transition temperature relative to the tetrafunctional epoxy ,37 and t


In order to better understand how the immune system interacts with environmental triggers to produce organ-specific disease, we here address the hypothesis that B and plasma cells are free to migrate through the mucosal surfaces of the upper and lower respiratory tracts, and that their total antibody repertoire is modified in a common respiratory tract disease, in this case atopic asthma. Using Adaptive Immune Receptor Repertoire sequencing (AIRR-seq) we have catalogued the antibody repertoires of B cell clones retrieved near contemporaneously from multiple sites in the upper and lower respiratory tract mucosa of adult volunteers with atopic asthma and non-atopic controls and traced their migration. We show that the lower and upper respiratory tracts are immunologically connected, with trafficking of B cells directionally biased from the upper to the lower respiratory tract and points of selection when migrating from the nasal mucosa and into the bronchial mucosa. The repertoires are characterized by both IgD-only B cells and others undergoing class switch recombination, with restriction of the antibody repertoire distinct in asthmatics compared with controls. We conclude that B cells and plasma cells migrate freely throughout the respiratory tract and exhibit distinct antibody repertoires in health and disease. The respiratory tract mucosa serves as a physical barrier composed principally of pseudostratified cells, with ciliated columnar epithelial cells resting on a basement membrane along with mucin-producing goblet cells. The cellular composition of the lamina propria underneath the epithelial layer varies according to the site within the respiratory tract but includes blood vessels and lymphatics, structural cells and a range of immune cells including innate immune cells, antigen-presenting and effector cells, cartilage and, in the lower respiratory tract, smooth muscle cells. Lymphoid tissue surrounds the upper respiratory tract , while lymph nodes are abundant adjacent to the lower airways in the mediastinum and adjacent to the trachea and bronchi. Lymph flows towards the hilum and ultimately returns to the systemic venous circulation via the right lymphatic and left thoracic ducts.The respiratory tract is located inside the body but, like the digestive and reproductive tracts, is constantly exposed to the external environment with its plethora of stimuli from microorganisms, proteins including allergens and other particulate matter including particulate pollution. The respiratory tract is divided into upper and lower regions above and below the larynx, the former comprising the nasal cavity, paranasal sinuses and pharynx and the latter the trachea and bronchial tree. It is also contiguous with the skin , is still debated, although it is widely accepted that BALT can be “induced” (iBALT) in association with many chronic inflammatory diseases of the airways such as COPD –4. In advia the lymph nodes and lymphatics is relatively well established, little is known about the capacity of individual immune cells such as B cells and plasma cells to migrate freely between the mucosal surfaces and lymphatics of the upper and lower respiratory tracts. According to the concept of the “United Airways Disease” (UAD) treatment of disease at one site in the respiratory tract, such as allergic rhinitis, will result in improvement in symptoms in another, such as the lower respiratory tract in allergic asthma  and one non-atopic, non-asthmatic (NANA) control subject using Adaptive Immune Receptor Repertoire sequencing (AIRR-seq). Detailed analysis of the sequences provided conclusive evidence that B cells are able to migrate both locally and to more distal sites within the bronchial mucosa of the same lung . The stuIn the present study a primary goal was to extend these observations by addressing the hypothesis that B cells in the upper  and lower  respiratory tract mucosa of both lungs are directly related. We compared the repertoires of the B cells obtained near contemporaneously from nasal biopsies (NBx) and bronchial mucosal biopsies (BBx)  and the peripheral blood from patients with atopic asthma (AA) and non-atopic, non-asthmatic control subjects (NANA). Our secondary aim, through detailed analysis of somatic hypermutation of the B cell antibody genes, was to attempt to align their directions of travel with their maturation history. Thirdly and finally, we wished to confirm and extend our previous observation of IgD-only cells in the bronchial mucosa in a larger cohort and to compare IgD-only cells of the nasal and bronchial mucosa.Six atopic patients with asthma (AAXX) and nine non-atopic, non-asthmatic controls (NANAXX) were enrolled in the study . From ea6 PBMCs from each blood sample was carefully thawed and RNA extracted using the Promega Maxwell HT simply RNA custom kit. RNA recovery and quality were estimated using Agilent TapeStation and HSRNA ScreenTapes.Total RNA was extracted from one of the paired nasal biopsies and one bronchial biopsy from each of four sites within the bronchial mucosa using a Qiagen RNEasy plus kit. One aliquot of 10The RNA was used for heavy chain immunoglobulin amplification and high-throughput sequencing using the previously described AbSeq protocol . Brieflyhttp://immcantation.org) with the Presto v0.5.7 .We enrolled six atopic asthmatics (AA) and nine non-atopic, non-asthmatic control subjects (NANA) into the study see Figure 1versus 54 only found in the three lung sites). These expanded clones were often large (up to 512 unique sequences) compared to the local clones found in only one site . Connectivity analysis , we analyzed the pattern of trafficking events between tissues along B cell lineage trees . Briefly, given the tissue  associated with each observed sequence in a lineage tree, the algorithm determines the set of internal node tissue states requiring the fewest number of trafficking events along the tree. The types of trafficking events  that occur within each tree are then compared to those obtained from the same tree topologies with randomized tissue states . We firsThe clonal sequences confined to the left or right bronchial tree or carina shared similar characteristics in terms of mutation frequency, expansion, isotype distribution and numbers of isotypes within a clone . Estimating the selection strength separately for nasal and bronchial biopsy samples, we still found statistically lower selection pressure scores in the CDRs of asthmatics compared to controls   and a negative selection pressure in the framework regions (FWR) consistent with antigen-driven selection in both asthmatics and control subjects (Diversity in antigen specificity and effector function is caused by the recombination of a variety of different IGHV-genes and constant region gene (IGHC), respectively. We first examined the isotype usage and found that in all subjects, atopic asthmatics as well as non-atopic controls, the nasal mucosa was dominated by IgA  followed by IgG (mean 10.3% of unique sequences), but also included small amounts of IgM (mean 2.4% of unique sequences) and IgD (mean 1.4% of unique sequences)  . This indicates that sequences from the same tissue  were more similar than would be expected by chance. Using the same method, we further identified a significantly greater proportion of trafficking events from nasal to bronchial mucosa than in randomized trees in 6/6 atopic asthmatics, and in 7/9 non-atopic control individuals  – sequences from clones found in the nasal but not in the bronchial mucosa, global nasal sequences (NBx_G) – the nasal mucosa sequences from clones that spanned the nasal and bronchial mucosa, global bronchial sequences (BBx_G) – the bronchial sequences from clones that spanned the nasal and bronchial mucosa and non-global bronchial sequences (BBx_NG) – sequences from clones found in the bronchial but not nasal mucosa. We used the previous evidence of predominant directional trafficking from the nasal to the bronchial mucosa to track selection working on B cell clones leaving the nasal mucosa and clones entering the bronchial mucosa. We first observed that the B cell clones leaving the nose (NBx_G) are similar to the ones only found in the nasal mucosa (NBx_NG) in terms of isotype composition with a mean of 86% IgA, 10% IgG and smaller amounts IgD and IgM  compared to the non-global clones from both tissues (NBx_NG and BBx-NG) for IgA and IgG . This region is not only the most variable region of an antibody including significant variability in length, but it is also usually critical for antigen binding and therefore a major site for affinity maturation. We found that the CDR3 length was significantly increased in both nasal and bronchial global sequences of IgA and IgG isotype   than in the bronchial mucosa and was similar between the asthmatics and the control subjects (p=0.48). A mean of 20% of the IgD-only clones from each individual asthmatic, and 15% in individuals from the control group, were shared between the nasal and bronchial mucosa. The IgD-only mutation frequency was very high (11-12% on average) in both groups and at both sites . This revealed that IgD-only B cells from both the bronchial and nasal mucosa showed evidence of a significantly stronger negative selection pressure in the CDR1 and CDR2 regions compared with that of switched isotypes (IgA and IgG) and IgM  (We previously found that the ratio of replacement to silent mutation rates in the CDRs (a measure of selection strength) was negatively correlated with the number of mutations within a clone . This inIn this study we employed Adaptive Immune Receptor Repertoire (AIRR) analysis to compare the antibody repertoires of B cells at various sites in the upper (nose) and lower (lung) respiratory tract mucosa of atopic asthmatic patients and non-atopic controls in order to gain deeper insight into the mechanisms underlying immune defense and disease pathogenesis in asthma. In summary, our data and analysis clearly demonstrate passage of B cells between the upper and lower respiratory tract mucosa, dominantly from the nasal to the bronchial mucosa, and mutually between the mucosal surfaces of the right and left bronchial trees. Our observations that B cell clones undergo somatic hypermutation and may undergo isotype switching during the trafficking suggest that it is predominantly activated B cells that traffic. We have also uncovered evidence of site- and maturation-specific selective forces shaping the local B cell repertoire. Furthermore, we confirm the presence of a special subset of IgD-only B cells, most prominent in the asthmatic bronchial mucosa, characterized by a very high degree of somatic hypermutation and with features of selection pressures distinct from those operating on the B cells of other isotypes.We have further clarified the practicability of determining the B cell repertoire in the respiratory mucosa from a limited number of mucosal biopsies from the upper and lower respiratory tracts. This corresponds to our previous findings , 24, buti.e. the nasal mucosa, four different sites within the bronchial mucosa ranging from the carina down to the 4th generation bronchi of the lower lobe as well as the peripheral blood. There was no detected directional bias for trafficking between the two lungs but clones shared between the left and right bronchial mucosa were more likely to be related to cells in the nasal mucosa, suggesting that the shared clones were selected for mobilization, most likely following activation by exposure to specific antigen  mucosa of non-atopic control subjects compared with the atopic asthmatics .en route to the bronchial mucosa or following entry, but selection against IgA did not seem to occur at the stage of clones leaving the nasal mucosa. Our observation that global clones tended to have a greater mutational load (driven by SHM and affinity maturation) is fully compatible with a scenario of antigen-driven migration of B cells which have undergone proliferation, isotype switching by CSR and affinity maturation. However, as for CSR, it was not possible to determine whether SHM took place en route to the bronchial mucosa, for example in local, draining lymph nodes or other systemic lymphoid tissue, or locally within the mucosa itself. We also searched for directional bias in B cell migration using a parsimony-based phylogenetic test, which takes into account the progressive accumulation of mutations from the germline toward the tips of the clonal linage trees   might co+IgD+ B cells that produce both IgM and IgD with identical VDJ rearrangements through alternative mRNA splicing. These B cells have not yet “experienced” antigen activation, SHM and affinity maturation or CSR, and the vast majority will never do so or survive long enough to be observed. In contrast, the antibodies from IgD-only B cells that we have identified in the bronchial, and to a significantly lesser extent the nasal mucosa, in this and our previous study . IGHJ6 harbors 5-6 tyrosine residues at the 5’ end and since tyrosine residues are the amino acids most commonly involved in antibody-antigen interactions because of their ability to recognize and interact with antigens both by hydrogen bonding and hydrophobic interactions , 33, untStaphylococcus aureus  had detectable serum concentrations of anti-enterotoxin-specific IgEs, evidence that S. aureus had been present in those subjects.In light of our results and reports from clinical studies , 35, we s aureus , 37, eves aureus . Two of Moraxella catarrhalis and Haemophilus influenzae express IgD-binding proteins that can bind to the Cδ1 domain of IgD on naïve IgD-expressing B cells  can be found below: The studies involving human participants were reviewed and approved by London Central Ethics Committee, REC number 15/LO/1800. The patients/participants provided their written informed consent to participate in this study.LO-L and HJG designed the research. CJC, RL and EBC collected the samples. LO-L, BSS and NEGU processed the samples. YJ, CC, TGJ and RH performed the sequencing and initial data processing. HM, KBH, NN and SHK analyzed the data and produced the figures in discussion with LO-L and HJG. LO-L, HJG, SHK and CJC. wrote the paper. All authors contributed to the article and approved the submitted version.LO-L, BS, NU, CC and HG were supported by a research grant from The Medical Research Council, UK (grant number MR/M022943/1) and by the National Institute for Health Research (NIHR) Clinical Research Facility at Guy’s & St Thomas’ NHS Foundation Trust and NIHR Biomedical Research Centre based at Guy’s and St Thomas’ NHS Foundation Trust and King’s College London. The work of SK was supported in part by the National Institute of Allergy and Infectious Diseases of the National Institutes of Health (award number R01AI104739). The work of HG and LO-L was also supported by the award of a Leverhulme Emeritus Professorship to HG (award number EM-2019 027).The views expressed are those of the authors and not necessarily those of the NHS, the NIHR or the Department of Health.SHK receives consulting fees from Northrop Grumman. KBH receives consulting fees from Prellis Biologics. YJ, CC, TGJ and RH are employed by Bristol Myers Squibb.The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.The handling editor has declared past collaborations with one of the authors SHK at the time of this review.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


The ovary is an indispensable unit of female reproduction and health. However, the study of ovarian function in mammals is hindered by unique challenges, which include the desynchronized development of oocytes, irregular distribution and vast size discrepancy of follicles, and dynamic tissue remodeling during each hormonal cycle. Overcoming the limitations of traditional histology, recent advances in optical tissue clearing and three-dimensional (3D) visualization offer an advanced platform to explore the architecture of intact organs at a single cell level and reveal new relationships and levels of organization. Here we summarize the development and function of ovarian compartments that have been delineated by conventional two-dimensional (2D) methods and the limits of what can be learned by these approaches. We compare types of optical tissue clearing, 3D analysis technologies, and their application to the mammalian ovary. We discuss how 3D modeling of the ovary has extended our knowledge and propose future directions to unravel ovarian structure toward therapeutic applications for ovarian disease and extending female reproductive lifespan. Female reproduction hinges on the development of the ovary, the organ which generates mature oocytes and produces hormones to regulate reproductive functions. The ovary harbors the oocyte reserve which supplies mature eggs for the duration of reproductive lifespan and concludes with the cessation of ovulation at menopause. The loss of ovarian function and subsequent drop in gonadal hormones has been associated with enhanced risk for cardiovascular disease , osteopoThe multifaceted functions of the ovary are accomplished by the coordination of diverse cell populations. The operational unit of the mammalian ovary is the follicle, which consists of an oocyte surrounded by granulosa and theca cells. The maturation of follicles from the non-growing or primordial stage toward large antral or ovulatory follicles is governed by signaling between follicles, within the supportive cells of the follicles, and the greater ovarian microenvironment . In addiIn vivo imaging methods such as ultrasound and tomography provide 3D visualization [reviewed in in toto analysis of ovaries  as a group of about 40 primordial germ cells (PGCs) located in the primitive streak near embryonic endoderm of the embryo . DiploidFoxl2 in early fetal stages, develop to granulosa cells in the medulla, and contribute to initial follicle formation during early stages of reproduction , to which a cohort of coelomic epithelial cells migrate and proliferate; these two precursors establish the supporting steroidogenic and interstitial cells of the genital ridges [reviewed in oduction . A seconollicles . Anotheronephros . Signalsonephros .Tie2-expressing endothelial cells revealed that small branches from the vessels at the border of the mesonephros and gonad extend into the primordial gonad at E11.5. With subsequent sex differentiation of the gonads, distinct vascular structures become evident in developing ovaries; without further endothelial cell migration from the mesonephros, blood vessels grow by branching morphogenesis, penetrate the developing ovaries and undergo local organization and remodeling within  . AlthougMvh, Gdf9, and Zp3 to reveal spatiotemporal dynamics of the first wave of folliculogenesis in 3D. Mvh-EGFP was detected earliest in the fetus, persisted in primordial follicles in postnatal ovaries, and extinguished in later stages of folliculogenesis. Gdf9-mCherry positive oocytes were detected starting at PN0. Zp3-AmCyan expression was first seen in oocytes located in the anterior-dorsal region of PN4 ovary, suggesting formation of primary follicles initially occurs in the anterior-dorsal part of the ovary  the need for appropriate microscopes and objectives with long working distances to accommodate large samples, (2) computation and storage required for big data sets, and (3) the opacity of most organs. The invention of confocal microscopy  that have different refraction indices, meaning that the light propagates through them at different speeds. Thus, light waves passing through the ovary scatter heterogeneously between different tissue layers, conferring an opaque or milky appearance. The underlying mechanisms of light scattering in biological samples and different clearing techniques are explained in detail by others . To incrMore than a century ago, The main steps of solvent-based protocols are tissue dehydration, removing lipids, and impregnating the sample with clearing solution to homogenize the refractive index of remaining structures . Zucker in situ relied upon organic solvent-based clearing in BABB to render mouse ovaries transparent after immunostaining  and tetrahydrofuran (THF), known as 3DISCO solvent-based clearing , 2014, oleA2 is an aqueous-based clearing approach that uses detergents instead of hydrophobic solvents to remove lipids while maintaining tissue hydration with urea and glycerol  protocol   , hydrogeYdrogel) . Feng et ovaries . Spatial ovaries . Within Prox1 promoter in order to characterize the spatiotemporal development of the lymphatic network in the ovary , the same hormone treatment contributed to defects in ovarian vasculature, and ultimately decreased ovarian weight and reduced number of oocytes ovulated compared to wild-type; this functionally demonstrated the dependence of follicle maturation and successful ovulation upon vascular remodeling.CLARITY also enabled the mapping of dynamic changes in ovarian blood vessels during folliculogenesis, which would be difficult or impossible to visualize with traditional histology. Work by leA2 were combined to establish the Whole Organ Blood and Lymphatic Vessel Imaging (WOBLI) technique , the broad morphology and follicular structures were maintained in the ovary. In pubertal mice, this protocol allowed for quantification of total vessel length, vessel diameter, vessel straightness and total number of branching points within the ovary  simple immersion in high-refractive index matching solutions, (ii) iDISCO, (iii) CUBIC, (iv) Ethyl cinnamate ‘ECi,’ and (v) combined CUBIC and ECi ‘C-ECi’  could be induced to initiate meiosis by co-culture with fetal ovarian somatic cells, which lead to the formation of reconstituted ovaries (rOvaries) and ultimately the generation of healthy offspring from these in vitro derived oocytes  in 3D. The comprehensive view of the ovary provided by 3D analyses highlights the dynamic changes occurring during development and will inform the roadmap for generating oocytes outside of the body , the field of 3D ovary modeling has adopted a limited number of these techniques. Despite the relative ease of clearing the fetal and adult ovaries, increased fibrosis and stiffness of the ovary with aging  make theThe pioneering studies of the last 15 years paved the way for deciphering the 3D structure and function of the mammalian ovary. In the longer term, the development of non-toxic clearing reagents would enable 3D visualization of specimens for downstream analysis and manipulation in the lab or treatments in the clinic. With the evolution and widespread adoption and combination of tissue clearing technologies, imaging and analysis tools with other cutting edge genomic and machine learning technologies, the field will be poised to unravel the complexity of the mammalian ovary, and ultimately to advance female reproductive health and aging.BS and DJL conceived and wrote the review. Both authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


In this work, a parametric analysis of a dual-mode applicator for microwave ablation treatments is proposed to optimize the geometric parameters of the structure. The dual-mode concept means that the applicator comprises an additional sensing mode to determine the dielectric properties of the surrounding tissue before and during the ablation procedure. Based on numerical electromagnetic-thermal coupled simulations the most optimal design in terms of applicator efficiency as well as ablation zone volume and shape is determined that further fulfills the sensitivity requirements of the sensing mode. The multiobjective optimization problem is solved graphically with the so-called Pareto-optimization method. The resulting Pareto-optimal dual-mode applicator designs are characterized by electromagnetic and thermal simulations and discussed.A multiobjective optimization method for a dual-mode microwave applicator is proposed. Dual-modality means that microwaves are used apart from the treatment, and also for the monitoring of the microwave ablation intervention. (1) The use of computational models to develop and improve microwave ablation applicator geometries is essential for further advances in this field. (2) Numerical electromagnetic–thermal coupled simulation models are used to analyze the performance of the dual-mode applicator in liver tissue; the sensitivity evaluation of the dual-mode applicator’s sensing mode constrains the set of optimal solutions. (3) Three Pareto-optimal design parameter sets are derived that are optimal in terms of applicator efficiency as well as volume and sphericity of the ablation zone. The resulting designs of the dual-mode applicator provide a suitable sensitivity to distinguish between healthy and tumorous liver tissue. (4) The optimized designs are presented and numerically characterized. An improvement on the performance of previously proposed dual-mode applicator designs is achieved. The multiphysical simulation model of electromagnetic and thermal properties of the applicator is applicable for future comprehensive design procedures. Liver cancer is the sixth most frequently diagnosed cancer worldwide, with an increasing incidence rate . Thanks The MWA system consists of a needlelike applicator that works as an antenna at the operating frequency, a power generator, and a low-loss coaxial cable between those devices. The applicator is typically made of a semirigid coaxial cable modified by one or more slots and/or metallic ground in order to achieve monopole or dipole radiation characteristics . CommercTo date, data on the performance and duration for ablation therapies are summarized in tables from the manufacturers of the respective probes and are based on clinical studies of ex vivo and/or in vivo experiments. These data are not standardized and do not take into account patient-specific characteristics that are important for optimal ablation performance . Thus, tThe use of computer-assisted numerical methods is an important component for the development of novel MWA applicators and optimization of existing structures. Experiments, even on ex vivo tissues, are extremely resource- and time-intensive, and for in vivo experiments, there are ethical considerations that complicate the establishment of experimental protocols. For these reasons, the use of numerical simulation methods is particularly suitable for the investigation of the applicators performance. In , a BayesIn this work, an optimization procedure based on a parametric analysis for the design of a dual-mode microwave ablation applicator is proposed. The optimization goals are the minimal reflection coefficient of the antenna at the operating frequency as well as the maximum volume and sphericity of the ablation zone. The optimization is constrained by the microwave sensing mode to enable the detection of a tumor within the host tissue. Previous studies of the dual-mode applicator were designed by optimizing the antenna efficiency only . In thisThe numerical optimization of the dual-mode microwave ablation applicator was conducted with CST Studio Suite (Dassault Systemes). The software comprises the opportunity of full-wave electromagnetic–thermal coupled simulations. Further, biological materials can be implemented in the simulation model. In this section, the simulation model of the open-ended coaxial slot applicator, as described in , with anon zones . The parThe development of accurate simulation models to determine the microwave ablation characteristics and sensitivity are essential for the numerical optimization of the dual-mode ablation applicator. CST Studio Suite offers different modules for a precise three-dimensional simulation of the applicator embedded in so called biological materials. These materials can be defined by their mechanical, dielectric, and thermal properties. The multiphysics electromagnetic–thermal coupled simulation model includes the numerical calculation of dielectric losses in tissue based on the electromagnetic field distribution according to Maxwell’s equations. The spatial distribution of the resulting losses is then transferred into distributed temperature sources for the thermal simulation model. The generated heat is determined by the so-called bioheat equation introduced by Pennes .The structure of the dual-mode applicator is based on a coaxial slot antenna with an open end, as shown in In this study, unidirecional electromagnetic–thermal coupled simulations were performed to obtain information about the antenna efficiency and ablation zone characteristics. For the simulations, the applicator was inserted into a material under test (MUT) block, depicted in As a starting point for parametric analysis of the applicator geometry, an initial design parameter set was determined via an implemented optimization tool in CST. The optimization goal was to maximize the absolute value of the reflection coefficient The boundaries of the simulated region were set to open (add space) for electromagnetic and thermal simulations. Further, symmetry planes of the rotational-symmetric model were set in order to reduce the simulation time. The mesh settings of the hexahedral mesh were adaptively set until the energy within the simulation cell was converged. The resulting global mesh settings with a maximum cell side of The sensitivity of the dual-mode applicator was evaluated by means of the numerical accuracy of the relative permittivity extraction at the operating frequency of ribed in . It is bribed in . The relThe complex coefficients As reference materials, sodium chloride solutions with NaCl concentration of Equation  at the oribed in  and tranThe dielectric contrast between tumor and healthy liver tissue varies per patient ,29. For In this study, the efficiency of the applicator and the ablation zone characteristics in terms of shape and size were optimized under constraints of the applicator’s sensitivity. In the following, the objectives of the optimization are described.The applicator efficiency is related to the computed reflection coefficient It is desirable to achieve preferably large and spherical ablation zones. These properties can be described by the two objective metrics—volume and sphericity of the ablation zone. The heating pattern of each design parameter combination of the applicator inserted into liver tissue was simulated according to loss calculations from the electromagnetic simulations. The input power was set to 10 The optimization of the dual-mode applicator performance comprises multiple object functions x)≥f(x^) .The parametric analysis includes 225 permutations of the applicator geometry. Each electromagnetic simulation required approximately 25 min ± 5 min, and thermal simulations took approximately 1 min ± 0.5 min. All simulations were performed remotely on a CST solver server with 2 × 12 cores @ The Pareto-optimal solution of the parametric analysis can be graphically determined. The values of each objective function The simulated reflection coefficients from 0 GHz to 10 GHz of the Pareto-optimal dual-mode applicator designs inserted into a liver tissue block show a good matching at the operating frequency of w −15 dB . The ParRegarding the results of the corresponding thermal simulations, the axial temperature distribution varies slightly between the parameter sets. With increasing sleeve length, the sphericity of the ablation zone increases and the volume decreases; whereas, the radial temperature distribution practically shows no differences between the three parameter sets. The resulting temperature distributions along the axial (LA) and radial (SA) direction are shown in In this work, a parametric analysis of a dual-mode microwave applicator was conducted with the aim to optimize the applicator’s performance in terms of efficiency as well as the size and shape of the ablation zone. Moreover, the dual-mode functionality was considered in the optimization by constraining the simulation results. In the past, many MWA applicators have been proposed in literature that were designed by matching the antenna to healthy liver tissue properties as surrounding. For a comprehensive optimization of the applicator design parameters, a multiphysical approach must be applied that incorporates electromagnetic–thermal coupled simulations. The starting point of the optimization was selected by an optimization of the applicator efficiency only within liver tissue. The following multiobjective optimization under constraints of the sensitivity resulted in different applicator geometries.This study presents a multiobjective optimization procedure based on numerical simulations of the dual-mode applicator inserted into liver tissue as MUT. The sensitivity investigations are a key factor of the optimization procedure as it is a fundamental property to ensure a proper dual-mode functionality. The sensitivity was defined as the error between the extracted permittivity from the simulated reflection coefficient and the actual reference permittivity. Therefore, electromagnetic simulations of the applicator designs inserted into three reference materials were conducted. By doing so, the applicator performance inserted in materials with different dielectric properties to liver tissue was considered. Regarding applicator-matching of the Pareto-optimal design parameter, the optimal matching frequencies of the antenna were shifted compared with the operating frequency of From the resulting Pareto-optimal design parameter sets, a certain optimal design of the applicator with a distance between tip and slot of This study is limited due to the sole use of numerical solutions to derive the electromagnetic and thermal properties of the microwave applicator; therefore, the calculated extraction errors include numerical uncertainties. The next step will be the experimental verification of the proposed applicator geometries.


To review the role of extracellular vesicles (EVs) released from the male reproductive tract and their impact on developing sperm. We discuss how sperm exiting the seminiferous tubules, although developmentally mature, require further modification. Acquisition of various functions including increased motility, transfer of cargoes and ability to undertake the acrosome reaction is mediated through the interaction between sperm and EVs. A review of the literature identified that EVs are released from different portions of the male reproductive tract, notably the epididymis and prostate. These EVs interact with sperm as they pass from the seminiferous tubules to the epididymis and vas deferens prior to ejaculation.EVs are small lipid-bound particles carrying bespoke RNA, protein and lipid cargoes. These cargoes are loaded based on the state of the parent cell and are used to communicate with recipient cells. In sperm, these cargoes are essential for post-testicular modification.Interactions between developing sperm and EVs are important for the subsequent function of sperm. Prior to ejaculation, these interactions confer important changes for the post-testicular modification and development of sperm. Little is known about the interaction between EVs from the testes and the spermatogonial stem cell niche or developing sperm within the seminiferous tubules. However, the numerous roles of EVs in the post-testicular modification of sperm have led many to suspect that they may also play important roles in developing sperm within the testes.Sperm are crucial for successful fertility. In order to do this, they must be able to swim a large distance to meet the egg in the female reproductive tract and fertilise it. Once released from the testes, sperm may appear to be fully developed, but this is not the case. Several important modifications are required in order for them to swim and fertilise an egg. These modifications are carried out by sending sperm small packages from other cells which contain messages and cargo. We discuss the release of these small packages along with different parts of the male reproductive tract and how they change the way sperm behave and function. This article reviews the literature and known functions of these packages called extracellular vesicles, which are released by the male reproductive tract and modify sperm, transforming their function, before they are ejaculated. Extracellular vesicles (EVs) are a heterogeneous population of small, cell-derived particles, encapsulated by a lipid bilayer. They are secreted by all cell types with wide-ranging functions in both health and disease . SimilarThe study of EVs is a rapidly evolving field. First reported in the 1980s, EVs secreted from the reticulocytes of sheep were initially thought to be a means of waste excretion from cells . Key hormones driving pubertal development include luteinising hormone (LH) which stimulates Leydig cells and follicle-stimulating hormone (FSH) which stimulates Sertoli cells .Within the epididymis, sperm are transformed gaining numerous functions, many of which are mediated by cargo transported to sperm by EVs. These EV-mediated functions include forward motility, zona pellucida binding and the ability to fertilise an oocyte a. FollowClement & Giuliano 2015Epididymosomes like other EV populations show high levels of heterogeneity, and two distinct sub-populations have been reported in bovine models. The first is smaller and CD9-positive and fuses with sperm via their tetraspanin domains, while the larger is CD9-negative and contains higher levels of epididymal sperm-binding protein 1 (ELSPBP1) .b).The epididymis is a tightly coiled tube on the surface of the testis, conveying sperm from the seminiferous tubules to the ductus deferens, where it joins with the excretory ducts of the seminal vesicles. It has numerous roles including storage, maturation, transport of sperm to the ejaculatory duct and removal of defective sperm. This post-testicular maturation ensures the production of a homogeneous population of sperm. Failure to undergo this transformation produces sperm incapable of fertilising an oocyte  and originates from the epididymis   and prostatic acid phosphatase (PAP), but many had not been previously identified within the seminal fluid EVs  but also the corpus (n = 3) and cauda (n = 3) of the epididymis  were found to be absent within the seminal fluid pellet which instead contained heat shock protein-70 and CD55, both of which are known EV markers , the precursor for testicular germ cell tumours, may allow early diagnosis and treatment in the pre-malignant stage.Currently, the release of EVs has not been reported in the testis in humans at different stages of development but has been extensively characterised in the epididymis and prostate, in the adult reproductive tract. Identification of EVs released in the foetal and pre-pubertal testis may give insights into the regulation of germ cell development and testicular pathologies across development, in a similar way to the modification of maturing sperm in the adult testis . DevelopAlthough their release has only been investigated in the transport and storage components of the adult reproductive tract, EVs from these tissues have a clear and important role in the maturation and transformation of sperm. Failure of sperm to undergo this process and acquisition of new functions, mediated by EVs, leaves them incapable of performing their most essential biological function: traversing the female reproductive tract and fertilisation of an oocyte. Further modifications occur in the female reproductive tract including the adaptation of the female immune response to sperm and preparation of the endometrium for embryo impanation, not covered in this review.Increasing knowledge regarding EVs is likely to provide exciting opportunities in the future for the development of EV-based diagnostic and therapeutic options. The potential role of EVs as novel vectors for drug delivery has recently been explored with numerous applications in a range of tissues  Future Leaders Fellowship MR/S017151/1.M P R conceived the idea for the article, undertook the literature search, wrote the manuscript, developed the figures and approved the manuscript for submission. C D G and R T M conceived the idea for the manuscript, undertook editorial changes and approved it for submission.


Significance: The non-destructive characterization of cardiac tissue composition provides essential information for both planning and evaluating the effectiveness of surgical interventions such as ablative procedures. Although several methods of tissue characterization, such as optical coherence tomography and fiber-optic confocal microscopy, show promise, many barriers exist that reduce effectiveness or prevent adoption, such as time delays in analysis, prohibitive costs, and limited scope of application. Developing a rapid, low-cost non-destructive means of characterizing cardiac tissue could improve planning, implementation, and evaluation of cardiac surgical procedures.Aim: To determine whether a new light-scattering spectroscopy (LSS) system that analyzes spectra via neural networks is capable of predicting the nuclear densities (NDs) of ventricular tissues.Approach: We developed an LSS system with a fiber-optics probe and applied it for measurements on cardiac tissues from an ovine model. We quantified the ND in the cardiac tissues using fluorescent labeling, confocal microscopy, and image processing. Spectra acquired from the same cardiac tissues were analyzed with spectral clustering and convolutional neural networks (CNNs) to assess the feasibility of characterizing the ND of tissue via LSS.Results: Spectral clustering revealed distinct groups of spectra correlated to ranges of ND. CNNs classified three groups of spectra with low, medium, or high ND with an accuracy of Conclusions: LSS and machine learning are capable of assessing ND in cardiac tissues. We suggest that the approach is useful for the diagnosis of cardiac diseases associated with changes of ND, such as hypertrophy and fibrosis. These light-tissue interactions are leveraged for tissue characterization by various optical modalities, including fluorescence imaging, multi-photon microscopy, spectroscopy, and tomography. Due to their diagnostic effectiveness combined with miniaturization and low cost, these technologies show promise as an effective means to improve patient care while reducing expense.Optical technologies for medical diagnosis have advanced rapidly over the last several decades. A broad category of these new technologies characterized as “optical biopsies” uses the interaction of light with tissue for its in-vivo pulmonary and cardiac applications. Fiber-optic confocal microscopy (FCM) and optical coherence tomography (OCT) are examples of optical technologies being explored for use in the heart.–,,,,Miniaturization of optical components such as lenses and the development of coherent optical fiber-optic bundles have enabled the integration of optical technologies into catheters for use in a variety of previously unattainable regions, such as ,,–Spectroscopic approaches using light in the visible and near-infrared spectrums are promising alternatives that could decrease overall cost and complexity while increasing the depth of tissue characterization by leveraging the physics of light transport and scattering in tissue.Many cardiac diseases such as myocarditis, amyloidosis, and other cardiomyopathies as well as allograft rejection are diagnosed by observing and quantifying microstructural abnormalities. The current gold standard for diagnosis of these diseases is retrieving a biopsy of cardiac tissue, a procedure associated with significant risk. LSS as a means of characterizing abnormal microstructural changes in cardiac tissue has the potential to decrease the risk and harm associated with diagnosing and monitoring these diseases by gathering information from the tissues in a non-destructive manner. An important foundation that needs to be established for the development of LSS toward application in these cardiac diseases is the sensitivity of the LSS spectra to changes in the scattering profile of tissue and its correlation with changes in the underlying tissue, such as the density of myocyte nuclei.–in situ.,,We investigated the ability of LSS to identify the nuclear densities (NDs) of the myocardium in combination with machine learning. Machine learning is increasingly utilized to overcome challenges related to analyzing complex signals such as spectra.,Here, we introduced a novel system for broad-spectrum LSS for applications in cardiology and cardiac surgery. This system measured spectra from cardiac tissues of an ovine model spanning a large range of gestational ages, including pre-term and adult. Previous work on cardiac tissues from ovine models showed a decrease in ND with age.22.1A customized LSS setup was developed for application in cardiac tissue. A custom-built spectroscopy probe  was used to both illuminate the tissue with broad-spectrum light and gather light from the tissue. The setup shown in The probe utilized a single 2.2The LSS system was used to gather spectra from ovine ventricular tissue according to the process outlined in 2.3After spectral measurements, tissue samples were sectioned within the probed region at the center of each sample. A vibratome  was used to section at A Leica SP8 confocal microscope with a Two or more different tile scans from each sample were used to calculate the ND. Manual segmentation was performed using the Fiji/ImageJ image processing software package.2.4Unsupervised and supervised machine learning approaches were used to explore whether spectra gathered from the LSS system allowed for differentiation between the ND of the preparations. We employed MATLAB  and its Deep Learning Toolbox for unsupervised learning evaluation of the correlation between LSS spectra and the measured ND.spectralcluster to identify clusters of spectra from preparations of different NDs using the Euclidean distance metric. From this spectral clustering analysis, we generated a similarity graph between nodes of the first and second principal components of the spectra.gscatter to visualize the spread of the spectral set, with the resulting groups identified by color. Groups were associated with the NDs of the spectra contained in each group. The principal components analysis (PCA) was performed, and the first two principal components were used to visualize any potential groups within the dataset. A one-way ANOVA and Tukey-Kramer post hoc test using a significance level of 0.05 was used to identify differences between the clustered spectra. A box and whisker plot was created from these data, which compared the NDs associated with the spectrum within each group produced from the spectral clustering. Significant differences between clusters were indicated by brackets.We explored normalized, filtered, and concatenated spectra from the paired collection fibers. Each spectrum was annotated with the ND of the sample from which it was measured. We applied the MATLAB function 2.5We applied CNNs to differentiate NDs based on spectra from LSS. This work was built upon our previous work on identifying fibrosis from in vivo confocal imaging.The CNNs were systematically trained and validated using a form of k-fold cross-validation called leave-one-out cross-validation (LOOCV).Two tests were performed to determine the potential sensitivity of the networks to specific wavelength ranges and to also determine the sensitivity of the networks to the resolution of the spectra. An initial network configuration was selected . HyperpaAlthough hyperparameter variation was not performed during these training sessions, a validation sample was still removed from the training dataset in addition to the testing sample. Network weights were saved during the training process when the loss value decreased. The network weights resulting in the lowest loss value were restored at the end of the training session. The predicted classes from the testing datasets for each fold were then compared to the assigned classes for each spectrum based on ND values. From this comparison of aggregated predictions, we calculated the accuracy for the CNN topology on the entire LOOCV as follows: To improve the accuracy of the predictions resulting from the LOOCV, the above training process was altered to introduce hyperparameter variations within each training fold. Multiple networks were trained using the base topology outlined in 33.1We gathered 440 spectra from two collection fibers from 22 samples. Raw spectra displayed a moderate degree of variability in intensity within samples and in-between samples. Spectra from R1 and R2 were different with respect to intensity range and shape . After n3.2Representative transmural confocal microscopy tile scans of RV free wall myocardium are shown in Manual segmentation using ImageJ indicated a 72.5% difference in ND between samples of 4.3 months and 10 months gestational age and a 78.2% difference in ND from 4.3 to 56 months. Automated segmentation yielded a 65.4% and 77.8% decrease in ND for the same age differences. The reduction of ND with age in our samples is summarized in 3.3Dimensionality reduction via PCA revealed that the first two principal components accounted for clusters . Two of clusters . For ins3.4We explored the sensitivity of the CNN to different wavelength ranges. Accuracies of 4,,The presented studies suggest that the developed LSS system can predict NDs in cardiac tissue samples within predefined ND ranges based on CNNs. The studies were based on microscopy and image processing, which showed decreases in ND on the order of 70% between hearts of 4.3 to at least ten months gestational age. This decrease is consistent with previously observed values.,LSS is particularly well suited to identify the change in the relative distribution of major cellular components such as nuclei, which have long been recognized as major sources of light scattering.,,LSS relies on the wavelength-specific absorption and scattering of photons by cellular and subcellular structures as well as constituents of the extracellular matrix.,It is well understood from animal models that the ND of heart muscle decreases in maturing neonates as the cardiac myocytes grow.,,,Analysis of LSS spectra is often a time-consuming endeavor based on simplified models and mathematical simulations of light-tissue interactions.,,Our second approach for the prediction of NDs from spectra was based on CNNs and histological quantification of NDs. The high accuracy of our predictions using LOOCV demonstrated that the CNN can reliably classify spectra into predefined ND groups. CNNs perform feature extraction and dimensionality reduction intrinsically, thus reducing or removing the need for preprocessing frequently required when working with spectral data. For these reasons, CNNs are well suited to handle raw signals.Monitoring of cell density, the fraction of extracellular components such as collagens, and the degree of cellular infiltration is essential to diagnosis and treatment planning for certain types of heart disease. Our studies indicate that LSS can provide a rapid means of tissue characterization in the heart. There is no destructive process required for this tissue characterization. Due to the simplicity of the developed LSS probe, it could be single-use disposable. This constitutes an advantage over many existing modalities such as FCM or OCT, where probes are expensive each due to their technical complexity and require sterilization protocols.,Potential applications of the developed LSS system include the diagnosis of heart diseases. Decreases in myocyte density and increases in cellular infiltrates are a hallmark of hypertrophy and myocarditis, respectively.The results of our experiments with a reduced resolution spectral dataset indicate that a spectrometer with nanometer wavelength resolution is not necessary to achieve a high level of accuracy for ND prediction via CNN. The LSS system could be based on spectrometers with lower wavelength resolution. This finding can inform the cost-effective design of future LSS systems.A limitation of our work is that we ignored intensity differences and normalized the spectra to the mean intensity before analyses. Our motivation was to reduce the effects of attenuation of spectra due to bending of the optical fibers and suboptimal connection of fibers to the spectrometer. Also, we applied a simple Gaussian filter to remove noise. The filter parameters were determined by visual inspection of a Fourier analysis of a representative sample of the dataset. The standard deviation of 20 spectral measures was selected due to it removing the majority of the noise from the signal while still preserving the more subtle fluctuations in the spectra. More advanced filters might improve the reduction of noise in the spectra. We note also limitations regarding the specific approach to machine learning used in this study. We only investigated CNNs for classification. Other methods of machine learning, such as fully connected neural networks and k-means clustering, could yield similar or higher levels of accuracy and performance. We also acknowledge the limitations associated with the size of the dataset used for this study. A larger dataset could improve the performance of the networks trained in this study, potentially resulting in a successful regression estimation of NDs from LSS data. Due to the size of the datasets used, our networks could overfit the training dataset during the training process. We used LOOCV, where the hyperparameter variation was performed on data that were separate from the training and testing datasets, to prevent biasing the network to the testing dataset. We also kept the number of trainable parameters small in our networks, 5Two machine learning approaches, spectral clustering and CNNs, allowed us to identify, with high accuracy, a range of NDs in cardiac tissues using spectra gathered by a customized LSS system. The described system is of low technical complexity and cost. The combination of LSS and machine learning constitutes a useful and affordable tool for the diagnosis of cardiac tissues for diseases hallmarked by microarchitectural changes. The presented approach has applications in monitoring and diagnosing heart diseases such as hypertrophy, fibrosis, and myocarditis. A further application is transplant rejection surveillance in lieu of traditional endomyocardial biopsy.


Contamination monitoring, scintigraphic imaging, and quantification of effective radiation dose provide the level of risk to the operator from the subcutaneous radioactive contamination. Findings demonstrated a very low dose to operator and no deterministic radiobiological effects. Delayed imaging demonstrated negligible biological clearance from the injury site. Implications of the findings for clinical practice are discussed, highlighting the need for a careful and calm approach to radiopharmacy activities.Attention to the implications of common needle stick injuries has focused heavily on the risk of cross-infection from blood-borne pathogens. An additional risk to the nuclear medicine healthcare worker is that of subcutaneous radioactive contamination from radiopharmaceuticals. This case report provides a rare opportunity to examine the clinical and operator causes of one such event during the dispensing of  Needlestick injuries (NSI) present healthcare workers with the risk of cross-infection from blood-borne pathogens such as hepatitis B virus, hepatitis C virus, and human immunodeficiency virus Previous studies have acknowledged the occurrence and risks of radiopharmaceutical NSI, highlighting the utility of re-capping devices to reduce such events ,6. Surpr99mTc-Tetrofosmin from a multi-dose vial, the operator (JE) incurred a needlestick injury to the thenar aspect of the left palm . Statistical analysis of the scintigraphic imaging .Full informed consent has been given for the publication of this case report.


The combination of planar chromatography with desorption/ionization mass-spectrometry (MS) techniques provides chemists with unique tools for fast and simple separation of mixtures followed by the detection of analytes by the most powerful analytical method. Since its introduction in the early 1990s, thin-layer chromatography (TLC)/matrix-assisted mass spectrometry (MALDI) has been used for the analysis of a wide range of analytes, including natural and synthetic organic compounds. Nowadays, new desorption/ionization approaches have been developed and applied in conjunction with planar chromatography competing with MALDI. This review covers recent developments in the combination of TLC with various desorption/ionization MS methods which were made in recent several years. The conjugation of thin-layer chromatography (TLC) and mass spectrometry (MS) is one of the promising hybrid analytical methods. Despite the seeming simplicity, TLC with different detection systems is actively used in analytical and organic chemistry for the express separation of various mixtures and preliminary quantification, for example, for the analysis of oil and petroleum products , monitorChlorella vulgaris and Spirulina platensis using high-performance TLC (HPTLC) and many others; the qualitative determination of the target components in the composition of complex mixtures can be reliably performed using suitable detection systems, and one of the main analytical methods in this case is conjugation to MS. At the same time, MALDI, among all the MS methods, occurred to be one of the most suitable methods for direct conjugation to planar chromatographic methods, opening broad possibilities for the identification of components of complex “omics” mixtures, screening, especially for small molecules, and development of methodological approaches, including new stationary phases for TLC, which might be suitable for direct MS detection of the separated compounds. MERCK KGaA, for instance, produces the so-called MS grade TLC plates which possess some important advantages; these TLC plates are characterized by a very low MS background due to the control of MS-detectable impurities, and the stationary phase layer thickness has been decreased to 100 μm, which is twice lower than that of standard TLC plates, as it was found that layer thickness influences the efficiency of laser desorption from the plate surface . Together with the minimal thickness of the stationary phase layer (1–50 μm) providing improved separation properties, these TLC plates were found to be suitable to the direct coupling to different MS detection systems due to covalent binding of the stationary phase and, as a result, the low bleeding level. However, together with the advantages, a decrease in the thickness of the sorption layer also resulted in the decrease in the resolution due to the small elution pathway. The solution of the problem was in increasing of the separation efficiency, so the UTLC and HPTLC plates were taken into consideration. Increase of the efficiency allowed the increase of the sensitivity of the method, reaching about a dozen pmol per sample using UV detection and a few pmol with MALDI-MS.via coating nonporous silica particles, chemically modified with polyacrylamide, as 15 mkm films, on glass or silicon. The authors used the model mixture of three proteins, namely, myoglobin, cytochrome c, and lysozyme, for testing the separation ability of the prepared sorbent, and have received the nearly baseline resolution realized by the hydrophilic interaction chromatography mechanism. The sorbent demonstrated the HETP value of about 3 mkm, giving the opportunity to develop the separation efficiency more than 3500 TP per plate. Zhang et al. also varied the silica particle diameter, and tested the 900-, 700-, and 350-nm particles. The typical SEM image of a UTLC slide with a coating with silica nanoparticles demonstrates the smooth particles with narrow size distribution and optimal layer arrangement.As it was found, the decreasing particle diameter improved the resolution but slowed down the separation. The plates with the optimized properties were used combined with MALDI-MS for the analysis of the proteins. MALDI for each protein was carried out at the region which was previously marked by the fluorescently labeled proteins under a microscope, and the mass spectrum for the center of each spot was recorded. The mass spectrum in each case agreed with the expected molar mass for each protein. The signal-to-noise ratio had an acceptable value, confirming the possibility of utilization of the approach.In 2013, Svec et al.  suggesteIn 2015, However, the main difficulties for this combination are the application of the MALDI-MS matrix onto the TLC plate and subsequent ionization. Kucherenko et al. suggested TLC plates with an organic polymer monolithic layer to work out the problem ; 2019. TOne of the important distinctions of MALDI-MS from other soft ionization methods is the key role of the matrix in the ionization process. There are a large number of matrix compounds suitable for the analysis of low molecular weight analytes . HoweverAnother example of the application of the inorganic matrix was proposed in the study by It is important to note, however, that the use of solutions of matrixes can cause secondary chromatographic processes on TLC plates and, hence, analyte spot blurring. This problem can be overcome by using solvent-free matrixes such as graphite-assisted laser desorption/ionization (GALDI) . The samComposite glycerol/graphite/aromatic acid matrices were also proposed to avoid spot blurring and enhance desorption/ionization of the analytes . MixtureDopants also sometimes play a very important role in ionization processes . In casePlanar chromatography is one of the first analytical methods where derivatization procedures were applied mainly for spot visualization . DerivatA similar approach was used for TLC/MALDI analysis of primary amines . In thisThe unique opportunities of the TLC/MALDI systems result in wide utilization of this method in the analytics of complex organic mixtures. One of the areas of interest in current analytical chemistry is lipid analysis. Leishmania major, was chosen by the authors as a model enzyme to screen biological activity in plant extracts due to its capacity to hydrolyze specific carbohydrates. The first part of the approach was in differential analysis by TLC densitometry to determine the zones in the plant extract between the blank and enzymatic reaction. Zones of interest were then subsequently investigated via TLC/MALDI-TOF-MS for the identification of bioactive molecules. The development of the method demanded the solution of different analytical problems, such as separation of isomers (glucose and fructose), elimination of the high-matrix effect, and the analysis of polar molecules with low molar masses (sugars), which is a challenging problem itself while interacting about MALDI. However, the method occurred to be feasible for the analysis of bioactive molecules in complex mixtures containing interfering compounds .Escherichia coli. The direct TLC-IR-MALDI-MS analysis allowed the successful visualization of the chromatographic separation of the various lipo forms of globotriaosylceramide and globotetraosylceramide. The developed method allowed the fast and reliable overview of the glycosphingolipid composition of the investigated cell line of high medical relevance. This possibility may also be useful in glycolipidomic studies of complex biological matrices; for example, for globotriaosylceramide imaging, TLC/MALDI is widely used for the analysis of lipids . Thus, KThough MALDI-MS is the most used approach for direct detection of analytes from TLC plates, there also several desorption/ionization methods which are also capable of such analyses. Thus, even in the 1980s, it was shown that fast atom bombardment  and secof. Nevertheless, the approach was used, for example, to develop methods for detecting components of thermochromic inks for forensic purposes (The first study demonstrating the power of the combination of TLC and ambient ionization MS was published shortly after the presentation of DESI . The metpurposes . A very purposes . In thisDART mass spectrometry operates using Penning ionization principles: excited atoms of gas (mainly helium) interact with molecules of ambient air components producing secondary ions, which desorb/ionize compounds from the surface of the analyzed object . This ioThe first version of the commercial DART ion source was inconvenient for TLC analysis because the gas stream was directed to the orifice of the MS interface and the analysis required cutting of the plates. Though a special adapter increases the efficiency of desorption/ionization and reproducibility of the results was proposed by Plectranthus amboinicus (Lour.) essential oil. Comparison of HPTLC-UV/Vis chromatograms before and after derivatization using the anisaldehyde–sulfuric acid reagent and HPTLC-Vis-EDA autograms allowed determining bioactive compound zones, which then were subjected to analysis by DART with a high-resolution mass analyzer. A similar approach was used for bioanalytical profiling of sunflower leaves (All abovementioned improvements have rapidly increased the popularity of TLC/DART, and now, it is the most common combination of ambient mass spectrometry with planar chromatography. Thus, this approach is often used for the characterization of plant extracts. For example, DART-MS coupled with the ion trap mass analyzer was used for express identification of alkaloids extracted from plants and separated by TLC . The comr leaves .TLC/DART analysis can also be used for quantitative analysis of various compounds. A very promising approach was developed by There are also a significant number of other home-built systems allowing desorption/ionization of analytes from TLC plates. Their descriptions are summarized in Reproducibility and high spatial resolution achieved by MALDI-MS are still keeping this method as the most widely used desorption/ionization technique for the detection of analytes from TLC plates. Further developments of this method, including new chromatographic sorbents and matrix compounds/mixtures, and novel sample preparation procedures guarantee its popularity for the analysis of various objects. Most notably, there are new studies introducing TLC/MALDI as a useful tool in new research fields such as, for example, petroleomics. However, the fast growing number of ambient ionization techniques offers a good alternative for MALDI. These methods allow developing cost-effective, robust, and express approaches for the detection of various classes of analytes from TLC plates. We believe that all these MS ionization approaches will keep one of the first chromatographic separation methods actual for a lot of purposes.