Ts (300–773 K) on the structural phase ordering, static and dynamic magnetization behaviour has been investigated in ion beam sputtered full Heusler alloy Co2FeAl (CFA) thin films on industrially important Si(100) substrate. The B2 type magnetic ordering is established in these films based on the clear observation of the (200) diffraction peak. These ion beam sputtered CFA films possess very small surface roughness of the order of subatomic dimensions (<3 Å) as determined from the fitting of XRR spectra and also by AFM imaging. This is supported by the occurrence of distinct Kiessig fringes spanning over the whole scanning range (~4°) in the x-ray reflectivity (XRR) spectra. The Gilbert damping constant α and effective magnetization 4πMeff are found to vary from 0.0053 ± 0.0002 to 0.0015 ± 0.0001 and 13.45 ± 00.03 kG to 14.03 ± 0.04 kG, respectively. These Co2FeAl films possess saturation magnetization ranging from 4.82 ± 0.09 to 5.22 ± 0.10 μB/f.u. consistent with the bulk L21-type ordering. A record low α-value of 0.0015 is obtained for Co2FeAl films deposited on Si substrate at Ts ~ 573 K.The influence of growth temperature  Tc),making them attractive for spintronic applications; high density magnetic storage1α), which describes the rate of relaxation of the magnetization. However, as the ferromagnetic (FM) film thickness is reduced, it becomes difficult to maintain a low value of the damping constantet al. reported ultra-low damping in Co-Fe binary alloy thin filmsTc (~1000 K) and 100% spin polarization due to the spin split band structure induced half-metallicityTc as well as the spin polarization2YZ, e.g., Ni2MnSi, Co2MnSi, Co2MnGe, Co2FeAl, etc., exhibit three different types of atomically ordered phases, namely fully ordered L21, partially ordered B2 and fully disordered A2 phases. In the L21 phase, all the three types of atoms  occupy their respective assigned sites. However, in the B2 phase, Y and Z atoms randomly share their positions, and the A2 phase corresponds to a completely random occupation of all existing sites of the L21 phase by any X, Y or Z atoms. The half-metallicity in Heusler alloys is sensitive to these atomic arrangements and degrades with atomic disorder, compared to the L21 phase which is perfectly half-metallic. Picozzi et al. reported that the atomic disorder might lead to additional states at the Fermi level, thus reducing the spin polarization92FeAl (CFA) is the most widely investigated Heusler alloy. Although several attempts have been made to grow fully ordered CFA thin films, especially on MgO substrate, using different growth techniques, the success in crystallizing the L21 phase in Co2FeAl Heusler alloy thin film is far from being achieved. To our knowledge, only Qiao et al., reported L21 films by MBEThe interest in the Heusler alloy thin films is widely increasing due to properties like tunable anisotropy, large spin polarization and high Curie temperature (filmsTc ~00 K and filmsTc ~00 K and Ts) and to use sufficiently high Ts so that on the one hand it promotes the formation of the L21 phase while on the other hand it avoids atomic disorder which is a thermally activated process. Therefore, synthesis of highly ordered CFA thin films continues to be a challenging task. Very few attempts have made to grow CFA thin films on Si substrate1213B2 ordered CFA filmsTs the surface mobility of the ad-atom during the growth of the films is sufficiently high so that they can lower their energy by reaching to their respective atomic sites. In this regard, the ion beam sputtering technique−5 to 10−4 Torr range). This makes this technique an energy enhanced process in which sputtered species carry substantially large energy, ~20 eV, compared to other deposition techniques. This high energy of sputtered species is responsible for high ad-atom mobility of the growing films and reduction of the phase formation temperature. In this report, we present a detailed study on the structural and magnetic (static and dynamic) properties of CFA thin films grown on Si(100) substrate at different Ts by employing ion beam sputtering. The observed results demonstrate a direct correlation between the structure and dynamic response of the CFA thin films, tuned by optimizing the substrate temperature during the ion beam sputtering.In order to realize the fully ordered CFA phase, it is necessary to optimize the growth temperature  chamber for deposition using a load-lock system. The substrates were then heat treated at 620 °C for 2 h for smoothening to achieve atomically flat substrate surfaces prior to film growth−7 Torr using a turbo pump. The 6″ dia. CFA alloy target (composition Co50Fe25Al25) fixed on a remote controlled water cooled turret was sputtered by ~4.5 inch dia. high energy Ar-ion beam (500 eV) extracted from a RF ion-source . During the deposition, the chamber was maintained at ~2.4 × 10−4 Torr by bleeding 3.5 sccm Ar gas directly in to the ion-source operated at 75 W. A series of Si/CFA(53 nm)/Ta(2 nm) thin films were prepared by varying Ts while keeping the other parameters constant . A 2 nm thick layer of Ta was used as a capping layer to prevent the surface oxidation. It is to be noted that the above mentioned thicknesses are only the nominal thicknesses derived from measurements on thicker films using surface profilometry employing a Bruker Dektak XT profiler system . The actual film thicknesses of the FM layer, Ta and its oxide (Ta2O5) were accurately determined by simulating the experimentally observed X-ray reflectivity (XRR) spectra (The CFA thin films of constant thickness were deposited on Si(100) at various temperatures using a ion-beam sputtering deposition system (NORDIKO-3450). Prior to the deposition, the Si(100) wafers were ultrasonically cleaned in acetone and propanol baths, and subsequently treated with HF (10:1 ratio) solution for 60s to remove the native SiO spectra .Scan Assist). The film thickness and interface roughness were investigated by simulating the specular XRR spectra using the PANalytical X’Pert Reflectivity software (ver. 1.2 with segmented fit). To determine the chemical state  of the top Ta as well as of the underlying FM layer, the binding energy shift employing X-ray Photoelectron Spectroscopic (XPS) measurements was carefully investigated. The XPS spectra on Si(100)/CFA/Ta bilayers were recorded by using a SPECS make system with an Al-Kα x-ray source (1486.6 eV) and a hemispherical energy analyzer with pass energy of 40 eV and a resolution of ~0.7 eV. The saturation magnetization was measured using the vibrating sample magnetometer (VSM) module of the Quantum Design make Physical Property Measurement System (QD PPMS-VSM) Evercool-II. The magnetic anisotropy behaviour was investigated by recording hysteresis loops at various azimuthal orientations by employing a homebuilt Magneto-Optical Kerr Effect (MOKE) set up in longitudinal geometry. A broadband lock-in amplifier (SR830 from Stanford Research Systems) detection based custom built ferromagnetic resonance (FMR) technique employing a vector network analyzer (Model 8719ES from HP) and a coplanar wave guide (see ref. The deposited CFA films were investigated by glancing angle X-ray diffraction (GAXRD). To ascertain the film morphology, topographical scanning was performed using the Bruker make atomic force microscope (AFM)  and (311) are expected to be present in the diffraction patternB2 phase. The presence of the (200) peak in our films , (400) and (422) diffraction peaks indicative of the polycrystalline nature of the CFA thin films. Ts. The observed intensity of the (220) diffraction peak exhibits an increase at Ts = 673K owing to the improved crystalline quality. Moreover, in accordance with the enhancement in peak intensity, a corresponding minimum in FWHM is also observed at Ts = 673 K, indicating a transition from short-range to long-range crystallographic orderTs is the result of strain relaxation at higher Ts. The lattice constant (a) evaluated using the (220) peak is found to decrease with Ts. The values of a are in good agreement with values reported for the bulk L21 phase (0.574 nm)Ts. It is inferred from the graph that the crystallite size increases with increasing growth temperature. It is easily understandable considering the diffusion process from an atomic perspective. With increasing Ts, the ad-atoms gain sufficient surface mobility which helps them to adhere to the nucleated islands thereby increasing the crystallite sizeL21 and B2 CFA phases by XRD is difficult, and requires other probes such as neutron diffraction, NMR, etc. However, there exists an indirect approach to distinguish among the L21, B2 and A2 phases of CFA. This essentially relies upon the density of states (DOS) based correspondence between the smallest α and the extent of structural ordering. The lowest damping corresponds to the perfectly ordered half-metallic L21 phase, and highest damping corresponds to the disordered phasending to 1 and  assuming a tri-layer model CFA/Ta/Ta2O5. The simulated XRR spectra can be seen to overlap remarkably well with the experimental data  of individual layers in the Si(100)/CFA/Ta samples, specular XRR spectra were recorded. tal data . The estTS increased to 673 K. The density decreases down to 6.34 ± 0.06 g/cc on further increase of the growth temperature to 773 K. This Ts dependent variation in film-density can be attributed to the Ts induced changes in the ad-atom mobility as explained above. For comparison, the surface roughness measured using AFM and XRR is shown in Ts. A thin layer of 2.2 nm of Ta2O5 was found to be formed on the top of the Ta layer due to the surface oxidation. As XPS is a powerful spectroscopic technique to examine the electronic properties such as electronic environment, oxidation state, multiplicity etc. of the various possible species formed near the surface of the films, in the next paragraph we present and analyse the XPS data recorded on the CFA/Ta bilayer sample deposited at Ts ~ 573 K.The density of the RT grown CFA layer is found to be 6.12 ± 0.06 g/cc, which increased up to 6.51 ± 0.06 g/cc as Ts ~ 573 K. The portion of recorded spectra corresponding to the binding energies of Ta, Co and Fe were de-convoluted in to their component metal and oxide-peaks and fitted by using the XPSPEAK 4.1 software which automatically takes care of the background during fitting using background correction tool. All spectra were calibrated using the C-1s peak (284.6 eV). The XPS investigation revealed the presence of Co, Fe, Ta and Ta2O5 in the Si(100)/CFA/Ta thin film. Whereas the select regions (in orange and blue) in viz., 4d5/2 and 4d3/2, respectively corresponding to the presence of Ta2O5, the binding energy regions plotted in green and violet evidence the existence of the Ta-metal layer. Similarly in 7/2 and 4f5/2 electronic states, respectively corresponding to the formation of Ta2O5. Likewise, the corresponding Ta-metal peaks are represented for clarity in green and violet colors. The relatively lower intensity in the Ta-metal peaks compared to that of Ta2O5 . Thus, the XPS data provide further experimental support to the formation of a thin passivating layer of Ta2O5 over the Ta and underlying CFA as revealed from the XRR simulations. Zhu et al.et al.2O5 layer for protecting the underlying Ta and FM layers.of Ta2O5  is consiof Ta2O5 . Figure Ts are shown in Mr/Ms = 1) indicating nearly defect free samples and domain wall nucleation controlled switching of the magnetization. Ts = 573 K, which endorses the existence of uniaxial anisotropy in these CFA thin films. The origin of the uniaxial anisotropy could comprise of several sources, such as strain due to film-substrate lattice mismatch or the intrinsic magnetocrystalline anisotropy effect. It is noted that a characteristic spike is present near the positive coercive field in these hysteresis loops  and coercivity (Hc) evaluated from MOKE as function of Ts are shown in Ms values are very much comparable to the reported bulk value (5.0 μB/f.u.)Ms indicates the improvement of local magnetic ordering of these thin films with the increase in TsTs, which also improves the local magnetic ordering within the lattice of the Heusler alloy. Hc decreases as the growth temperature Ts is increased. It may be pointed out that the Hc is influenced both by the pinning of the domain walls at the pinning centres/defects  as well as by the local variations in the magnetic anisotropy, viz., on the angle between the easy-axes of the domains with respect to direction of the applied magnetic field. Observation of higher (lower) Hc in room (high) temperature deposited films is attributed to the presence of relatively higher (lower) structural and magnetic disorder and smaller (larger) grain size of those films. This finds support from the observed increase in Ms  = ΔH2/{ΔH2 + (Hext−Hr)2} and FA(Hext) = ΔH(Hext − Hr)/{ΔH2 + (Hext − Hr)2} are symmetric and antisymmetric Lorentzian functions, respectively, with S and A being the corresponding coefficients. The experimental and fitted FMR spectra of the film deposited at 573 K are shown in Hr on the microwave frequency.Here, Hr and the frequency f can be expressed by the famous Kittel’s formula29The relationship between the resonance field γ = gμB/g being the Lande’s splitting factor and taken as 2, 4πMeff is the effective magnetization, and HK is the in-plane uniaxial anisotropy field  is expressed asThe frequency dependence of the linewidth ΔH0 is the frequency independent film inhomogeneity linewidth contribution, while the second term corresponds to the frequency dependent contribution associated with the intrinsic Gilbert relaxation.It is important to point out here that the first term ΔH vs. f plots together with the fits obtained using α for the films deposited at different Ts. The extracted values of the damping constant α for all the samples are plotted in α is found to be 0.0015 ± 0.0001 for the film deposited at 573 K. This value competes excellently with the smallest α-values of 0.001 and 0.0011 reported by Mizukami et al.et al.31α obtained in these CFA films is correlated with improved structural quality (evident from the observation of the (220) reflection, B2 ordered phase (evident from the presence of the (200) reflection, 4πMs  and film inhomogeneities33on, 4πMs , low pinon, 4πMs , and exton, 4πMs . The inhHinh; inhomogeneity of the sample which is independent of frequency, (ii) ΔHG; Gilbert damping term which is intrinsic in nature and dependent on frequency, and (iii) ΔHmag2; TMS term which is extrinsic in nature and frequency dependent. Thus the overall linewidth of the system can be written asTo rule out the possibility of a TMS contribution in our film samples, we have performed a detailed analysis including the TMS contribution as described in the theory of two-magnon scattering by Arias and Millswhere H vs. Hr plots were fitted using K, and the parameters b (height/depth of bump/pit defects), p (fraction of surface covered by defects), D (exchange stiffness), and Hs (surface anisotropy field << 4πeffM) are assumed to be constants for the material under study  by employing ion-beam sputtering deposition technique is reported. Based on the detailed investigations of the microstructure, surface morphology, X-ray reflectivity, and the static and dynamic magnetization properties, it is found that these CFA thin films exhibit the B2 ordered phase and possess improved crystallinity, very low surface roughness (<3 Å), and minimal anti-site disorder consistent with the observed higher saturation magnetization. Most importantly, a record low Gilbert damping constant of 0.0015 ± 0.0001 is obtained corresponding to the optimum substrate temperature of ~573 K. This lowest value of the damping constant observed in the CFA films deposited on silicon substrate compares excellently well with values reported for MBE grown CFA films deposited on single crystalline MgO substrates. The study clearly suggests that thermally stable half-metallic ferromagnetic Co2FeAl thin films sputtered on the industrially popular Si substrate have remarkable application potential owing to their very low magnetic damping for developing novel spin transfer torque devices.In summary, the growth of CoHow to cite this article: Husain, S. et al. Growth of Co2FeAl Heusler alloy thin films on Si(100) having very small Gilbert damping by Ion beam sputtering. Sci. Rep. 6, 28692; doi: 10.1038/srep28692 (2016).


Essential demographic and clinical data were collected to calculate the RESP score, the ECMOnet score, and the sequential organ failure assessment (SOFA) score before VV-ECMO. The predictive power of hospital mortality of each score was presented as the area under receiver-operating characteristic curve (AUROC). The multivariate logistic regression was used to develop an institutional prediction model. The surviving to discharge rate was 55% (n = 59). All of the 3 published scores had a real but poor predictive power of hospital mortality in this study. The AUROCs of RESP score, ECMOnet score, and SOFA score were 0.662 (P = 0.004), 0.616 (P = 0.04), and 0.667 (P = 0.003), respectively. An institutional prediction model was established from these score parameters and presented as follows: hospital mortality (Y) = −3.173 + 0.208 × (pre-ECMO SOFA score) + 0.148 ×  + 1.021 × (immunocompromised status). Compared with the 3 scores, the institutional model had a significantly higher AUROC . The 3 published scores provide valuable information about the poor prognostic factors for adult respiratory ECMO. Among the score parameters, duration of mechanical ventilation, immunocompromised status, and severity of organ dysfunction may be the most important prognostic factors of VV-ECMO used for adult respiratory failure.Despite a potentially effective therapy for adult respiratory failure, a general agreement on venovenous extracorporeal membrane oxygenation (VV-ECMO) has not been reached among institutions due to its invasiveness and high resource usage. To establish consensus on the timing of intervention, large ECMO organizations have published the respiratory extracorporeal membrane oxygenation survival prediction (RESP) score and the ECMOnet score, which allow users to predict hospital mortality for candidates with their pre-ECMO presentations. This study was aimed to test the predictive powers of these published scores in a medium-sized cohort enrolling adults treated with VV-ECMO for acute respiratory failure, and develop an institutional prediction model under the framework of the 3 scores if a superior predictive power could be achieved. This retrospective study included 107 adults who received VV-ECMO for severe acute respiratory failure (a P The venovenous (VV) mode is the preferred mode of ECMO in this setting, and yields a significantly higher survival rate when compared with venoarterial (VA) ECMO . However, ECMO has not gained full acceptance among medical institutions worldwide due to its invasiveness and high resource usage.,5 The fact that VV-ECMO itself fails to bring survival benefits in the conventional ventilatory support versus extracorporeal membrane oxygenation (CESAR) trial increases the controversy of VV-ECMO for ARF in adult patients. Furthermore, a delayed administration of VV-ECMO  is a known risk factor that may turn this salvage therapy into futile medical care.–9 Therefore, to establish consensus on the timing of intervention, the ELSO and other European ECMO organizations have retrospectively analyzed their databases of adult respiratory failure and launched some outcome-predicting scores to enhance the decision-making process; 2 of them are the respiratory extracorporeal membrane oxygenation survival prediction (RESP) score and the ECMOnet score. Except the 2 published scores, the sequential organ failure assessment (SOFA) score is also a common tool used to predict mortalities in adult ECMO.–14 Since the therapeutic experience and equipment may significantly differ among institutions, physicians are encouraged to validate the predictive power of these “packaged” scores with their own databases before using them to recruit candidates.,16 The primary aim of this study was to validate the predictive powers on hospital mortality of the 3 published models  with our patient cohort of adult respiratory ECMO, and the secondary aim was to develop an institutional prediction model under the framework of these packaged scores to see if a superior predictive power could be achieved.Extracorporeal membrane oxygenation (ECMO) is an effective respiratory support to correct hypoxemia in adult patients with severe acute respiratory failure (ARF), which is not improved by advanced mechanical ventilation (MV).22.1 The ethics committee of the Chang Gung Medical Foundation approved the protocol (CGMF IRB no. 104–6277B) and waived the necessity of individual patient consent.From March 2007 to April 2015, a total of 123 adult patients received VV-ECMO for advanced respiratory support at Chang Gung Memorial Hospital. This university-affiliated hospital is a tertiary referral center with 3400 beds. To reduce the heterogeneities in disease severity and patient management, 16 patients were excluded due to a requested transfer to another hospital (n = 1), switching to VA or mixed configuration of ECMO (n = 5), and death on VV-ECMO in the first 24 hours . Therefore, only 107 among the 123 patients who received a single run of VV-ECMO and survived on VV-ECMO for >24 hours were enrolled in this retrospective study. This study was conducted in accordance with the amended Declaration of Helsinki.2.2aO2/FiO2 ratio <70 mm Hg) under advanced MV (FiO2 >0.8 and peak inspiratory pressure [PIP] >35 cm H2O). Nevertheless, VV-ECMO was contraindicated in candidates showing (1) uncontrolled hemorrhages, (2) obvious brain damages , and (3) significant hemodynamic instability before the intervention.Our indication of VV-ECMO was a deteriorating hypoxia ; and the outcomes (weaning off VV-ECMO and surviving to discharge). For practical purposes, we made some differences in the RESP score to its original version. First, we defined patients having an “aspiration pneumonitis” rather than a “bacterial pneumonia” if the diagnosis of “aspiration” was made, although they had an identified pathogen in their sputum cultures before ECMO. Second, we assigned the patients with fungal pneumonia to the category of bacterial pneumonia. Third, we excluded 3 items  in the calculation of RESP score. As our inclusion criteria of VV-ECMO for ARF in adults, patients of VV-ECMO should be paralyzed with neuromuscular blockade agents before ECMO and have a relative stable hemodynamics. We excluded the item of nitric oxide inhalation because the information of nitric oxide inhalation was often missing in our EMR system before 2012. Fourth, we assigned a SOFA neurological assessment score to each patient according to his/her neurological status before sedation , since all of them were sedated before VV-ECMO.We retrospectively collected the demographic and clinical variables required for calculating the RESP score, ECMOnet score, and SOFA score before the administration of VV-ECMO in each patient, from our institutional electronic medical record (EMR) system. These medical records were registered from March 2007 to April 2015. The following variables were collected: age; sex; etiologies of ARF ; immunocompromised status 2.4,19,20Figure 2, and gradually increase the ECMO pump flow to achieve a steady flow that carries the best pulse oximetry-detected oxyhemoglobin saturation (SpO2). To rest the lung on VV-ECMO, we change the setting of MV to a lung-protective level step by step. At first, we use a pressure-control mode with a PIP ≤35 cm H2O and a moderate PEEP (often 10–14 cm H2O) to obtain an estimated tidal volume ≤6 mL/kg/min on VV-ECMO. Then we take arterial and the postoxygenator blood samples to adjust the sweep gas flow and the MV FiO2. We also adjust the pump speed dynamically to provide a best SpO2 (>90%) and SaO2 (>85%) to gradually taper the PIP to 30 cm H2O and MV FiO2 to 0.4. The hemoglobin is kept at a level of ≥10 g/dL to increase the capacity of oxygenation. Keeping a modest anticoagulation on VV-ECMO with systemic heparinization is necessary except in the hemorrhagic patients. The therapeutic range of activated partial thromboplastin time is 40 to 55 seconds. In patients showing significant improvements, we would try to wean the patient from VV-ECMO as long as the arterial oxygenation could be maintained under the lung-protective ventilation, with a MV FiO2 ≤0.6.We have thoroughly described our techniques and therapeutic protocol of VV-ECMO in our previous publications.2.5P < 0.05. The independent t test was used for univariate comparisons of the independent numerical variables. If the numerical variables were not normally distributed, the Mann–Whitney U test was used for univariate comparisons. The chi-square or Fisher exact test was used to compare the categorical variables. Data were presented as mean ± standard deviation (SD) for numerical variables with normal distribution or median (interquartile range) for numerical variables without normal distribution. The categorical data were presented as number (percentage). The predictive power of hospital mortality of each score was presented as the area under receiver-operating characteristic curve (AUROC). The AUROCs among prediction models were compared with a nonparametric approach suggested by Hanley and McNeil. The multivariate logistic regression method with backward stepwise selection was used to develop the institutional mortality prediction model. All variables with a P < 0.05 in univariate tests were tested by the multivariate logistic regression analysis. Statistical significance was assessed at the level of P < 0.05. The Hosmer–Lemeshow test was used to assess goodness of fit for the institutional model. The Kaplan–Meier method was used to calculate the survival on VV-ECMO. The endpoint for the survival analysis was death on VV-ECMO.Statistical analyses were performed with SPSS for Windows . For all analyses, the statistical significance was set at 33.1Staphylococcus aureus, Pseudomonas aeruginosa, and Acinetobacter baumannii); viral pneumonia ; trauma and inhalation injury (n = 21); aspiration pneumonitis ; and others . The median duration of MV before VV-ECMO was 3 (1–8) days. The mean values of pre-ECMO SOFA score, RESP score, and ECMOnet score were 11 ± 2, 0 ± 3, and 5 ± 2, respectively. Seventy-three per cent (n = 78) of the patients were weaned off VV-ECMO and 55% (n = 59) of them survived to hospital discharge. Eight patients died for the major hemorrhagic complications  during the support of VV-ECMO. The other nonsurvivors (n = 40) showed a dependence on respiratory supports, either VV-ECMO or MV, and died with sepsis or multiple organ failure. The median values of ECMO stay and hospital stay were 9 (5–15) days and 43 (26–47) days, respectively. The actual survivals of RESP score were 75% in class I (score ≥6), 68% in class II (score 3–5), 63% in class III (score −1 to 2), 24% in class IV (score −5 to -2), and 38% in class V (score ≥−6) in this study. Figure The mean age of the 107 patients was 55 ± 16 years and 73% (n = 78) of them were male. Figure jury n = 1; aspira3.2Y) = еX/(1 + еX); X = −3.173 + 0.208 × (SOFA score before VV-ECMO) + 0.148 × (MV day before VV-ECMO) + 1.021 × (immunocompromised status). The model explained 33.3% (Nagelkerke R2) of the variance in hospital mortality and correctly classified 69.2 % of the cases . This predictive model also well-fitted the dataset . Table Since the published scores showed unsatisfactory results, an institutional mortality prediction model was developed with the multivariate logistic regression analysis. Table 4,10 Both of them are derived from multicenter databases and verified with rigorous statistical methodologies. We also incorporated the SOFA score and compared its predictive power with the above mentioned scores, as the authors did in their original studies.,10The study was aimed to find a practical model to predict hospital mortality in adult patients receiving VV-ECMO for severe ARF. Such a tool may theoretically improve the successful rate of this resource-demanding therapy by excluding the poor-prognostic candidates from VV-ECMO, or by actively recruiting the good-prognostic candidates before clinical deterioration. To meet this purpose, we started to search the ideal model among the published scores that have been used for mortality prediction in adult respiratory ECMO. We chose 3 scoring systems  that estimate hospital mortalities based on patients’ characteristics presented before ECMO. The RESP score and the ECMOnet score are specific designed for adult respiratory ECMO and launched recently.,14,23  belonged to undetailed categories of “others acute respiratory diagnoses” and “other (diagnosis).” All of the definite diagnoses  have similar survival rates, from 64% to 70%. The overall survival rate of patients with definite diagnoses is significantly higher than the overall survival rate of the patients with undetailed diagnoses . Thus, RESP score may become more attractive to physicians if it has a simplified classification for the diagnoses in ARF. Except the complex definitions in score parameters, these “packaged” scores may also have decreased predictive powers in a specific disease because of neglecting factors which have significant impacts on prognosis. For example, studies using SOFA score to predict hospital mortality in ECMO-treated patients often yield unsatisfactory results. An obvious disadvantage of using SOFA score to predict hospital mortality in adult respiratory ECMO is not taking the length of pre-ECMO MV into consideration, which is an important poor-prognostic factor in this patient group.However, all the 3 scores showed a real but unsatisfactory predictive power of hospital mortality in this medium-sized cohort. This was not surprising since similar results are shown in some recently published single-center studies of VV-ECMO23 Table . In our  since the mortality rate may be extremely high.Despite none of the 3 published scores performing perfectly in this study, they do help physicians to see the big picture of adult respiratory ECMO. Rather than their predictive powers of mortality in a specific cohort, these published scores are more important in providing the comprehensive knowledge of important prognostic factors for adult respiratory ECMO. These scores therefore may guide investigators to find their future directions for outcome improvement in a more efficient way. During the validation process of the 3 packaged scores, we also needed to perform a detailed analysis of all the parameters included in the scores. We found that the duration of MV, the immunocompromised status, and the SOFA score were independent predictors of hospital mortality in adult patients who were ready to be treated with VV-ECMO. Therefore, we built our institutional model with the 3 independent predictors and yielded an increased discriminative power than the original scores in this cohort. The model could be easily applied at bedside with a table demonstrating the estimated risks of mortality according to the combinations of the 3 predictors Table . Based o,4 Many studies have separated these patients from the discussion of adult respiratory ECMO since they have a different risk of hospital mortality than the patients who could be stabilized by VV-ECMO.,13,14 In fact, no matter what configuration of ECMO is chosen, the prime task of the ECMO specialists is to quickly stabilize the patient. A pre-ECMO echocardiography is very helpful in deciding the ECMO configuration.,25 The VA rather than VV-ECMO should be used in patients showing a strained right ventricle with significant tricuspid regurgitation in bedside echocardiography. The VA-ECMO directly improves the arterial oxygenation and provides a better perfusion of vital organs than VV-ECMO. However, since the VV-ECMO provides a prepulmonary gas exchange and is more physiological, the VA-ECMO may need to be changed to VV configuration after a couple of days if the native lungs are not well recovered. Therefore, heart–lung interactions before and during different configurations of ECMO should be an important direction of research to define the optimal timing and configuration of adult respiratory ECMO in specific scenarios.The major limitations of this study are its retrospective design and limited sample size. It was a pilot study and the model would need to be validated in different settings and populations. It did not provide a comprehensive discussion of adult respiratory ECMO since some high-risk patients were excluded to increase the homogeneity in patient management. These high-risk groups were patients who died quickly on VV-ECMO and patients who required VA or mixed configurations of ECMO. Patients who died quickly on VV-ECMO often showed refractory hypotension soon after the device implantation, and there is often insufficient evidence to draw a solid conclusion about the cause of their deaths or to predict it. Patients requiring VA or mixed configurations of ECMO often have an advanced disease and showed unstable hemodynamics due to a combined cardiopulmonary failure.5The RESP score, the ECMOnet score, and the SOFA score provide valuable information about the poor-prognostic factors for adult respiratory ECMO. Nevertheless, some adjustments of the score parameters may be necessary to fit the clinical demand among institutions. Among the score parameters, duration of MV, immunocompromised status, and severity of organ dysfunction may be the most important prognostic factors of VV-ECMO used for adult respiratory failure.


Experimental evidence in animal models suggests that misfolded Amyloid-β (Aβ) spreads in disease following a prion-like mechanism. Several properties characteristics of infectious prions have been shown for the induction of Aβ aggregates. However, a detailed titration of Aβ misfolding transmissibility and estimation of the minimum concentration of biologically active Aβ seeds able to accelerate pathological changes has not yet been performed. In this study, brain extracts from old tg2576 animals were serially diluted and intra-cerebrally injected into young subjects from the same transgenic line. Animals were sacrificed several months after treatment and brain slices were analyzed for amyloid pathology. We observed that administration of misfolded Aβ was able to significantly accelerate amyloid deposition in young mice, even when the original sample was diluted a million times. The titration curve obtained in this experiment was compared to the natural Aβ load spontaneously accumulated by these mice overtime. Our findings suggest that administration of the largest dose of Aβ seeds led to an acceleration of pathology equivalent to over a year. These results show that active Aβ seeds present in the brain can seed amyloidosis in a titratable manner, similarly as observed for infectious prions. Accumulation of misfolded Aβ has been linked to cell and synaptic toxicity in both Sc) of a normally synthesized protein, termed PrPC . PMDs include several neurodegenerative disorders, such as Parkinson's and Huntington's diseases, and prion diseases or transmissible spongiform encephalopaties (TSEs), among othersrPC Ref. . The mecin vivo1112131415161718111213141615Several studies have shown that Aβ and other disease-associated misfolded protein aggregates are experimentally transmissible in vivo in a similar way as performed for infectious prions.One of the important characteristics of the prion agent is that its capacity to transmit pathology is directly proportional to the quantity of the misfolded protein inoculated and that transmission can be observed even after administration of highly diluted concentrations of prion-infected brain extracts. Titration of infectious prions is useful to estimate the load of infectious agent in specific samplesSeveral previous experiments have shown that brain amyloidosis can be accelerated after intra-cerebral (i.c.) injection of misfolded Aβ aggregates in transgenic mouse models of AD111213141516−2 to 10−7  inoculum and sacrificed the animals 21 days later. Brains from these animals were compared to the ones from mice receiving an equivalent inoculum but sacrificed 230 days later (285 days old) as described for the main experiment. We did not observe presence of anti-Aβ staining in animals sacrificed at the shorter time point , generated an extent of Aβ deposition as high as the one induced by the 10% brain extract. At the dilutions tested, we were not able to observe a saturation of the amyloid inducing activity.In order to extrapolate our results to the biological setting, and taking advantage that the spontaneous accumulation of amyloid deposits in these transgenic mice progressively increases with time, we sacrificed non-treated animals at different time points and measured the amyloid burden in their brains. Mice were sacrificed either at 15–16, 17–18 or 19–21 months old and brain slices analyzed histologically for the presence of Aβ aggregates . The amyAlthough similar total Aβ burdens were observed in some induced vs. aged groups of transgenic animals, the brain regional distribution of Aβ aggregates found in each case was different. In our experiments, induction of Aβ aggregation was performed by i.c. administration of seeds into the hippocampus. Interestingly, a higher burden of Aβ deposits was found in this anatomical structure compared to old animals having the same burden considering cortex and hippocampus together . On the It is thought that in the brain of AD patients accumulation of Aβ deposits leads to changes in the phosphorylation and aggregation of the Tau protein, resulting in the formation of neurofibrillary tangles. To investigate whether extensive deposition of Aβ aggregates induced tau hyperphosphorylation, we stained brain slides from animals in various conditions with an antibody that recognizes hyperphosphorylated tau. Under our experimental conditions, we were unable to detect hyperphosphorylated tau in the brain of animals injected with the most concentrated brain extract or even in non-injected old animals .in vitro and in vivo experiments exploring the prion-like features of Aβ aggregates, added to the current knowledge gathered on TSE-prions, have posited a novel mechanism for the spreading of these deleterious molecules during the disease. A strong debate currently exists on whether Aβ and other misfolded protein aggregates are prions in the sense of their mechanism of transmission, principally due to the lack of evidence of natural inter-individual transmission. So far, no epidemiological data suggest that misfolded Aβ is able to transmit disease between individuals. However, it is important to highlight that human prion diseases are not easily transmissible by standard routes and the onset of clinical disease normally occurs after years or decades of a silent incubation period.During recent years there has been great interest to explore the putative prion-like features of protein aggregates involved in PMDs other than TSEs. Among them, tau and Aβ have taken a highlight place due to their involvement in AD, the most common type of senile dementia. Previous Sc properties have still not been explored, feeding the current controversy on whether misfolded Aβ is actually a prion. Among the non-explored prion features of misfolded Aβ we can include the titration of the agent, which is a common test to measure the specific PrPSc transmissible activity. Prions can efficiently maintain infectivity even after subjecting infected samples (i.e. brain tissues) to high dilutions. The purpose of the current study was to titrate biologically active Aβ seeds in vivo and define the minimum dose capable to induce pathological changes. The experiments described in this article were done with tg2576 mice, which is a transgenic line that over-express the human Amyloid Precursor Protein (APP) harboring the Swedish mutation and as a consequence starts developing the first visible Aβ aggregates at ~9 months oldSeveral properties are often observed associated to prions and are considered necessary for them to behave as infectious agents8101112122627de novo generation of Aβ aggregates and not to the original inoculum −5 to 10−7  induced amyloid deposition to a comparable extent of that observed spontaneously in these transgenic mice at around 19–21 months of age. In other words, our results suggest that injection of ~3.43 ng of aggregated Aβ (the amount estimated in a 10−2 dilution) is able to accelerate brain amyloid pathology in an extent equivalent to 10–12 months. However, and in agreement with a previous report32The availability of a “titration curve” for the transmissibility of Aβ aggregates may be useful to assess the specific burden of Aβ seeds present in different tissues or biological fluids, as well as to quantify the effect of disease modifying strategies directed to understand or combat AD. Interestingly, the degree of the induction of amyloid pathology obtained with distinct dilutions of the brain extract was comparable to that observed in non-treated animals sacrificed at different ages . In our In conclusion, in this study we show that biologically active Aβ seeds are titratable and that the minimum amount able to induce pathological changes is in the same range as described for infectious prions. In the light of this data, it is important to further explore the prion-like properties of Aβ aggregates in order to fully understand how these particles spread and finally exert their deleterious effects in the brain. Comparing the mechanisms of misfolding and aggregation of Aβ and prions may be useful to design novel diagnostic and therapeutic strategies directed to attack the spreading of pathological alterations in AD.ad libitum) as groups of no more than five mice per cage. All animal procedures described in this article were in agreement with the regulations of the Center of Laboratory Animal Medicine and Care (CLAMC) and Animal Welfare Committee (AWC) of the University of Texas Medical School at Houston. 4–10 animals  were used per experimental group as indicated in Tg2576 transgenic mice express the human amyloid precursor protein gene harboring the Swedish mutation−1) was serially diluted in 10-fold dilutions using the same buffer until reaching a 10−7 dilution regarding the brain. Resulting inocula were stored at −80°C until used for animal injection.Brain halves from four 18–20 months old tg2576 mice  or 9 months old tg2576 mice were ultra-centrifuged at 100,000 × g for 1 h at 4°C using a L100K Beckman-Coulter ultracentrifuge . Supernatants were discarded and pellets resuspended in 200 μL of 70% formic acid. Samples were sonicated in a bath sonicator and centrifuged for 30 min in the same conditions listed above. Supernatants were collected, diluted 20 times in 1 M Tris buffer (pH 11) and snap frozen in liquid nitrogen. Resulting samples were stored at −80°C until tested for Aβ concentration using an Aβ42 specific ELISA kit  or an ELISA kit detecting total Aβ . Extractions and ELISA measurements were performed two times in duplicates by two different investigators. In order to assess the linearity of the diluted material, a 1.25% brain homogenate sample of the original inoculum was 2-fold serially diluted in PBS three independent times. Resulting samples were diluted 1/5 in 88% formic acid and further diluted 20 times in 1 M Tris buffer for neutralization. Aβ concentration was measured by ELISA as described.200 μL of 10% w/v brain homogenates from the original inoculum  days old tg2576 mice were i.c. injected with different tg2576 brain homogenate dilutions . Injecti2O2/10% methanol in PBS, for 30 min. Then, brain sections were incubated in 85% formic acid for 5 min for antigen retrieval. Sections were incubated  with the mouse 4G8 antibody , which recognizes the 17–24 sequence of Aβ, diluted 1:1000 in PBS/0.02% Triton-X100 , or anti-hyperphosphorylated tau AT8 antibody, diluted 1:200 in the same buffer (Pierce). After washing with PBS, sections were incubated for 1 h with an HRP-linked secondary sheep anti-mouse antibody at a 1:500 dilution . Peroxidase reaction was visualized using DAB Kit  following the manufacturer's instructions. Finally, sections were dehydrated in graded ethanol, cleared in xylene, and cover-slipped with DPX mounting medium . ThS staining was performed by incubating tissue slices with a ThS  solution (0.1% in 50% ethanol) for 15 minutes after deparaffinization. After incubation, sections were washed for 2 minutes in 80% ethanol, dehydrated and cover-slipped as described.Formalin fixed brains were dehydrated for paraffin inclusion. Brains were serially sliced in 10 μm thick sagittal sections from the midline. 5 slices (one every tenth slice) per animal were processed for immunohistochemistry. Briefly, sections were deparaffinazed and the endogenous peroxidase activity was blocked with 3% HFor quantification, brain slices were examined under a DMI6000B microscope  and image analysis was performed using the ImageJ software . 4G8 burden was defined as the antibody labeled area in each tissue slice per total area analyzed , and expressed as percentage.−1 to 10−5) or Mann-Whitney U-test (10−6 and 10−7) were used to compare Aβ burden in injected and non-injected animals. The values are expressed as means ± SEM. Data was analyzed using the Graph Pad prism software, version 5.0. Statistical differences were considered significant for values of P < 0.05.Kolmogorov-Smirnov (K-S) statistic test was used to confirm normal distribution of the data. According to their distribution, Student's t-test (10R.M. designed the experiments, performed most of the animal manipulations, analyzed the data, and wrote the manuscript. J.B.-A. performed most of the histological analyses and prepared the final version of the figures. C.D.-A. participated in some animal and histological analyses. C.S. designed the experiments, supervised the project and wrote the manuscript.Supplementary figures


A re-refinement of 4mwk, namely, the X-ray triclinic crystal structure at 100 K of cisplatin binding to histidine in a protein in a multi-temperature structural dynamics study, has been made. This is so as to correct clashes in the protein structure solute molecules model for which our attention was drawn by the valuable critique of this PDB deposition by We have considered the His 15 imidazole ring split occupancy (60%/40%) placements in 4yeo but conclude that there was insufficient 2Fo-Fc electron density at the1.2 rms contour level and no Fo-Fc electron density contoured at the COOT default cut off of 5σ for such a precise interpretation. But, as we stated in our paper , the HisIn 4mwk, we had imported the PDB's “CPT” ligand and then deleted ligand atoms that did not fit the electron density. We realise that this was due to the PDB's CPT having an incorrect, tetrahedral, rather than square planar, geometry. In preparing this revision of 4mwk, we have now removed the CPT labels on the atoms that we had and instead assigned individual atoms and which is hopefully clearer in our PDB files.In checking the PT labels, so as to ensure the same labelling nomenclature between the PDB files and the article, again for clarity, we noticed an error in Tables II and III concerning the carboplatin 4oxe NE side PT4 which in fact is PT1. Since 4oxe does not feature in Figure 2, being entirely cisplatin at the three temperatures, this label error in Table III does not affect Figure 2.The revised-4mwk-cycle-27 PDB validation server report led to our deleting the top three listed poor RSR and LDF moieties, namely, “ACT N 1,” “NO3 I 4,” and “PT L.” The fourth peak DMS H 1 has a good density; we therefore retain this moiety. The fifth peak, CL J, makes a sensible ligand distance to PT C at 2.30 Å away; so we retain this ligand atom. There are some “blobs of electron density” in the COOT final Fo-Fc “validation difference map”  that we Structural Dynamics article is a comparison of identically prepared crystals' diffraction data measured at three different temperatures, and carboplatin binding in the triclinic crystal form at one temperature, we have also revisited 4mwm, 4mwn, and 4oxe to again (i) remove any clashing solute molecules and (ii) replace the potentially confusing use of the incorrect “CPT.” We have removed solute molecules which we are not certain as to their chemical identities. In all these cases above , the as previously published processed structure factor amplitudes' data have been used. Table Since In our re-refinements above, we have harnessed the validation of the protein molecular model incorporated in the refinement program PHENIX_REFINE .The new PDB codes are listed in Table


Stroke is the fifth leading cause of death and the primary cause of long-term adult disability in the United States. Increasing evidence suggests that low T3 levels immediately following acute ischemic stroke are associated with greater stroke severity, higher mortality rates, and poorer functional outcomes. Prognosis is also poor in critically ill hospitalized patients who have non-thyroidal illness syndrome (NTIS), where T3 levels are low, but TSH is normal. However, data regarding the association between TSH levels and functional outcomes are contradictory. Thus, this study investigated the role of TSH on stroke outcomes, concomitantly with T3 and T4.In this work, blood was collected from patients with radiologically confirmed acute ischemic stroke at 24±6 hours post-symptom onset and serum levels of TSH, free T3, and free T4 were measured. Stroke outcomes were measured at discharge, 3 and 12 months using the modified Rankin scale and modified Barthel Index as markers of disability. Though we found that lower levels of free T3 were associated with worse prognosis at hospital discharge, and at 3 and 12 months post-stroke, none of these outcomes held after multivariate analysis. Thus, it is likely that thyroid hormones are associated with other factors that impact stroke outcomes, such as sex, age and stroke etiology.This study found that lower levels of free T3 were associated with poorer outcomes at hospital discharge, and at 3 and 12 months post stroke, however, these associations diminished after correction for other known predictors of stroke outcome. Thyroid hormones have a complex relationship with ischemic stroke and stroke recovery, which merits further larger investigations. Stroke is the fifth leading cause of death and the primary cause of long-term adult disability in the United States . PerturbThe relationship between thyroid hormones and functional outcomes post-stroke is complex. Current data has shown that low T3 levels immediately following acute ischemic stroke (AIS) are associated with greater stroke severity and mortality, and poorer functional outcomes –8. This This study examines the association between thyroid hormone levels  and ischemic stroke outcomes as determined by the modified Rankin scale (mRS) and modified Barthel index (mBI) at hospital discharge, and at 3 and 12 months.This study was conducted at an 868-bed community-based teaching hospital with a Neurology Residency Program. This center has The Joint Commission’s Gold Seal of Approval and the American Heart Association’s Advanced certification as a comprehensive stroke center and has an annual stroke admission rate of approximately 1000. This study was approved by the Institutional Review Board.n = 129) at 24 ± 6 h post-symptom onset under an approved IRB from December 2011 to May 2013. Patients with active malignancy or treatment with immunosuppressants were excluded from the study. Serum levels of TSH, free T3 (fT3), and free T4 (fT4) were quantified by solid-phase chemiluminescent immunometric assay .Blood was collected from patients over 18 years of age with radiologically confirmed acute ischemic stroke (AIS)  was collected. Risk factors such as history of hypertension, atrial fibrillation, coronary artery disease, previous stroke or transient ischemic attack, diabetes mellitus, hypercholesterolemia, cigarette smoking and alcohol use were also collected. Primary outcomes were in-hospital mortality and National Institutes of Health Stroke Scale (NIHSS) score on admission. Secondary outcomes included death or hospice after discharge, and modified Rankin scale (mRS) and modified Barthel Index (mBI), at 3 and 12 months post-stroke. The mRS is a scale for measuring the disability of patients post-stroke. It has a scale of 0–6, with 0 designating no limitations or symptoms, 5 being severe disability requiring constant care, and 6 being dead . The mBIContinuous data, such as age, are presented as means (standard deviation). Non-normally distributed continuous and ordinal data are presented as medians (interquartile range) and were analyzed with non-parametric tests (Spearman’s Rho). Categorical data are presented as proportions, and group differences were assessed with chi-square tests of proportion. Several variables with non-normal distributions were dichotomized and analyzed with chi-square test of proportions. These included the mBI, dichotomized into “independent” (score of 15 or greater), or “dependent” (14 or less), and mRS, dichotomized into “disabled” (score of 3 or greater) or no disability to slight disability, termed “abled” (2 or less). A “poor outcome” composite measure was defined as death or disabled with an mBI ≤14 or a mRS ≥3 as defined above at 3 and 12 months after AIS. A “favorable outcome” composite measure was defined as mBI ≥15 or mRS ≤ 2. Significant findings identified in the univariate analyses that pertained to more than one thyroid marker were reanalyzed using multivariable logistic regression to control for all significant confounding variables. Thus, a unified list of predictive factors was applied to all three thyroid markers in multivariate analysis. This included atrial fibrillation (AFIB), depression, urinary tract infection (UTI), intra-arterial devices (IA), age and NIHSS on admission. The criterion of statistical significance was set at 0.05. All analyses were performed using Statistical Package for the Social Science v14.Patient demographics are listed in Table As expected TSH was inversely correlated with fT4, though there was no significant correlation between TSH and fT3. fT3 and age were inversely correlated, but there was no significant association with age for TSH or fT4. There was no significant difference between males and females in TSH, fT4, or fT3 levels Table .Table 2Ar = −0.393, p < 0.001) and lower mBI scores  , but not TSH or fT4. Hospital mRS was inversely associated with TSH , and directly associated with fT4 . There was no association with fT3. Admission mBI score was unrelated to fT3, fT4, or TSH levels (Table The NIHSS on admission was inversely associated with fT3 (p = 0.015). There was no association between hospital mortality and fT3 or fT4. However, patients who were alive at discharge had significantly higher fT3 levels than patients who either died in hospital or went to hospice (p = 0.01) . Patients who died at 3 months had lower TSH values than those who were alive at 3 months. Higher 24 h fT3 levels were predictive of good outcome at 3 months as measured by mBI > 15 (p = 0.004). In addition, patients with a favorable composite outcome at 3 months had higher TSH levels than those with a poor composite outcome . This was confirmed by an inverse association between mRS and fT3 levels, showing that the higher the initial fT3, the better the outcome, even a year after the index event (p = 0.042). Neither mBI nor the mRS at 12 months was related to TSH or fT4 levels (Table The same pattern was seen at 12 months. Patients with the highest fT3 levels were the most likely to be independent as measured by the mBI 1 year after ischemic stroke (p = 0.004) with a mean rank of 83 for small vessel disease, 59 for large vessel disease, and 53 for cardioembolic stroke etiology.There were differences in fT3 based on stroke etiology determined with the TOAST classification. Patients with small vessel disease had higher fT3 values than those with large vessel or cardioembolic stroke etiology (H(2) = 11.06, A multivariate logistic regression model was used to control for the potential confounders identified in the univariate analyses as described above. When controlling for AFIB, depression, UTI, IA, age and NIH on admission, thyroid markers were not associated with mortality or functional outcomes Table , suggestThis study is the first to look at TSH, fT3, and fT4 concurrently and their association with stroke severity and long term functional outcomes. This study found that low TSH and fT3 were associated with poor function at 3 months, and that low TSH and fT3 were associated with higher rates of death in the hospital. However, these results were not seen in the multivariate analysis when other known predictors of outcome such as stroke severity (NIHSS) were controlled. This is likely due to the relatively small sample size of the study, and the fact that the majority of patients sampled were euthyroid. Though prior studies have demonstrated an association between low T3 levels and poor outcomes, it is likely that this study was underpowered to detect an association. Outcomes at 12 months were only available in a minority of patients. Future studies with an increased number of patients with hyper- and hypothyroidism and hypothyroidism with hormone replacement should be done to assess a wider range of thyroid values and outcomes. In addition, assessment of hormone levels over multiple time points is needed to demonstrate changes in thyroid status over time, as only one time-point was evaluated here. However, the significant association between thyroid hormone levels and outcome in the univariate models suggest that this is worth pursuing vigorously in larger studies, as strategies to replace thyroid levels could be developed for therapeutic trials.Although it is unknown whether administration of exogenous thyroid hormone in stroke patients improves outcomes, administration of T3 resulted in increased mRNA expression of reelin and brain-derived neurotrophic factor in rats, both of which are important in nervous system regeneration . In hosp


The present work reveals that at the sub-millimeter length-scale, molecules in the liquid state are not dynamically free but elastically correlated. It is possible to “visualize” these hidden elastic correlations by using the birefringent properties of pretransitional swarms persistent in liquids presenting a weak first order transition. The strategy consists in observing the optical response of the isotropic phase of mesogenic fluids to a weak (low energy) mechanical excitation. We show that a synchronized optical response is observable at frequencies as low as 0.01Hz and at temperatures far away from any phase transition (up to at least 15°C above the transition). The observation of a synchronized optical signal at very low frequencies points out a collective response and supports the existence of long-range elastic (solid-like) correlations existing at the sub-millimeter length-scale in agreement to weak solid-like responses already identified in various liquids including liquid water. This concept of elastically linked molecules differs deeply with the academic view of molecules moving freely in the liquid state and has profound consequences on the mechanisms governing collective effects as glass formation, gelation and transport, or synchronized processes in physiological media. Liquids differ from solids by a delayed response to a shear mechanical solicitation; i.e. they have no shear elasticity and exhibit a flow behavior at low frequency < 1Hz). This postulate might be not verified at all length scale. Several recent experimental observations Hz. This  highlighThe article is constructed as followed. We first examine the optical behavior of the isotropic liquid of two liquid crystalline fluids upon low frequency mechanical strain. We reveal that it is possible to strain-induce a spectacular low frequency birefringence. This so far unknown property indicates that the isotropic liquid loses entropy to gain orientation . The anaTwo liquid crystalline fluids (LCF1 and LCF2) selected to represent the liquid behavior of a small molecule and of a large molecule, are examined in their isotropic liquid phase. LCF1  is a 40 -5s). The strain amplitude is defined by the displacement length δl divided by the sample thickness(γ0 = δl/e). Typical displacements and strain amplitudes are of the order of 4 to 250μm for e = 250μm and γ0 varying from 2 to 100% respectively.The optical experiments were performed in transmission mode under crossed polarized microscopy (Olympus BX60). Oscillatory shear motion was applied by a home-improved CSS450 Linkam cell (with a temperature gradient < +/- 0.05°C) equipped with quartz plate-plate fixtures. The used wavelengths were λ = 420nm and 614nm respectively. The transmitted intensity I was measured with a CCD camera (Vision Technology) and normalized to the uncrossed polarizer intensity I0. The averaged birefringence 〈Δn〉 is calculated using the formula 0). An optimized transfer of the motion to the liquid was achieved by ensuring total wetting boundary conditions between the liquid and the substrate  and by probing sub-millimeter gaps . T–8. T1–8]ce curve  characteThe present results confirm that the isotropic phase is an elastically correlated “self-assembly” in agreement with stress measurements indicating a weak elastic modulus at low frequency –8. In li


Conjugates of the photoactivated rhodamine dyes with biopolymers  are important tools for microscopic investigation of biological tissue. In this study, a precursor of the photoactivated fluorescent dye (PFD) has been successfully used for staining of numerous mammalian cells lines and for conjugate formation with chitosan (“Chitosan-PFD”) and histone H1 (“Histone H1.3-PFD”). The intensive fluorescence has been observed after photoactivation of these conjugates inside cells . Developed procedures and obtained data are important for further application of novel precursors of fluorescent dyes (“caged” dyes) for microscopic probing of biological objects. Thus, the synthesized “Chitosan-PFD” and “Histone H1-PFD” have been successfully applied in this study for intracellular transport visualization by fluorescent microscopy. Design of various conjugates of fluorescent dyes and their nonfluorescent precursors with proteins, polysaccharides, nucleic acids, and other biomolecules are the rapidly growing fields at the “edge” of molecular biology and chemistry, biophysics, and biochemistry –3. Such The main goal of this work was to study multifunctional precursor's conjugates of the fluorescent dye (PFD) with chitosan and histone H1 in order to visualize the transport of these biopolymers into the cells of different types.The synthesis and properties of the “caged” PFD dye  have beeμg/mL in the cell culture media from the stock solutions in DMF at 10 mg/mL and in DMSO at 2 mg/mL. Cells were incubated for 30 min at 37°C in CО2 incubator. During this incubation (15 min after beginning), the nuclei staining dye Hoechst 33342 was added to cell cultures. After incubation, cells were fixed for 15 min and washed with phosphate buffer (pH 7.0). These samples were mounted on glass slides using Mowiol 4.88  and stored at 4°C overnight for the complete polymerization. All samples were analyzed using confocal microscope Eclipse TE2000 (Nikon) with EZ-C1 3.90 Free Viewer software. The images were compared by color intensity in different channel using software “ImageJ” and plugin “Colocalization Finder.”The following PFD concentrations were used: PFD solutions from 0.02 to 10 μg/mL. Cells on coverslips were washed with phosphate buffer and mounted using Mowiol 4.88 on slides after incubation for 1 hour.A chitosan with a molecular weight of 50 kDa (degree of deacetylation of 96%) and PFD-NHS were used for the “Chitosan-PFD” conjugate preparation. In particular, chitosan amino groups were acylated with PFD-NHS to form an amide bond and N-hydroxysuccinimide. PFD-813 and PFD-NHS with chitosan solutions (for comparison and verification) and the “Chitosan-PFD” conjugate were added to the plate for 1 hour pregrown on coverslips cell monolayers at final concentration of 2.2 μL DMSO and added to 200 mL of bicarbonate buffer (50 mM), containing 2 mg of histone H1.3; (2) the resulting obtained mixture was stirred for 1.5 hours at 20°C; (3) the mixture was neutralized by 0.1 N acetic acid and distilled water was added to 500 mL final volume. Succinimide group of PFD promotes formation of the covalent bonds with primary amino groups (lysine and arginine residues) of histone H1.3 to form the conjugate “Histone H1-PFD”  with PFD (tetramethylrhodamine derivative), capable of bright fluorescence in the red region of the spectrum (after photoactivation), was synthesized by standard procedure: (1) 0.5 mg PFD-NHS was dissolved in 50  H1-PFD” . The conμg/mL in all cases. High resolution images of various cell types were obtained after light-induced activation of PFD  was published in the recent papers , 18, 20.n of PFD .The best micrographs were obtained after photoactivation of “live” cells stained by PFD. The advantages of such methodology (making preparations of “live” cells) are the following: the absence of the fixing agent ; replacement of the washed buffer by fresh culture medium; and the exclusion of a polymerizable material . The only disadvantage of such methodology is the necessity of fast measurements of such samples because of the rapid evaporation of the culture medium between the glass plates.As can be concluded from the previous experiments, PFD is concentrated mainly in the intracellular organelles of the “live” cells  as compaThe rhodamine derivatives can be used to study biopolymer transport using fluorescent microscopy. Chitosan is important biopolymer for modeling and applied studies of the BAS transport in various cells. Chitosan heterogeneous clusters in the cells were observed around the nucleus after photoactivation . The imaμg/mL “Histone H1-PFD”. Microphotographs of “native” and fixed cells in order to evaluate the intracellular distribution of the conjugate were obtained using the confocal microscope . It is found in all cases that the obtained conjugates after cell penetration can be seen as small aggregates (size about 1 μm or less) which are clearly distinguishable in all studied cells  and drug delivery systems into human and animal cells.Properties of the novel PFD before and after photoactivation to fluorescent rhodamine derivative (Rho813) have been characterized in the fixed and “live” cells. The PFD demonstrated better cell staining properties as compared to other rhodamine caged dyes. Such dyes are very stable during photoactivation, have intensive fluorescence even at low dye concentration, and easily penetrate various cell types, selectively staining some organelles (mainly mitochondria and lysosomes). Conjugates of chitosan or histone H1.3 with PFD were obtained as examples to investigate the distribution of different biopolymers within cells. The intensive fluorescence has been observed after the photoactivation of these conjugates inside different cell cultures . The synthesized “Chitosan-PFD” and “Histone H1-PFD” have been successfully applied in this study for intracellular transport visualization by fluorescent microscopy. The PFDs potential for nanoscale optical resolution renders them particularly suited for special tasks in biology and bionanotechnology and human and animal medicine.


The method is based on Fourier domain pre-processing via the Born approximation, followed by the Multifractal Detrended Fluctuation Analysis. The approach is experimentally validated in synthetic multifractal scattering phantoms, and tested on biopsy tissue slices. The derived multifractal properties appear sensitive in detecting cervical precancerous alterations through an increase of multifractality with pathology progression, demonstrating the potential of the developed methodology for novel precancer biomarker identification and tissue diagnostic tool. The novel ability to delineate the multifractal optical properties from light scattering signals may also prove useful for characterizing a wide variety of complex scattering media of non-biological origin.Multifractal, a special class of complex self-affine processes, are under recent intensive investigations because of their fundamental nature and potential applications in diverse physical systems. Here, we report on a novel light scattering-based  Studies on structures and processes that exhibit self-similarity have evoked intensive investigations recently because of their fundamental nature and potential applications in diverse areas ranging from complex biological systems, material structures, electrical circuits to various optical phenomena1212578inverse problem; that is, given a tissue light scattering signal (either as a function of angle or wavelength), is it possible to extract its multifractal properties? In tissues, the wide range of spatial scaling of the inhomogeneities and the complicated nature of the spatial correlations present between them contributes in a complex intertwined way to the light scattering signal. The method we propose is based on Fourier domain pre-processing via the Born approximation, followed by the Multifractal Detrended Fluctuation Analysis. The approach is experimentally validated in synthetic multifractal scattering phantoms, and tested on tissue slices (tissue sections of human cervix of different pathology grades). The light scattering-derived multifractal tissue optical properties yield valuable information on subtle changes in the index inhomogeneity distribution of tissue; they also appear sensitive in detecting cervical precancerous alterations through an increase of multifractality with pathology progression. The potential of the developed methodology for novel precancer biomarker identification as a tissue diagnostic tool is thus suggested.Elastically scattered light contains information about the spatial frequency spectrum of the investigated object, providing a useful route to probe the self-similarity in the spatial distribution of its refractive index (RI). Indeed, such light scattering measurements have been explored for characterizing the fractal structures of biological tissues12H (0 < H < 1)244Y(i)  is divided into Ns = int (N/s) non-overlapping segments m of equal length s. The local trend of the series (my(i)) is determined by least square polynomial fitting of each segment m of the series. These are then subtracted from the segmented profile to yield the detrended fluctuations, and the resulting variance is determined asq) dependent fluctuation function is then constructed by averaging the variances over all the segments as q(s) ~ sh(q). Using this approach, any non-stationary multifractal signal can be characterized via two sets of scaling exponents: (i) the generalized Hurst exponent h(q), and (ii) the classical multifractal scaling exponent τ(q), along with the singularity spectrum f(α). These are related asA statistically homogeneous fractal  series is one whose variance exhibits a power law scaling described by a single global Hurst exponent,  0 < H < 123. A muα is the singularity strength and the full width of f(α); Δα (taken at f(α) = 0) is a quantitative measure of multifractality.Where inverse analysis strategy for extraction and quantification of multifractality from the corresponding light scattering signal. Consider a weakly fluctuating scattering medium having normalized RI fluctuations 0 is the average RI of the medium; the fluctuating part of index Δn(r) gives rise to phase distortion and scattering. In the first order Born approximation (weak variations of Δn(r)), the elastic scattering field for scalar excitation can be related to Δn(r) via a Fourier transformation171819β is the scattering vector with modulus β = 2k sin(θ/2), θ is the scattering angle, λ is the wavelength ; σ = n0δn represents the index fluctuation strength and η(r) is the corresponding spatial distribution of index inhomogeneities. It is evident that any signature of multifractality hidden in η(r) would be encoded in I(β), in the Fourier domain. Consequently, for media exhibiting statistical self-similarity in index fluctuations, the information on multifractality can in principle be obtained from the scattering signal as η/(ρ) can be interpreted as index inhomogeneity distribution with spatial scale ρ, representing the randomness of the mediumin statistical sense, ρ = |r − r/| is the distance between any two medium points. In what follows, we demonstrate that η/(ρ) contains the essential multifractal features of index fluctuations in complex systems such as tissues, which can be extracted by employing multifractal analysis. For this purpose, (i) we employ MFDFA directly on the spatial variation of tissue RI, obtained from differential interference contrast (DIC) images of human cervical pathology ; and (ii) quantify the multifractal tissue optical properties by employing MFDFA on the Fourier pre-processed light scattering data η/(ρ) .The MFDFA approach can be directly employed on spatial variation of refractive index to quantify signature of multifractality from any random medium such as tissue−1) is shown in P(ν) ≈ νγ−) (γ) is not uniform throughout the entire ν range  ≈ ν−γ) . As seen ν range , suggest ν range , the ligexponent . A factoforward analysis are shown in qF(s) vs logs with varying moment q furnish supporting evidence of multifractality (h(q) (qF(s) vs logs) and the width (Δα) of the singularity spectrum f(α) (h(q) and considerable magnitude of Δα ( = 0.42) demonstrate strong multifractality in the spatial variations of tissue RI.The corresponding results of MFDFA actality . Its strrum f(α) . Signifiinverse problem of extracting the multifractal optical properties from the light scattering signal. We employ MFDFA on the Fourier pre-processed light scattering data η/(ρ) through fluctuation series synthesized either by fractional Brownian motion  or binomial multifractal model2h(q) spectrum derived from the experimental data and from the theoretical one shows excellent agreement. The experimental value h(q = 2)  =  0.70, that obtained from the theoretical data (h(q = 2)  =  0.67) and the known input for the fluctuation series (h(q = 2)  =  0.69), are in close agreement  correspond to the Hurst exponent of an equivalent monofractal series2inverse extraction of multifractality, and may be relevant for tissue examinations.We now turn to the more challenging ata η/(ρ). While tinverse analysis performed on the light scattering spectra recorded  vs logslogF (h(q) spectrum and the singularity spectrum f(α)   =  0.58] is in good agreement with the forward analysis result [h(q = 2)  =  0.53]. This suggests self-consistency in inverse analysis of multifractality from tissue light scattering spectra. Note that even though the parameter η/(ρ), extracted via Fourier pre-processing, represents the index variations in a statistical sense over the probed tissue volume, it contains contributions of sub-micron level spatial index fluctuations (as evident from q) and the large scale (via the positive moments q) fluctuations and quantify their statistical self-similar properties via h(q) (and τ(q)) and f(α)h(q) (τ(q)) for negative q -values (f(α) (f(α). Note that f(α) contains relatively fewer data points for smaller values of α. This non-ideal nature of f(α)  arises due to the strong contribution of the background Brownian type random fluctuations in the Fourier pre-processed light scattering data η/(ρ). Such background random fluctuations in turn originates from the associated randomness in the spatial index variation (as evident from η/(ρ). Moreover, one may also additionally employ methods such as polarization gating to reduce the signature of tissue absorption on light scattering spectra13 vs logs . The derrum f(α)  exhibitsanalysis . Moreove -values  and the es (f(α)  underscomultifractal tissue optical properties were investigated for discriminating different grades of cervical precancer. Twenty two precancerous and four normal connective tissue sections are analyzed in (i) the values for h(q = 2) or τ(q = 2) are higher in normal tissues and decrease with increasing grades of precancer  = 0.77 ± 0.01, 0.56 ± 0.01 and 0.42 ± 0.01 for normal, Grade I and Grade III tissues respectively); (ii) the magnitudes of Δα are larger for higher grades of precancer . Note that the limiting values for h(q = 2) of unity and zero correspond to smooth Euclidean random field/marginal fractal (implying long range correlations) and a space-filling field/extreme fractal (anti correlated behavior) respectively. Reduction in the value for h(q = 2) with increasing pathology thus likely indicates increasing tissue roughness, or effectively the predominance of index inhomogeneities having smaller dimensions. Connective tissue is comprised of collagen fiber network with complex organization of constituent micro-fibrils; statistical self-affinity of fibrous network may manifest as the observed multifractality in spatial variation of RI. Reduced h(q = 2) may originate from volume fraction reduction of the fibrous network and/or fibers shortening/disorganization (both processes occurring with disease progression)232425Δα) at higher grades of precancer, on the other hand, may be a manifestation of the increased heterogeneity and the different scaling behavior of the small scale  and the large scale (fibrous network) index fluctuations. This suggestion was initially tested in extracted Bovine collagen samples treated with 5% acetic acid (results not shown); this is known to degrade organized collagen fibrous structures by breaking cross-links2325The newly derived inverse method is developed, validated and explored for extraction and quantification of multifractality in the spatial RI fluctuations of biological tissues. Experimental validation of this inverse analysis method in synthetic multifractal scattering phantoms, and the agreement between the forward analysis on differential interference contrast image of tissue and inverse analysis from light scattering spectra, establishes self-consistency. The latter demonstrates that subtle changes in multifractal nature of tissue RI fluctuations can indeed be probed and quantified from light scattering spectra, despite numerous complexities associated with the wide range of spatial scaling of the index inhomogeneities, complex spatial correlations, and the resulting intertwined nature of their contribution to the tissue light scattering signal. The demonstrated ability to delineate multifractal properties may provide a valuable noninvasive tool for characterization of tissue and wide variety of other complex scattering media of non-biological origin. Observed differences in the multifractal tissue optical properties between different grades of precancers show considerable promise as potential biomarkers for early detection. The ability to probe and quantify subtle morphological and structural alterations associated with precancer using the elastic backscattering spectra bodes well for in-vivo deployment, because the backscattering geometry is clinically convenient and the measurement technique is fairly straight-forward. Exploiting the elastic backscattering spectra (singly or weakly scattered) recorded from superficial tissue depths, in-vivo applications of this approach can be realized with a suitably designed fiber optic probes and/or by other methods  to eliminate multiply scattered lightin-vivo detection of precancers remain to be rigorously evaluated. We are currently initiating such investigations in our laboratory.In conclusion, a novel light scattering-based total = 26, with ngrade I = 12, ngrade II = 4, ngrade III = 6; four biopsies from the normal counterparts, nnormal = 4). Standard histological preparation of the excised tissues involving fixation, dehydration, imbedding in wax, sectioning under a rotary microtome , and subsequent de-waxing was employed. The control experiments were performed on the extracted Bovine collagen  samples. The collagen was treated with 5% (weight by volume) acetic acid and the light scattering spectra were recorded both before and after 30 minutes of treatment. Consent for the use of all the biopsy samples of human cervical tissues with normal counterparts in our study was obtained from the Ethical Committee, G. S. V. M. Medical College and Hospital, Kanpur, India. The methods were carried out in accordance with the approved guidelines.Histopathologically characterized  biopsy samples of human cervical tissues with normal counterparts were obtained from G. S. V. M. Medical College and Hospital in Kanpur, India  microscope  was used to measure the spatial distribution of tissue refractive index . Images ents see . Brieflyanalysis .transmissive liquid crystal-based twisted nematic spatial light modulator . The grey levels of the individual SLM pixels were modulated using user controlled inputs of either mono or multifractal fluctuation series. Fractional Brownian motion algorithm was used to generate monofractal fluctuation series with user controlled Hurst exponent (H) and multifractal fluctuation series with user controlled generalized Hurst exponent h(q) was generated using binomial multifractal model (see f), and an EMCCD camera  was at its back focal (Fourier) plane. The recorded intensity distribution at the CCD (x-y) plane represented the spatial frequency , analyzed data  contributed in writing the manuscript. A.V. and A.P. contributed in theoretical analysis/interpretation of data and in writing the manuscript. P.K.P. contributed in theoretical multifractal analysis, interpreting the data and writing the manuscript. S.K. contributed in the control validation studies on synthetic mono/multifractal scattering phantoms .Tissue multifractality and Born approximation in analysis of light scattering: a novel approach for precancer detection


New optical fiber based spectroscopic tools open the possibility to develop more robust and efficient characterization experiments. Spectral filtering and light reflection have been used to produce compact and versatile fiber based optical cavities and sensors. Moreover, these technologies would be also suitable to study N-photon correlations, where high collection efficiency and frequency tunability is desirable. We demonstrated single photon emission of a single quantum dot emitting at 1300 nm, using a Fiber Bragg Grating for wavelength filtering and InGaAs Avalanche Photodiodes operated in Geiger mode for single photon detection. As we do not observe any significant fine structure splitting for the neutral exciton transition within our spectral resolution (46 μeV), metamorphic QD single photon emission studied with our all-fiber Hanbury Brown & Twiss interferometer could lead to a more efficient analysis of entangled photon sources at telecom wavelength. This all-optical fiber scheme opens the door to new first and second order interferometers to study photon indistinguishability, entangled photon and photon cross correlation in the more interesting telecom wavelengths. New emerging quantum optics technologies are mostly influenced by the possibility to design realistic proposals to implement and control quantum correlations between photonsIn this regard, single self-assembled Quantum Dots (SAQDs) are well known solid-state semiconductor nanostructures that offer key advantages as single or entangled photon emitters fabricated on a GaAs substrate. SAQDs show confinement in all dimensions, leading to a 0-dimensional density of states similar to single atoms. The optical emission in the biexciton to neutral exciton cascade has been proposed as a deterministic polarization entangled photon sourceAmong the possibilities offered by SAQDs engineering, the QD confinement potential can be controlled by the design parameters and the growth conditions, leading to a direct control of electronic states and optical properties, as for example to redshift the photon emission towards longer wavelengths (1300 or 1550 nm). However, in some cases the change in the design of the QD based structure and the growth techniques necessary to achieve such a wavelength tuning can deteriorate the optical quality: reduction of coherence times, increase of the homogeneous linewidth and spectral diffusion effects by the presence of fluctuating charges in the QD surroundings. These factors will invariably induce a reduction of fidelity and photon indistinguishability. At the same time, Fine Structure Splitting (FSS) from electron and hole exchange interaction usually increases with the QD sizeFrom a technological point of view, conventional Si detection technology is not useful for telecom wavelengths, as the detector efficiency drops dramatically beyond 1000 nm and InGaAs photodetectors are routinely used in photonic devices instead those based on Si, despite their higher noise characteristics. The use of those infrared detectors introduces a strong limitation in the experimental determination of single photon emission, particularly when the Signal to Noise Ratio (SNR) decreases towards unity, even if the detection scheme is enhanced by use of either gated InGaAs Avalanche Photodiodes (APDs) or changing InGaAs by superconductor based photodetectors212In this letter, we propose a new all-optical fiber Hanbury Brown & Twiss (HBT) interferometer containing a tunable Fiber Bragg Grating (FBG) optical filtering stage coupled to Geiger mode InGaAs Avalanche Photodiodes (APDs) for single photon detection . The useTypical μ-PL spectra of our large size SAQDs are composed by a relatively high number of optical transitions, which originate from the multiple carrier configurations through the different QD levels. The number of available confined states increases as the QD becomes wider and hence charge carriers can recombine through multiple state configurations. In order to identify and label all these transitions, we performed power excitation, time resolved and polarization resolved μ-PL experiments. 0; one electron and one hole in conduction and valence band S-levels, repectively, i.e. [1Se1Sh]), with a slope of 0.98. At an emission energy higher than X0, i.e. with negative binding energy, we identify a positive Trion  and a doubly positive charged exciton, a positive Quarton . X+2 recombination is composed by single unpolarized transition (XUn+2) at a slight lower energy than X+1, and two doublets at lower energy (XD1+2 and XD2+2), as it has usually found in literature17+1 and X+2 transitions are larger than the one of X0 (+2 transition (XD2+2) appears at −3.3 meV with respect to the unpolarized recombination0, and hence with positive binding energy, we identified the neutral biexciton emission , with slope ≈1.68, and a single positive charged biexciton , with slope ≈1.9. XX+1 recombination is produced between electrons and holes in S-levels (Se-Sh) populating the hot positive trion state . As recently demonstrated in refs +1* transition is composed by singlet and triplet states, whose degeneracy is broken by the hole-hole exchange interaction, whereas the triplet degeneracy is broken by electron and hole exchange interaction. The complete description of the X+1* state diagram, containing all exchange effects is presented in the +1* triplet state emission is characterized by a slope ≈1.5 for the power dependence of the μ-PL integrated intensity that is smaller than that of XX+1, but slightly larger than the one of X+1. Finally, we observe a biexcitonic transition with a slope = 1.9, matching the low energy sideband of the X0 transition. We were not able to extract enough information to propose a clear excitonic assignation, although here we tentatively assign to the singlet state transition of the hot positive trion X+1*.The integrated intensity versus power excitation Inset in  providesne of X0 , as extrOn the basis of our previous experience in the study of selectively charged quantum dots2325+1T, XX0, X−1, X0, XUn+2 and X+1 optical transitions, but not for the other transitions, as their μ-PL signal intensity under pulsed excitation was below the noise level. 0 transition, that coincides with the decay time for XX0 (0.73 ns), thus indicating that the recombination of X0 must wait for the radiative decay of XX0. These data reinforce our previous excitonic labeling. As we found in previous μ-TRPL measurements in a similar SAQD28+1* and XX+1 transitions, which will be discussed below. Within our experimental spectral resolution (≈46 μeV), we do not observe any significant FSS for X0 and XX0 optical transitions. The mean linewidth measured in our SAQD is about 100 μeV, which is slightly larger than those measured in standard SAQDs emitting at the first telecom window  are expected to exhibit a FSS of the same nature as that observed for X0, but it is characterized by a larger magnitude  and dark counts (Ni) from detection channel i (Start/Stop), ω is the time bin of the correlation board (190 ps) and T is the total integration time (11 hours). We fitted the normalized photon coincidence plot using the expression of the second order correlation function of a two level system, (g2(τ)), convolved by a Gaussian function representing the detector time resolution:where c is the FWHM of the instrumental response (400 ps), τd is decay time of the transition and n is the average number of photons in the photon stream. Following this procedure our fitting routine returns g2(0) = 0.30 ± 0.12. When the photon correlation is measured with small SNR it must be considered the noise effect from dark count coincidences. In our experiment we found where τ1 = 0.99 and ρ2 = 0.69. Under these detector conditions, an ideal single photon emission, (g2(0) = 0), would produce dark count correlations of CN(0) = 0.29 ± 0.04, which coincides with the experimental value extracted above by the fitting procedure. Therefore, we conclude that the X0 transition of our SAQD is compatible with g2(0) = 0.01 ± 0.16 after noise subtraction, which produces n = 1.01 ± 0.16 average photons.where 0 transition under pulsed excitation at 780 nm. The process to obtain the information for g2(0) is similar to that developed above for CW excitation, except that under pulsed excitation the photon correlation counts are distributed periodically along the delay time axis and separated by the laser period. To normalize the coincidence counts now we use the expression:m is an integer value corresponding to the pulse number. C(m) was obtained performing a multi Lorentzian peak fitting to calculate the integrated area of every m-peak, iH are defined as before, θ is the laser repetition period (13 ns) and T is the integration time (8 hours). After this normalization we obtain CN(0) = 0.44 ± 0.05. This result compares well with the theoretical value CN(0) = 0.51 ± 0.04, which is deduced by considering the effect of the dark counts correlation assuming an ideal situation (g2(0) = 0). Again, the g2(0) value for the X0 transition after noise subtraction is close to the ideal case.where +1*). Contrarily, we did not observe any measurable FSS related to neutral exciton (X0) and positive quartons (X+2). Such a small FSS for X0 is compatible with the nominal low lattice mismatch between these InAs/InGaAs SAQDs and the confining layers (6.34%), which has been proved to cause a decrease of the FSS32We have presented a complete optical study of a single metamorphic InAs SAQD by means of a novel all-optical fiber intensity interferometer containing a FBG-filtering stage coupled to Geiger mode InGaAs APDs. We analyzed and labeled the most relevant excitonic and biexcitonic transitions and identified a FSS ≈160 μeV associated to the single positive charged hot-trion state (X0 transition at the second telecom window (≈1300 nm) has been characterized, showing that the g2(0) value after noise subtraction analysis is close to the ideal case. This represents the first time where single photon emission at 1300 nm has been demonstrated by using InGaAs avalanche photodetectors under Geiger mode operation, which was possible by using a FBG filtering device integrated into the signal collecting optical fiber. The use of FBGs in conjunction with APDs in Geiger mode at 1300 nm is a novelty that offers a relatively simple and compact experimental architecture to study photon indistinguishability and coherence times. As an example of application, it has been recently found new and rich features in the simulation and measurement of frequency filtered N-photon correlation coincidences39Single photon emission of the XIn summary, our All-Optical Fiber proposal resembles the conventional procedure used in the analysis of SAQDs emitting below 1000 nm, where Si APDs photodetectors do not exhibit any SNR limitation and are used in the conventional Geiger mode. Our strategy represents a simpler, low cost and efficient tool to connect single/entangled photons into photonic architectures for telecommunications, as for example to develop new N-photon correlation experiments or more compact optical fiber based single and entangled photon devices. Single metamorphic QDs studied with our all-optical fiber based experiment shows single photon emission at 1300 nm with no evidence of neutral exciton Fine Structure Splitting, and hence represents a viable structure design to develop the more interesting deterministic entangled photon source at 1300–1550 nm telecommunication wavelengths.0.15Ga0.85As layer of 500 nm was deposited, followed by 5 nm of GaAs. Then a sub-critical coverage of 1.5 ML of InAs was deposited at 490 °C followed by 20 s annealing at the same temperature under As flux; SAQDs were finally capped by a 20 nm thick layer of In0.15Ga0.85As. Photon emission from these single SAQDs in the long wavelength range has been previously demonstrated168 cm−2)0.15Ga0.85As metamorphic layers the diameter of the QDs (dQD) has been estimated by AFM to be in the 30–45-nm range45In How to cite this article: Muñoz-Matutano, G. et al. All-optical fiber Hanbury Brown & Twiss interferometer to study 1300 nm single photon emission of a metamorphic InAs Quantum Dot. Sci. Rep.6, 27214; doi: 10.1038/srep27214 (2016).