Ensuring manufacturing reliability is key to satisfying product orders when production plants are subject to disruptions. Reliability of a supply network is closely related to the redundancy of products as production in disrupted plants can be replaced by alternative plants. However the benefits of incorporating redundancy must be balanced against the costs of doing so. Models in literature are highly case specific and do not consider complex network structures and redundant distributions of products over suppliers, that are evident in empirical literature. In this paper we first develop a simple generic measure for evaluating the reliability of a network of plants in a given product-plant configuration. Second, we frame the problem as a multi-objective evolutionary optimisation model to show that such a measure can be used to optimise the cost-reliability trade off. The model has been applied to a producer’s automotive light and lamp production network using three popular genetic algorithms designed for multi-objective problems, namely, NSGA2, SPEA2 and PAES. Using the model in conjunction with genetic algorithms we were able to find trade off solutions successfully. NSGA2 has achieved the best results in terms of Pareto front spread. Algorithms differed considerably in their performance, meaning that the choice of algorithm has significant impact in the resulting search space exploration. One of the major challenges in production planning is delivering orders to customers reliably whilst also minimising the costs involved in setting up and running the network of production across multiple-plants  with a set of Nodes N and Links L between the nodes and study topological connectivity after potential disruption events. As many real life networks have heterogeneous distribution of connectivity, removal of certain nodes or links would result in greater damage to the network than others. Global measures that assess vulnerability include operational pairs  – if this supplier fails, the assembly cannot be made. An alternative configuration might include the OEM procuring directly from Suppliers 2 and 3, both of which procure from Supplier 4. In this configuration Product 4 is multi-sourced to two suppliers in order to incorporate redundancy in the design. Suppliers are assigned multiple products. This design seems to be more reliable, but is it the most reliable and cost effective? A systematic method that considers trade-offs in both the cost of the configuration, and the reliability of alternative configurations needs to be developed. In this section, we first create a measure that evaluates the reliability of the design (Section 3.1) and next formulate an optimisation problem that systematically balances costs against reliability (Section 3.2).A graphical depiction of the reliable supply network design problem is illustrated in Fig. L, whereConsider that an assembler needs to procure a set of Products m is the number of plants. Further, the products provided by each Plant can be denoted by matrix F, where:n is the number of products.r.If an assembler can access to a plant through a set of links, then that manufacturer has a “path” to the products of that plant. The products in facilities that supply directly to the assembler have a path length of 1 to the assembler. These plants might procure products from other plants, creating path lengths of 2; who have a direct link to a plant of path length 1, but no direct link to the assembler. Thus, let us call the number of links the assembler needs to traverse to access a certain product, path length r, Nr) is the reliability score of the link between i and j.An adjusted matrix The adjusted r. For example we may choose to consider the reach to products from path lengths of 1 to 2, with a decrease in the significance of path length with weights 1 to 0.5.Here, each column sum would denote the reliability adjusted access level of each plant to all products in the network. Since we are interested in the assembler’s access to all products, the first column of F and the first row of F is 0 as the assembler does not produce products but only procures them. In the case that the assembler also produces products, we represent the assembler by a supplying plant, and add a dummy assembler in row 0 with a single link to the actual assembler whose reliability is 1.Note that the assembler is also represented in L incurs a cost of procurement, denoted by a matrix M. Then.K. The production cost is an aggregate value that the decision maker can model based on manufacturing variables such as labour, raw material and operational costs, and holding costs. ThenSuppose now that each link of L with minimal cost can be laid out as a bi-objective optimisation problem:The problem of finding the maximally reliable network configuration L must be 0:Three constraints need to be designed. First of these is that plants cannot supply to themselves. Hence the diagonal of the Secondly, the network must be a connected network i.e. every plant must have at least one link to either another plant or the assembler.L is positive  is the second smallest eigenvalue.The second constraint thus is:The final constraint is that every supplier must produce at least one product:To summarise, the optimisation problem is:Subject to:The formulation presented in the previous section constitutes a multi-objective network optimisation problem, which is not well handled in network literature Newman . HeuristL and F inherent to the problem.We opt for the use of GA as this class of algorithms are reported to handle unconventional search spaces well. Additionally, at each iteration the GA returns a population of solutions rather than an individual, which would help reduce the computational cost of the problem. Furthermore, the bit string representation used in GA can be leveraged for the binary matrices Chromosome: GA operate by defining an initial population of candidate solutions each of which is called an individual. Individuals are encoded as a digital “chromosome” that represents the variables of the optimisation problem to be solved. Each bit is called a gene. Several genetic encoding styles are possible. In our case we opt for binary string representation as this would handle the adjacency matrix and the binary product-plant matrix naturally. The first part of the bit string is used for L whereas the last part is used for the F. As L is symmetric and has a diagonal of 0, only n(n-1) bits are used, where n is the number of plants. As F the first row of F is 0, m(n-1) bits are used where m is the number of products.Fitness function: The fitness function is essentially the objective function or functions used to assess the “fitness” of the chromosomes. These in our case are α andC. Fitter chromosomes have a higher probability to pass their genes onto the next generation.Selection: Selection is the mechanism by which fitter chromosomes are given a bias to pass on their genes to the next generation. We use the Binary Tournament selection operator. Because we have two objective functions, our problem constitutes a multi-objective optimisation problem. In multi-objective optimisation there is no single optimum solution, but a set of Pareto optimal solutions. Pareto optimal solutions are trade-offs between different objectives and are also called non-dominated solutions, meaning that there is no other solution which would improve an objective without causing a worsening in at least one of the other objectives  over half the bits that differ between the two parent chromosomes. This is done by first finding the bit positions that differ between the two parents and then randomly selecting a differing position to swap the bits. The process is repeated until half of the differing bits have been swapped.Mutation: The mutation operator produces random changes in the chromosome. The operator is used to explore previously unexplored search space and helps diversify the population, preventing premature convergence. We use the BitFlip mutation, which takes the chosen chromosome and inverts the bits. In BitFlip mutation rather than selecting a single bit to mutate, the operator finds two random points in the string and reverses the order of the bits between those points.The GA formulation of the problem includes 5 parts Fig. :Chromosomax to 25,000 and the population size to 100 achieved a good balance between solution quality and computational efficiency. Figure Experiments were carried out on an Armari Magnetar, 3.5 GHz, 64 GB RAM Workstation. Computational trials indicated that setting the total number of generations NThe approach is tested with an automotive light and lamp producer’s production network : This metric represents the fraction of the objective space that is dominated by the obtained solutions, indicating the coverage of the real Pareto front. The larger the hypervolume, the better the optimiser : This metric calculates the proximity of the solutions obtained to the population of the real Pareto front. The smaller the GD, the better the optimisation result (Veldhuizen and Lamont Spread (S): This indicator estimates the diversity of the set of solutions obtained on the Pareto front Deb . A smallFurther tests were conducted on progressively larger problem instances Table . While PWe also report the CPU time for finding best solutions. Due to the probabilistic nature of GA, algorithms were run 30 times for each problem instance. Expectedly, as problem size increases so does CPU time, GD, and S, while HV decreases. PAES performs consistently worse than NSGA2 and SPEA2 in all metrics, while NSGA2 outperforms SPEA2 in HV and CPU time, albeit slightly. It appears that NSGA2 maintains its performance across the three problem instances, dealing with the product-plant configuration problem better than the other two algorithms. The different performances of these algorithms highlight the fact that the optimisation algorithm choice has an important role in achieving a good trade off solutions in the reliable supply network design problem formulated in this paper.Although supply chain design literature offers many bottom up models for plant-product configuration, the reliability of resulting designs have received little attention. Bottom up models require detailed models of material flow, and thus are complex to build and solve, and difficult to generalise beyond the case they are designed for. Furthermore, supply chain designs typically focus on chain like structures and do not take network formations into account, despite a growing number of empirical work suggesting that real life supply chains contain network structures.In this paper we developed a top down model for evaluating the reliability of production over a network of plants in a given supply chain configuration. We assumed that the supply chain is reconfigurable, hence the problem may be viewed as a product- plant network configuration problem. The model is generic as it works with minimum information, and can handle network structures. We then used the model as a basis to frame the configuration problem as a multi- objective optimisation problem that balances cost against reliability. We opted for the use of genetic algorithms because of their ability to handle unknown search spaces at reasonable computational speed. Of the three algorithms NSGA2 has achieved the best results in terms of Pareto front spread. Algorithms differed considerably in their performance, meaning that the choice of algorithm has significant impact in the resulting search space exploration.Several assumptions underlie the problem formulation presented in this work; each of which present an opportunity for further improvement to the model.Firstly, we assume that the reliability of links between suppliers can be a priori estimated and that this estimation is accurate. Thus the application of the developed model would benefit from a structured decision process that enables a manufacturer to carry out such estimations such as the incorporation of historical data analysis.Secondly, perfectly substitutable goods are assumed, although in real life supplier offerings may differ in their quality and therefore products may not be perfectly substitutable. Remedying this assumption could be done with additional costs to represent the substitutability of the products.In addition, it is assumed that the reliability of a supplier is the same for each product it offers although different production constraints may result in different reliability scores for each product. In this case the model could be further developed with multiple links to products at each supplier.The assembler is assumed to have no cost for accessing to alternative sources of products although buying from an alternative supplier or production plant could mean additional transaction costs. These could be easily incorporated into the model.As our aim has been to create a generic model we deliberately ignored bottom up details such as material quantities and associated cost models. These could be built in as extensions to the model specific to the case study problem being addressed. Finally the formulation does not allow precedence constraints; meaning that the sequence with which resources must arrive at the assembler is not incorporated in the model.Although the model has been designed with product-plant network configuration in mind, it also fills a gap in network science literature where a focal node’s ability to access resources distributed over a network is assessed.L ∈ Rm × m – binary matrix representing connections between production plants.F ∈ Rn × m – binary matrix that describes what products are produced by each plant.r between plants i and j in the network L.r between plant j and product kRel(Lij) – reliability score of the link between plants i and j in the network LL that takes into consideration the reliability of linksα – objective function representing the reliability of the manufacturer in accessing all products in the networkr – weight in the objective function α that corresponds to the path of length rwM – matrix representing the cost of procurement for each link between any two plantsN – matrix that describes the cost of each link in the network L* – element-by-element multiplicationK – matrix representing the cost of producing each product in each plantG – matrix that describes the cost of production in the network LC – objective function corresponding to the total cost of the network configuration LL in a symmetrical unidirectional formi (degree of node) in the network D – matrix describing the degree of each node in the network λ2(·) – second smallest eigenvalue of a matrix


Men suffering from azoospermia can father a child, by extracting spermatozoa from a testicular biopsy sample. The main complication in this procedure is the presence of an abundance of erythrocytes. Currently, the isolation of the few spermatozoa from the sample is manually performed due to ineffectiveness of filtering methods, making it time consuming and labor intensive. The spermatozoa are smaller in both width and height than any other cell type found in the sample, with a very small difference compared with the erythrocyte for the smallest, making this not the feature to base the extraction on. However, the length of the spermatozoon is 5× larger than the diameter of an erythrocyte and can be utilized. Here we propose a microfluidic chip, in which the tumbling behavior of spermatozoa in pinched flow fractionation is utilized to separate them from the erythrocytes. We show that we can extract 95% of the spermatozoa from a sample containing 2.5% spermatozoa, while removing around 90% of the erythrocytes. By adjusting the flow rates, we are able to increase the collection efficiency while slightly sacrificing the purity, tuning the solution for the available sample in the clinic. To assist men lacking spermatozoa (sperm cells) in the ejaculate  in fathering a child, a method has been developed to separate spermatozoa from erythrocytes (red blood cells). With azoospermia, a biopsy is performed to extract spermatozoa, which are used for intracytoplasmic sperm injection (a sperm is directly injected into the egg). The main complication with this procedure has been the abundance of erythrocytes, which have to be manually separated owing to the ineffectiveness of filtering methods. A team headed by Loes I. Segerink at the University of Twente, the Netherlands devised a technique for separating the two cell types using the tumbling behavior of spermatozoa, which is based on their length being about five times greater than the diameter of erythrocytes. The authors were able to extract 95% of spermatozoa while removing around 90% of erythrocytes. If the sperm count is very low, intracytoplasmic sperm injection (ICSI) might be necessary. In more rare cases , there are no spermatozoa in the ejaculate (azoospermia), making it necessary to perform a biopsy for obtaining spermatozoa2. This method is called testicular sperm extraction (TESE), after which the spermatozoa are extracted from the biopsy sample and used in ICSI2. Azoospermia can be split into two classes: obstructive and non-obstructive. In the first class, the cause for the lack of spermatozoa in the ejaculate is due to an obstruction in the reproductive tubes that prevents the spermatozoa from ending up in the ejaculate. In the second class, there is no obstruction, but there is a problem with the production of spermatozoa. This means that even in a biopsy sample, the sperm count is very low.Assistive Reproductive Technology is used all over the world to help couples with fertility problems conceive. It is estimated that 1 in 20 males has a low sperm count2. Moreover, the selected spermatozoa can differ in quality, thereby reducing fertilization and pregnancy rates. Some techniques have been introduced to better select spermatozoa for ICSI, such as the hyaluronic acid-binding assay3 and intracytoplasmic morphologically selected sperm injection4. Both techniques are not fully validated and therefore their value for improving the outcome of ICSI is doubtful. The development of a technique that reduces TESE sperm processing time in combination with sperm quality selection is our ultimate goal, which will lead to a next step in more efficient and effective fertility care.During a TESE, samples that are obtained from a patient with non-obstructive azoospermia contain only a very small number of spermatozoa, submerged in a mixture full of other tissue cells, other compounds, and a large amount of blood cells that are introduced due to blood vessel damage. The largest tissue cells and leukocytes are currently filtered out, e.g., by density gradient centrifugation, after which the remaining sample, containing mainly erythrocytes and spermatozoa, is imaged under a microscope. The sample, typically containing very few spermatozoa (<1%), is then searched for spermatozoa, which is a time-consuming manual process and can take many hours depending on the available numbers of suitable spermatozoa and oocytes7 for a better quality selection in single sperm for ICSI. Although this research is still running, we started the second part: more efficiency in TESE work-up. In first instance, we are focusing on the separation of spermatozoa from erythrocytes. Finally, we will bring these two parts (the faster TESE work-up and the single sperm selection) together in developing a microfluidic chip that automatically selects the highest quality spermatozoa for ICSI.Until now, our focus was on individual trapping of sperm for analysis using chip technology8 or result in the loss of a significant percentage of the initial sample or low purity (cross flow filters)9. As the spermatozoa in a testicular biopsy might be not developed enough to showcase swimming, we cannot rely on this phenomenon for separation, requiring an approach that uses an imposed flow. Son et al.10 reported being able to separate spermatozoa from erythrocytes in their work, where they used a spiral channel to separate spermatozoa. In their reported results, one can see that the spermatozoa are broadly distributed in the channel. Their method could not get high purity of the spermatozoa due to this effect without using multiple steps and sacrificing retention. Liu et al.11 used a pinch flow device to separate epithelial cells from spermatozoa for forensic purposes. They noticed poor focusing of the spermatozoa in the channel, but due to the large difference in size between epithelial cells (50 µm diameter) and spermatozoa , the results were good enough for this application. Their sample contained 30% spermatozoa and their final purity was 94.0 ± 4.7%, with a retention rate of 41.1 ± 2.9%. The poor focusing of spermatozoa in the pinch flow device is a problem for the separation of particles that are slightly bigger than spermatozoa, but we think this phenomenon can be used for the separation spermatozoa from of smaller particles. Therefore, we propose to use pinch flow fractionation (PFF) for the separation of spermatozoa from erythrocytes.The large amount of cells in the sample generated by the TESE procedure requires a microfluidic separation method that can tolerate a large sample throughput (billions of cells of very different sizes from ~2 to ~40 μm) without clogging. The standard filter types either show a lot of clogging (dead end filters)12. In this work, boar spermatozoa are used as a model for human spermatozoa and these are 45 μm long, 4 μm wide at the head, and have a height of 1 μm13. In comparison, human spermatozoa are 55 μm long, 3 μm wide at the head, and have a height of 1 μm14. This means that the cell sizes in a testicular biopsy, especially that of erythrocytes, are comparable with that of a spermatozoon and therefore the long shape of the spermatozoon needs to be utilized. It is mentioned by Samuel et al.15, in their review on the possible usage of microfluidics for TESEs, that PFF is not ideal for the separation of particles of multiple sizes when compared with other techniques, although the non-focusing of the spermatozoa, caused by their shape, can offer us an advantage when using PFF. In our application, a PFF device allows for the retention of most particles, while preventing clogging, as it uses hydrodynamic methods to sort the particles instead of steric hindrance.The dimension of erythrocytes are typically 7.5–8.7 μm and 1.7–2.2 μm in diameter and height, respectivelyIn this study we show that PFF can used to retrieve the few present spermatozoa from a testicular sample, by making use of the typical shape of spermatozoa.The observed behavior of the beads in the microfluidic chip showed good agreement with our model in Comsol of beads traveling through an identical geometry. The observed behavior of spermatozoa however deviates from that of spherical beads in three noteworthy ways. First, the spermatozoa appeared in a broadened section with, on average, a larger distance from the wall than expected from their width 1–3 µm). Second, the distribution of the spermatozoa in the broadened section is also considerably wider than that of round particles  and, therefore, also showcase tumbling behavior in a shear flow. Due to the large aspect ratio, the spermatozoa will mostly be oriented in their equilibrium orientation, which is aligned with the flow, either backwards or forwards depending on the vertical position in the channel21. However, in the PFF device, they are forced out of their equilibrium orientation due to the compressing and widening of the streamlines, inducing a tumble in the transition from the pinched section to the broadened section . The high shear then causes them to tumble20. The second form of tumbling is mostly in the x–y plane of the device when the streamlines broaden after the pinch. The latter has an influence on the final separation process in the chip. This rotation of the spermatozoa happens at a critical time point in the device, when the streamlines are diverging and carrying particles of different sizes along their respective streamlines. PFF fractionation works with the principle that particles are forced into a specific streamline at the pinched section based on their size and will follow this line when the streamlines diverge, separating particles of different sizes. However, as the tail of a spermatozoon is deformable, but not so flexible that it can completely follow the streamline the head is occupying, the effective diameter of the spermatozoa will be larger. For the spermatozoon visible in Fig. Two forms of tumbling have been seen in the PFF device. The first form occurs in the This larger effective diameter due to rotation makes the separation of the spermatozoa from larger cells more difficult. However, it is an advantage for the separation from cells that have equal smallest dimensions, but do not have such a large aspect ratio. Normally, a filter cannot be used for these separations, as a filter is based on the smallest dimension of a particle. Although erythrocytes also show tumbling behavior, the dispersion of these cells is less, as the larger axis of an erythrocyte is approximately one-fifth of that of a spermatozoon and therefore they will appear closer to the wall of the device. This tumbling phenomenon in the PFF device can therefore be used to separate spermatozoa from the abundant erythrocytes that reside in a typical biopsy sample.We have shown that due to the tumbling behavior of spermatozoa, these cells end up at a different position in the broadened channel than erythrocytes. The next step is to show that we are also able to isolate the spermatozoa from the sample.n = 3) of the spermatozoa are viable after our separation and we can improve this by shorting the tubing, as this causes most of the damage to the cells. In our experiments, we have also discounted the presence of leukocytes. These make up a small percentage of the sample (<0.2%) and might have been in our product outlet, which would reduce the purity ratio. For simplicity in our experiments, we have not classed these as a separate species from erythrocytes. For comparison with the results from Liu et al.11 who has a CE of 41.1 ± 2.9% and an EP of 97.0 ± 2.3% for separation of spermatozoa from epithelial cells of 50 µm, we have obtained higher CEs, but lower EPs, with a CE higher than 52 ± 0.3% for an EP of 81 ± 8%, and up to over 94 ± 8% for an EP of 31 ± 9%. This is due to the different application, as we are not looking for identification of a subject but for the harvesting of spermatozoa for fertilization. Next to this, Liu et al.11 separated spermatozoa from epithelial cells that were 50 µm in diameter, whereas we separate spermatozoa from erythrocytes, which have a very similar smallest dimension.A whole blood sample, spiked with 2.5% spermatozoa, is made to mimic the composition of a testicular biopsy after gradient filtration. The 2.5% in the sample has been chosen to have enough spermatozoa to be able to do statistics on the results. This sample is running through the system, while the percentage of flow to the waste outlet P3) is varied between 3% and 5%. If 3% of the total liquid is removed from the sample after the expansion, most spermatozoa are retained, but also a considerable percentage of the erythrocytes is kept in the collected sample. When larger percentages are removed, the amount of erythrocytes that are not separated out goes down dramatically. However, this also causes a larger percentage of spermatozoa to be removed from the sample  or less spermatozoa in a purer sample. This is dependent on the wishes of the clinician, who decides on an estimate of the amount of spermatozoa that are available in the total sample. This estimate is made during diagnosis of the severity of the condition and is known before the biopsy sample is processed. With this estimate, it is possible to choose the percentage of liquid to be directed toward the waste outlet (adjusting the pressure setting of the pump according to our script), with more liquid to be removed if one wants a purer sample and less removal of liquid if one wishes for a large amount of the spermatozoa to be retained.Due to the tumbling behavior of spermatozoa in a pinched flow, we are able to retrieve the spermatozoa from the erythrocytes in a mock sample of a testicular biopsy. By adjusting the flow rates, a high CE can be achieved for samples with very few spermatozoa (<20). We are able to obtain 95% of the spermatozoa, while removing ~90% of the erythrocytes. In samples with more spermatozoa, a high CE can also be sacrificed for a higher purity. They can choose to collect 85% of the spermatozoa, while removing 99% of the red blood cells to make the final selection of spermatozoa easier for the final application.The microfluidic chip consists of a PFF design, containing two inlets width: 50 µm): one for the sample and one for the buffer joining into a single channel (the pinched section) that is 50 µm wide and 125 µm long . The erythrocytes were counted in the blood samples (~5 × 109 cells ml−1) and the sample was mixed with spermatozoa containing solution to end up with 2.5% of spermatozoa in the sample. This concentration is slightly higher than in clinical samples, to obtain a better fidelity for the analysis. The sample was then diluted five times for handling with Beltsville Thawing Solution .To mimic a pre-filtered biopsy sample, as typically found in the clinic, we use a sample that consists of whole blood, spiked with boar spermatozoa, which is diluted five times. Fresh boar semen was obtained from a local artificial insemination center  at a concentration of 20 × 10−1 (0.17 for the sample flow) and the total separation time is ~15–20 min for a sample of 200 µl. Experiments were done with 6 and 15 µm beads , erythrocytes and boar spermatozoa. The erythrocytes and spermatozoa were imaged and counted for the fraction totals on a Nikon TE2000-U microscope  equipped with a ×10 phase contrast objective. A high-speed camera  was mounted onto the microscope for image recording with the Photron software (Photron Fastcam Viewer). This high-speed camera imaged the cells passing through the transition from pinched to broadened section. Prior to each experiment, the chip was activated with oxygen plasma to prevent bubble formation during filling. The PDMS and glass surfaces of the chip were coated with poly(l-lysine)-grafted-poly(ethylene glycol)  to prevent bead and cell adhesion during the separation experiments. PLL-g-PEG was rinsed through the PDMS micro channels at a concentration of 100 μg ml−1 in deionized (DI) water for at least 15 min. All experiments were carried out at room temperature to prevent swimming of the boar spermatozoa.Two sets of experiments are carried out. The first set is to characterize the behavior of spermatozoa and erythrocytes after PFF. The second set is to show the efficiency of spermatozoa isolation from a sample containing erythrocytes. For all these experiments, the same chip geometry is used, while changing the outflow ratios. The chip was connected to a Fluigent pump  using four channels and four equal length in- and outlet tubing. The fluidic resistances of the channels in the chip were calculated and measured with stagnation pressures. The in- and outlet pressures (P1–4) were then adjusted for a flow rate ratio of 1:20  for the inlets and 3/3.5/4/4.5/5:100  for the outlets. For this we used a Matlab script , which translated the wanted flow percentages to input pressures for the pressure pump. The total flow rate is 3.3 µl sThe influence of the chip on the viability of the spermatozoa was assessed with a SYBR 14/propidium iodide (PI) live/dead staining. The spermatozoa were incubated in a 1000× dilution of SYBR 14  for 20 min and a 100× dilution of PI  for 5 min at room temperature. To obtain the influence of the chip on the viability of the cells, the ratio of live/dead spermatozoa in the treated sample was divided by the ration of live/dead spermatozoa of the control sample.A first set of experiments was done with 6 and 15 µm beads and boar spermatozoa, to characterize the particle distribution in the separation channel. A mixture of microspheres and spermatozoa was prepared and flown through the device to characterize the particle distribution in the channel. From this distribution, the preferred outlet flow rates were estimated and used in the erythrocytes/spermatozoa experiments. The particle distribution of erythrocytes was also characterized. This was done with a Matlab script, which counted and circled the beads/erythrocytes, after which the spermatozoa (and any beads/erythrocytes that were missed) were counted by hand. The positions relative to the channel wall were saved and plotted in a histogram.Extraction purity (EP): the proportion of spermatozoa in the collection outlet, relative to the total number of cells extracted in this outlet. Therefore, EP reports the composition of the extracted sample.Collection efficiency (CE): the proportion of spermatozoa or erythrocytes in the collection outlet, relative to the total number of that cell type in the sample. This gives an indication of the retention of the spermatozoa and the percentage of erythrocytes that cannot be separated in that same experiment.Enrichment ratio (ER): the proportion of spermatozoa in the extraction outlet compared with the proportion at the inlet.After determining the preferred flow rates for the spermatozoa and erythrocytes experiments, the separation of spermatozoa from the sample was characterized. For this, we defined three parameters.The flow ratio of buffer to sample in the inlet was 95:5 for each experiment, whereas the ratio of waste flow/total flow in the outlet was varied between 3% and 5%. To obtain the EP and CE, the spermatozoa and erythrocytes were counted in the inlet and each outlet with the same script as mentioned above, supplemented by manual counting. The ER was then calculated from the sample composition before and after separation.Simulations have been performed with Comsol 5.1  to obtain simulation data for the position of 6 and 15 µm beads. The model uses fluid structure interaction in the pinched section, where the size of the bead (6 or 15 µm) is significant compared with the channel dimension (50 µm) to consider the effect that the particle has on the flow profile. Due to the large computation time of this method, the particle position after passing the pinched section was taken and further modeled with particle tracing to obtain the position over time in the broadened section. Particle tracing models the particle as just one point in the channel, with no major effects on the flow profile, which is now allowed due to the small size of the particle relative to the channel.Video S1: High speed video of spermatozoa in pinch flow fractionation deviceVideo S2Video S3Supplemental Material


Experimental observations, such as the bias polarity dependence of adatom removal and the observation of an intermediate state, indicate that the adatom penetrates the surface oxide layer. Through the comparison of finite element method simulations with various experimental relationships, the electric field is concluded to be the sole contributor to adatom penetration into the surface oxide layer. The energetic barrier to this process is estimated to be approximately 0.45 eV in magnitude. Furthermore, the resolution of this phenomenon is on the atomic scale: individual adatoms can undergo surface penetration whilst their nearest neighbour adatoms, separated by 5 Å, are unaffected. The mechanism reported here has the advantages of not strongly influencing the substrate and is exceptionally localised, which can be beneficial for the synthesis of single atom devices.Oxygen adatoms on the MoO Its compounds are utilized in a wide variety of applications such as fertilisers3, nanowires4 and catalysts5. Molybdenum oxides are important catalysts, catalytic applications include hydrotreating of oil to reduce the sulphur, nitrogen and aromatics content, yielding cleaner more efficient fuels6. MoO2 is a conductor which has been reported to act as a catalyst in several different reactions including the dehydrogenation of alcohols7, the reformation of hydrocarbons9 such as biodiesel10 and is used for the partial oxidation of aviation fuel11. The catalytic activity shown of MoO2 is due to the transfer of nucleophilic oxygen ions from the oxygen sub-lattice to the catalytically active surface where they are then consumed to sustain the redox cycles12. The oxygen content is an important factor in catalytic activity. For example, the catalytic activity of MoOx in converting methanol to formaldehyde is determined by its oxygen coordination13: MoO2 is a poor catalyst in comparison to MoO3. The presence of Mo6+ is the critical factor in obtaining a high formaldehyde yield rate. The ability to control which MoOx species is present on the surface has important applications for oxidation and reduction reactions. MoO2 can also be used as a nanostructure template14.Molybdenum is a crucial element in many high-performance alloys and steels15. Adsorbates on metal and semiconductor surfaces can be manipulated using the electric field generated between the STM tip and the surface16, tunneling electrons17 from the tip to the surface or a combination of both. Electric field induced diffusion19, desorption20 and switching22 of atoms and molecules on metal and semiconductor surfaces have been previously observed using STM techniques, however, adatom penetration into the surface has yet to be realised. The vertical movement of an atom through a surface has important applications in the field of quantum computing: in order to manipulate and read out a quantum state, such as the nuclear spin of an atom, requires the precise placement of atoms within a lattice23. Current methods involve creating a high precision hydrogen mask on a Si surface using the STM tip and then dosing the system with PH3 and subsequently growing Si over-layers24. Other methods involve firing high energy (14 keV) P ions at a Si(100) substrate to implant them within the Si lattice26. Atom manipulation with the STM tip offers the high resolution required to precisely place an atom within a lattice, provided the tip can impart a sufficient force to result in the atom penetrating the surface when a voltage pulse is applied.Individual atom surface modification using the Scanning Tunneling Microscopy (STM) tip was first famously demonstrated by ‘writing’ “IBM” with Xe atoms on a Ni(110) surfacex2+/Mo(110) surface. In that work, oxygen adatoms were removed from the surface via pulsing with the STM tip27. Pulsing was carried out in the tunneling regime, where both tunneling electrons and the electric field between the tip and surface could contribute to adatom removal. In that regime, the oxygen adatoms were concluded to be removed from the surface via inelastic electron tunneling. However, we will demonstrate that is also possible to observe the removal of the oxygen adatoms when not in the tunneling regime. In the current work, we present results based on pulsing adatoms on the MoOx2+/Mo(110) surface outside of the tunneling regime, where the electric field is the only candidate mechanism for adatom removal. There are two possibilities for the observed removal of oxygen adatoms from the surface, either the adatom escapes from the surface to the vacuum via desorption or the adatom penetrates the surface. The desorption of atoms from a surface to a vacuum using an electric field is demonstrated in Field Ion Microscopy (FIM). Protruding atoms evaporate from the microscope tip in a process called field evaporation. Field evaporation can be employed to examine the binding energy of surface atoms28, typically in the range 2–5 V/Å29. Metals and their compounds can have a potential barrier to the incorporation of adsorbates such as oxygen into and beneath the surface layer. The magnitude of this barrier affects surface phenomena such as catalysis, corrosion, friction, the kinematics of oxidation and the transition from physisorption to oxidation states. If a barrier to oxygen penetration is present Cabrera-Mott theory describes the growth of a thin film oxide layer30. The transport of oxygen ions through an oxide film has applications in the field of memristors32. These circuit elements have the unique ability to vary their resistance based on the applied current.In a previous publication, we have demonstrated the ability to ‘write’ on the MoOx2+ surface outside of the tunneling regime and to understand why the structure of the oxide layer allows for this process to occur. We have studied the effect of applying an electric field to an oxygen adatom on the MoOx2+/Mo(110) surface with no influence from the tunneling current and we conclude that it is possible to induce the transition of an oxygen ion from the surface to an underlying oxide layer. Finite element method (FEM) simulations are employed to show that an electric field alone can replicate the experimental results. Inducing adatom penetration through a surface by means of an applied local electric field is a new mechanism to the best of our knowledge and this method has potential applications in the fabrication of single-spin devices such as qubits for quantum computing as well as in the field of single molecule engineering34, which strives to assemble atomic-scale structures from their individual atomic components using the STM tip.The purpose of this study is to understand the mechanism behind the removal of adatoms from the MoO2 forming on the surface. Oxidising the MoO2/Mo(110) sample at room temperature results in the formation of an oxygen-rich MoOx2+/Mo(110) surface. The additional oxygen takes the form of adatoms on top of the O-Mo-O trilayer structure. Applying a single voltage pulse (positive sample bias) with the STM tip when outside of the tunneling regime can lead to a number of oxygen adatoms being removed from the surface at once, as seen in Fig. 27 where larger scale images are presented. No tunneling current is present during a bias pulse due to applying a sufficiently large vertical offset between the tip and surface, further details relating to the pulsing conditions and the sample preparation can be found in the methods section. The diameter of the circular area from which oxygen adatoms are removed depends on the magnitude of the pulse voltage. Individual adatoms can also be removed even when neighbour adatoms were separated by just 5 Å. The high resolution of oxygen adatom removal is depicted in Fig. Annealing a single crystalline Mo(110) sample in an oxygen partial pressure results in the formation of a thin film of MoOAfter pulsing is performed the oxygen adatom is either removed from the surface or still in its original location on the surface, depending on the magnitude of the bias pulse. However, on occasion pulsing has been observed to change the relative height of the oxygen adatom as observed by STM. In Fig. The minimum voltage required to remove an adatom is referred to as the threshold voltage, which depends on conditions such as the tip-surface separation. The threshold voltage for the removal of oxygen adatoms can be measured in two ways. Firstly, by systematically increasing the bias voltage and monitoring the increase in the spot diameter, a non-linear dependence is obtained as indicated by the black circles in Fig. The red triangle in Fig. W. The magnitude of this barrier can be modified by applying an electric field. If the probability of an oxygen adatom being removed follows the relationship in Eq. (V vs ln(Δt) graph contains two terms. The temperature dependent term provides only a small contribution, as the intercept only changes by ~0.1 V for the measurements at 121 K and 81 K, and hence, this term is omitted. Therefore, the intercept provides an approximate value for W is be determined to be 0.45 eV.To penetrate the surface, an oxygen adatom needs to overcome a potential barrier, p in Eq. , measurip in Eq. , the int35 of the electric field generated between the tip and surface have been performed in order to shed light on the mechanism behind the removal of oxygen adatoms from the MoOx2+/Mo(110) surface. Figure FEM simulationsMoO2+x/Mo10 surfacx2+/Mo(110) surface can be realised without the influence of tunneling electrons.A reference threshold value of 3.3 V for an offset of 1 nm was taken from the threshold voltage vs. applied offset experiment, black circles in Fig. Oxygen adatom removal is only observed when a positive bias is applied to the sample. Given that the oxygen adatoms are negatively charged and that the electric field acts perpendicular to the surface due to its conductive nature, the net force on the adatoms acts perpendicular and towards the surface. Large surface area scans before and after pulsing indicate that the density of adatoms in the regions surrounding the pulsed areas does not change upon pulsing outside of the tunneling regime, indicating there is no lateral diffusion in this regime. Pulsing at high voltage/current in the tunneling regime can lead to lateral diffusion, likely due to local heating of the surface. When the polarity of the pulse voltage is reversed, no adatoms are observed to be removed, when outside of the tunneling regime. Oxygen adatoms have been observed to transition to an intermediate state before being fully removed from the surface. This state occurs when pulsing close to the threshold voltage and results in a reduction in the apparent height of the oxygen adatom in STM images, indicating that either, the oxygen adatom is now within the surface layer or has changed its electronic state, likely due to chemical bonding.37. The energy barrier for oxygen incorporation depends on the oxygen adatom distance to the underlying surface layer and the bond lengths of the atoms within the layer which the oxygen adatom will penetrate, and hence, the substrate electron density. A larger electron density results in a larger barrier. Several factors affect the electron density at the point where the adatom crosses into the substrate, such as the lattice constant, lattice structure and material. In transition metals and their oxides, a 10% increase in lattice constant reduces the barrier by almost 30%37. Surface relaxations thus play an important role in determining the substrate electron density. Several different crystal lattice structures have been studied37, the rocksalt structure is very homogeneous and close-packed, leading to high energy barriers. While in the rutile, bixbyite, and spinel structures there are “open channels”, which can be envisioned as vacancies, through which the oxygen can diffuse easily and thus have lower energy barriers for oxygen incorporation37. Introducing a defect in the third layer of NbO(111) reduces the energy barrier from 4 eV to 1.2 eV37. Furthermore, the presence of an oxide-overlayer can increase or decrease the barrier, depending on the induced modification of the metal lattice substrate. For example, in the case of Nb(110), the barrier increases from 1.2 eV to 7.4 eV after oxidation. Whereas the barrier for incorporation into the Cu and CuO2 lattices are 0.9 and 0.04 eV37, respectively.The penetration of oxygen atoms through metal and metal oxide surfaces has received attention from a theoretical viewpoint2 surface without any adatoms did not result in any oxygen atom transitioning below the surface. The relative ease  at which oxygen adatoms on the MoO2 surface can penetrate the surface is concluded to be due to how the adatoms modify the nearby MoO2 lattice. Density functional theory (DFT) calculations have previously been performed in order to understand the structure of the molybdenum dioxide layer38 and the over-oxidised MoOx2+ surface27. These calculations can shed light on the observation of the removal of oxygen adatoms via pulsing with the STM tip. The simulations, seen in Fig. 39. The activation energies for such ionic conductivity’s were found to be in the range of 0.7 to 1.6 eV40, which is of the same order of magnitude as the potential barrier height obtained in our experiments (0.45 eV).Pulsing of the MoOa, in the simulations was intuitively taken as the distance between an oxygen adatom and the topmost molybdenum layer, this assumes the potential barrier is a maximum in the plane of the molybdenum layer, referred to as the crossing point. However, in a non-uniform system, such as this one where the oxygen adatom deforms the topmost molybdenum lattice structure, the crossing point may not lie in the same plane as the topmost molybdenum atoms. Variations of charge distribution on the atomic scale can affect the spatial distribution of the electrostatic field generated by STM tip which can influence the potential barrier. Nevertheless, the simulation is in good agreement with the experiment, suggesting that our model, which is based solely on the electric field, is correct.The FEM model simulating the electric field effect yields convincing results despite its simplicity. Disparity between the FEM simulation and experimental results are suspected to be due to the differences in the simulation parameters such as the tip shape. Furthermore, the surface is neither perfectly flat or conducting, which would lead to a different potential energy being present in the vicinity of the surface. The value for the adatom-surface distance, 42 on resistive switching and filament formation in memristor structures. Memristor structures  are mostly based on doped oxide insulator films32. Memristor switching is usually initiated via oxygen vacancy diffusion induced by an electric field in an interface layer of 1–10 nm in thickness. In order to reveal the memristor like resistive switching of the MoO2 structure, we have performed experiments on a molybdenum single crystal with a silver point contact (see supplementary section). The MoO2 layer in this memristor experiment has oxygen vacancies which support the diffusion of oxygen ions through it. Resistive switching has been observed at bias voltages in the range of 0.5–1.5 V, which again supports the energy barrier previously obtained in our experiments.Local modification of oxide compounds via oxygen diffusion induced by an external electric field has been previously observed in numerous experimentsx2+/Mo(110) surface demonstrate high mobility. Pulsing the oxygen adatoms with the STM tip leads to the removal of the adatoms from the surface. This only occurs at positive sample bias and is observed in the absence of the tunneling current. The bias polarity dependence, the absence of lateral surface diffusion and the presence of an intermediate state indicates that the adatom penetrates the topmost Mo layer due to the electric field. FEM simulations of the electric field generated between tip and surface accurately describe the experimental results, indicating that the electric field is solely responsible for the observed removal of the oxygen adatom, which is pushed below the terminating layer. DFT calculations show that the oxygen adatom modifies the underlying Mo layer structure, which in turn opens up a “channel” in the Mo layer, which can reduce the energetic barrier for the adatom to transition to the oxygen layer below.The oxygen adatoms on the MoOx2+/Mo(110) was found to be approximately 0.45 eV. This localised mechanism allows for the implantation of atoms without strongly influencing the substrate.The atomic resolution capability of this method of oxygen adatom removal is due to the exponential dependence of the probability to transition on the applied bias voltage. The minimum spot diameter in which adatoms are removed from the surface is quite small, just 0.5 nm in diameter. The potential barrier for oxygen adatom incorporation into MoOin-situ annealing at 1275 K. The preparation chamber used in experiments has a base pressure of 5 × 10−11 mbar at 300 K. The sample was heated by electron beam bombardment and temperatures were measured using an optical pyrometer . The clean Mo(110) surface was verified by low-energy electron diffraction (LEED) and STM before oxidation. Subsequently, the sample was oxidised, using O2 gas with 99.998% purity, at 1250 K in an oxygen partial pressure of 1 × 10−7 mbar for 2 minutes and allowed to cool to room temperature, resulting in a clean MoO2/Mo(110) surface termination43. The MoO2 (010) surface, which is detailed in length by Radican et al.43 and depicted in Fig. 1 direction of the Mo(110) substrate. In order to obtain oxygen adatoms on the MoO2/Mo(110) surface, the sample was then exposed to an oxygen environment of 1 × 10−7 mbar for 2 minutes at room temperature. This results in the formation of an oxygen-rich MoOx2+/Mo(110) surface exhibiting oxygen adatoms27 which is depicted in Fig. x2+ surface and make up the vast majority of the observed features. They can be seen in greater detail in the images presented in Fig. x2+/Mo(110) surface, the MoO2/Mo(110) surface was exposed to different doses of oxygen and there is a strong link between the number of these small bright dots and the dose of oxygen which the surface was exposed to. Further details correlating the small bright features to oxygen adatoms can be found in the supplementary section. Only these features, which can be easily distinguished from others, were the subject of the pulsing experiments.An atomically-clean single crystalline Mo110) (Surface Preparation Laboratory) surface was prepared by 10 (Surfa−11 mbar). An electrochemically etched single crystalline tungsten tip45 was utilized in the following pulsing experiments. The bias voltage is applied to the sample with respect to the tip. No drift corrections have been applied to any of the STM images presented in this work. Images were obtained in constant current mode. Further details of the preparation of the MoOx2+/Mo(110) surface and STM experimental conditions are outlined elsewhere27.The STM experiments were performed at liquid nitrogen temperature (78 K), using a commercial STM from Createc, in an ultra-high-vacuum (UHV) system with a base pressure of 2 × 1027. Following pulsing, the surface is scanned again to determine if the adatom has been removed. The minimum voltage required to remove an adatom is referred to as the threshold voltage.Pulsing involves applying a bias, for some period of time, to the surface whilst the tip is in its vicinity. Different bias voltage and tip-surface distance change the probability of removing oxygen atoms and the area from which the adatoms are removed. Prior to pulsing, the surface is scanned. The tip is then positioned directly above an oxygen adatom. In the tunneling regime the thermal influence of the tunneling current, inelastic electron scattering and the electric field generated between the STM tip and the surface contribute to the removal of oxygen adatoms. In order to isolate the contribution of the electric field, a vertical offset is applied to the STM tip to remove it from the tunneling regime for the duration of the applied voltage pulse. The vertical offset is not the absolute tip-surface distance but is the distance the tip is moved from its initial position in the scanning tunneling regime. In order to ensure consistency in this height, the initial scanning bias voltage and tunneling current were kept constant throughout all experiments  and a tip offset of 1 nm (sufficient to remove the tip from tunneling regime) was used unless otherwise stated. The pulse duration for the following experiments was selected as 500 ms. Increasing the pulse duration above this value does not cause a significant change in the pulses’ effectiveness at removing an adatom47 on atom manipulation using the same microscope. The tip is approached into the tunneling regime and the feedback is then switched off. No changes in the tunneling current were detected during a 100 second interval. The error in applying a vertical offset is estimated to be no more than 0.1 Å. All experiments were performed 30 hours after filling the STM with liquid nitrogen, ensuring that the temperature is stable and therefore no thermal drift is present. The accuracy of the tip can be verified by scanning an area before and after pulsing and comparing the target position (the position selected on the first(before) STM image) and the real position (the position adatoms are removed from on the second(after) STM image). The target and real positions are within 1 Å, this high of precision can be observed in Fig. The stability of the tip position has been previously tested in our experimentsW. The probability of the oxygen adatom overcoming this barrier and penetrating the molybdenum surface layer to the oxide layer below depends on the frequency of attempts to overcome the barrier denoted by f (phonon frequency) and the time interval Δt in which attempts are made.To penetrate the surface, an oxygen adatom needs to overcome a potential barrier, A is the pre-exponential factor. The electric field generated by applying a negative bias to the tip with respect to the surface reduces this barrier. If the sample metal surface is taken to be a reference point at 0 potential, the potential barrier height reduction can be written as W − qψ, where q is the effective charge of an oxygen ion and ψ is the electric potential at the point of the oxygen ion. It follows that the probability of the oxygen adatom penetrating the surface in the presence of electric field E generated by the STM tip is:d, is approximately 1 nm) the electric potential can be be approximated by a simple model in which we consider the tip and surface as a parallel-plate capacitor. Then the potential at a distance of a from the plate is given by ψ:a corresponds to the distance between the oxygen adatom and the metallic surface, taken to be the topmost Mo plane. d is the tunneling distance, while Zoff is any additional offset. Substituting for ψ in Eq.  surface. In the FEM model, the tip is approximated as a conductive sphere and the MoO2 a perfectly flat metallic plane. An electric potential is applied to the tip and the electric field between tip and surface can then be determined. The electric potential at any point can be then calculated as a function of tip-surface distance, tip radius of curvature and bias voltage. This allows one to estimate the trends of the experimentally observed relationships, such as the threshold voltage for the removal of oxygen adatoms as a function of tip-surface distance. The electric potential is calculated at a distance of 0.4 nm above the first Mo layer of the O-Mo-O tri-layer according to DFT results27 seen in Fig. FEM simulationsIn order to simulate the relationship between the spot diameter and the applied bias voltage, which used a vertical offset of 1 nm, an experimental reference point is required. A reference point of 1 nm offset for an applied bias voltage of 3.3 V was taken from the threshold voltage vs. applied offset experiment, black circles in Fig. To simulate the dependence of the threshold voltage on the tip-surface distance, the electric potential directly below the tip at the adatom height of 0.4 nm above the surface is calculated as a function of the applied bias voltage for various tip-surface offsets. These dependencies can be interpolated in order to select a potential and determine the bias voltage required to generate said potential at different tip offsets. Comparing the resulting data for a wide range of potential values to the experimental results not only allows us to find the best match to experiment but also to estimate the potential required for the adatom to transition below the surface.Supplementary information


International harmonization of postgraduate medical education is gaining importance in the globalization of medical education. Harmonization is regarded as the establishment of common standards in education, while maintaining regional or local freedom to adapt training to contexts. During the development of a harmonized curriculum, tensions between standardization and contextualization may surface. To allow future harmonization projects to recognize these tensions and deal with them in a timely manner, this study aims to gain insight into tensions that may arise when developing a harmonized curriculum for postgraduate medical education in Obstetrics and Gynaecology in Europe.Focus groups were held with international curriculum developers to discuss challenges that resulted from tensions between standardization and contextualization when developing a harmonized European curriculum for postgraduate medical education in Obstetrics and Gynaecology. Data were analyzed through conventional content analysis, using the principles of template analysis.Tensions between standardization and contextualization in the development of a harmonized curriculum were apparent in two domains: 1) Varying ideas about what the harmonized curriculum means for the current curriculum and 2) Inconsistencies between educational principles and the reality of training. Additionally, we identified ways of dealing with these tensions, which were characterized as ‘negotiating flexibility’. Tensions between standardization and contextualization surfaced in the development phase of harmonizing a curriculum, partly because it was anticipated that there would be problems when implementing the curriculum. Tensions between standardization and contextualization are experienced in medical education. They may surface in the development of a harmonized curriculum in particular, which may challenge the process since harmonization is aimed at providing standards while respecting local autonomy. This study is the first to gain insight into the tensions that may arise when developing a harmonized curriculum for postgraduate medical education in Europe, to allow future harmonization projects to recognize these tensions and deal with them in a timely manner.Globalization of medical education has received growing attention –7. WhileIn medical education in general, balancing standardization of education with allowances for contextual variation is increasingly a subject of debate , 19. BatBefore we can build on this debate, the terms ‘standardization’ and ‘contextualization’ require further explanation. For the purpose of harmonizing postgraduate medical education, standardization of training refers to the use of common learning outcomes, strategies for training, and systems of assessment, for instance in an outcome-based curriculum , 23. In As suggested, tensions between standardization and contextualization may surface in medical education, especially when harmonizing postgraduate medical education. An important phase in harmonizing postgraduate medical education is curriculum development. Ideally, in this phase, curriculum developers recognize challenges and deal with them to ensure feasibility and effective implementation of the curriculum. Therefore, we aimed to explore if and how tensions between standardization and contextualization surface during the development of a harmonized curriculum in postgraduate medical education to better understand the process of harmonization and the challenges that it may bring. We do not aim to find straightforward solutions, rather we are convinced that understanding the challenges in harmonization may offer starting points for consideration to improve feasibility and sustainability of harmonization of postgraduate medical education in the future. Thus, taking into account the increasing globalization of medical education, this research is valuable for future projects and the evolvement of globalization.Recently, the European Board & College of Obstetrics and Gynaecology (EBCOG) took steps to harmonize postgraduate medical education in Obstetrics and Gynaecology (OBGYN) in Europe . We tookThe research questions were: 1) Do curriculum developers recognize and/or deal with tensions between standardization and contextualization in the development of a harmonized curriculum in postgraduate medical education, and if so 2) Where do such tensions surface?’We performed a conventional content analysis study with focus groups to gain an understanding of curriculum developers’ experiences of tensions between standardization and contextualization during the development of the European OBGYN curriculum . In focuAs explained, EBCOG has initiated the harmonization of postgraduate medical education in OBGYN in Europe . ParticiAll 38 curriculum developers were identified as potential participants for this study and were contacted by email. They received information about the study and an invitation to participate. To include as many of the curriculum developers as possible, the focus groups were scheduled during international project meetings. The first focus group consisted of the advisory committee of the project, representing EBCOG members and members of subspecialty societies in OBGYN. The second focus group consisted of ENTOG members, representing trainees in OBGYN from different European countries. Trainees were deliberately invited as a separate group to prevent hierarchical relations affecting group dynamics. The third focus group consisted of the project’s working group, including gynaecologists, trainees, educationalists and representatives of stakeholder groups. During each focus group session, two moderators facilitated the discussion (FS & JEA or AJG & JEA). All focus groups were held in English, which is the second language of most participants.The focus group moderators encouraged participants to share any challenges they had experienced during curriculum development. A lot of room was offered for reflection and elaboration. The topic guide of the focus groups was limited to the challenges experienced, although this was slightly structured by suggesting that challenges may be experienced at three levels . The moderators made no direct references to the concept of tensions between standardization and contextualization, since this was considered too conceptual and too restrictive for collection of valuable data. The topic guide did not evolve throughout the study, although the discussions in the three focus groups were very divergent because of their free flowing nature.Eventually, three focus groups were held, each with a duration of approximately one hour. In the first, second and third focus group there were seven, four and nine participants, respectively. The total of twenty participants represented twelve different European nationalities.Data were collected between November 2017 and February 2018. The focus groups were audio recorded and transcribed verbatim. Audio recordings and transcripts were stored in a secured environment. After analyzing the data, the recordings were destroyed. The main researcher (JEA) moderated all three focus groups and coded the transcripts with the help of MaxQDA2007. We performed conventional content analysis according to the steps of King’s template analysis to allow structured and hierarchal organization of the coding . ThroughParticipants were contacted in person and informed about the purpose of the study, the role of the participant in the study and the reason for selecting the participant. The voluntary nature of participation was stressed. The researchers ensured that participants were aware that if they decided to withdraw from participation after the focus groups had taken place, their input could not be eliminated from analysis. Also, the researchers stressed the confidential nature of the information that was shared within the focus groups. Consent for participation in the study was obtained orally and in writing and there was an opportunity for questions from participants and further explanation by the researchers. There were no personal advantages nor possible risks in participating in this study. The study was approved by the Ethical Review Board of the Netherlands Association for Medical Education in 2016 (NVMO-ERB file number 795). Only the main researcher (JEA) had access to the original data, and all research material was coded to guarantee anonymity.Three of the researchers  were also involved in developing the European OBGYN curriculum. Although they did not participate in the focus groups but moderated them, the study may have been influenced by their positions, beliefs and values regarding harmonization of postgraduate medical education in OBGYN. For instance, the main researcher (JEA) initially believed that harmonizing postgraduate medical education would improve the quality of training in all European countries. However, her involvement in exploring the process as a researcher resulted in new insights that have considerably changed her initial stance. The majority of the participants also shared a positive perspective on harmonization, which was underlined by their involvement in the development of the European OBGYN curriculum. Variation in their geographical and professional backgrounds ensured that the issues under study were addressed from different contexts.Analysis of the data revealed that the developers of the European OBGYN curriculum recognized tensions between standardization and contextualization during the development of the harmonized curriculum. We identified two domains in which these tensions were apparent. Additionally, we identified ways to deal with the tensions that emerged from our data. Whenever we refer to the ‘harmonized curriculum’, we mean the European OBGYN curriculum that was developed for the purpose of harmonization of postgraduate medical education in OBGYN in Europe. Whenever we refer to the ‘current curriculum’ we mean the OBGYN curriculum of a country or region with which the participants were familiar.Throughout the process of curriculum development, the harmonized curriculum gradually began to take shape. In this process, the participants automatically compared the harmonized curriculum with their current curriculum. Participants increasingly empathized with the way people from their country or region might ‘feel’ about the harmonized curriculum, which was based on participants’ understanding of the current curriculum. This ‘feeling’ was identified as a general feeling about the entire harmonized curriculum, rather than about specific elements of the harmonized curriculum. This gave participants an idea of how their current curriculum should change, in order for it to comply with the harmonized curriculum. While participants discussed these ideas with each other, they discovered that there were differences between countries. To illustrate this, we provide three examples of different ideas about the harmonized curriculum and we explain which strategies for change are implied in these ideas.I think in [country] there will not be a huge change and they won’t be open for a huge change because they have an elaborate curriculum and they will stay with it. Because there is a lot of work behind [the current curriculum] and it is established.For instance, one participant thought that her colleagues would mainly compare their current curriculum with the harmonized curriculum to confirm the quality of their current curriculum. In her view, medical educators in her country only need confirmation that their curriculum is already of high quality and therefore do not need the harmonized curriculum to show how to improve their current training. This was supported by the following quote:Another participant explained that she felt that her country considered the harmonized curriculum to be a benchmark to show which elements of the current curriculum could be improved. Therefore, her idea for evolvement of their current curriculum was that selected elements of training might be improved, for which the harmonized curriculum would be a good reference. For yet another country, a participant explained that she felt that the harmonized curriculum is so far from her country’s current curriculum that her idea of change towards the current curriculum is thorough reform and extensive improvement.These three examples reveal that tension is experienced between standardization and contextualization, since participants collectively struggled with the realization that there were different ideas about what the harmonized curriculum means for the current curriculum.Throughout curriculum development, specific educational principles were selected as the foundation for the harmonized curriculum. Examples of educational principles that the harmonized curriculum relies on are an outcome-based curriculum, a soft-skills framework, a focus on simulation and teamwork training and assessment through entrustment to ensure gradual independence.If the professor or specialist see I can do it, he will leave me do it. But if he has to sign I’m confident to do it, then maybe he will start making problems about it. Because he will put the signature there and then feels responsible for it. So it takes longer to reach the competences that already have [the competences] informal.Participants explained that in their experience, some principles might not be consistent with the reality of training in their constituent workplaces and some principles might even have an adverse effect if they were implemented in specific countries. For instance, participants described that one of the educational principles of the harmonized curriculum is that a trainee should gradually gain independence in practice, and that this should be officially documented in a portfolio using entrustment decisions. One trainee feared that by formalizing this process of gaining gradual independence, some supervisors might be more hesitant to leave trainees to practice independently, rather than to stimulate it, because of the formal process:Staff members, (…) they are subspecialized more and more nowadays to teach their trainees, again, about advanced things in perinatal medicine or reproductive medicine, urogynaecology. And obviously that’s going to be rectified now in this new curriculum. But it needs again a change of perception and that may in itself be, also be a challenge to change that.This type of tension between standardization and contextualization was recognized in more cases. As a second example, two participants pointed out that the harmonized curriculum describes a broad standard of general competencies in the curriculum content, while at the same time the current workforce of gynaecologists in Europe seems to be differentiating and subspecializing more and more. According to the participants, this tendency to subspecialize would lead to a workplace that offers mainly learning opportunities that are more subspecialty-specific, which may compete with learning opportunities for the development of more general competencies. This view was supported by the following quote:But still we do visits in many, many countries in Europe. And that also may contribute to getting more information as to how new curriculums will be received and how they think they will able to fit it into let’s say a local organization.In addition to the two domains of tensions, we identified in our data that participants already tried to deal with the tensions in the curriculum development phase. Participants’ understanding of the existence of tensions seemed to initiate considerations of how to deal with them. Participants suggested that curriculum developers should engage in conversations with national postgraduate medical education bodies as well as local sites to explore the different needs of the different contexts. One participant explained:I think that’s important to talk about to each other and discuss about it. Are we doing it as we thought we are doing it?And within the tailoring process there might be a stepwise change of course.Participants experienced that consideration of differences subsequently led to their understanding that flexibility of the harmonized curriculum is needed to allow the standards to align with different contexts. Therefore, the harmonized curriculum should be developed further, based on more understanding of specific needs for flexibility. Participants described the harmonized curriculum as a ‘document in progress’ for which flexibility should be negotiated. This is supported by the following quotes:And then the other thing is if, for example, a country says, we can’t actually deliver that part of the curriculum, does that matter?For instance, one of the participants remarked that a ‘pick-and-choose’ strategy for specific elements of the harmonized curriculum may undermine the desired effects of quality improvement of postgraduate medical education throughout Europe. This is a consideration of flexibility that requires negotiation.If you also accept that things take time. And maybe it will take five to ten years to reach [it]. And everyone will have their own from now on until ten years. And you have to accept that. The squares getting rounder and rounder you know? You have to accept that, because change, management of change is really hard.Overall, participants agreed that the harmonized curriculum should be regarded as a strong recommendation for OBGYN training, while it should still allow for flexibility to align it with different contexts. This shows that the curriculum developers experienced tensions between standardization and contextualization when developing the harmonized curriculum, but also searched for ways to deal with them, and felt that negotiating flexibility of recommendations deals with these tensions. The following quote stresses how participants perceived implementation and further development of the harmonized curriculum.The aim of this study was to gain insight into tensions that may arise in the development of a harmonized curriculum for postgraduate medical education in Europe. Our findings show that curriculum developers recognized tensions between standardization and contextualization when developing a harmonized curriculum. Tensions surfaced due to varying ideas about what the harmonized curriculum means for the current curriculum, and due to inconsistencies between educational principles and the reality of training. Also, curriculum developers shared how these tensions may be dealt with.Reflecting further on the first domain in which tensions surfaced, we identified a challenge that requires some further exploration. The objective of the development of the harmonized curriculum is to improve training throughout Europe, although the tensions in the first domain show that different countries have different ideas about what the harmonized curriculum means for the current curriculum. This raises the question whether the intention to improve training across Europe can be fulfilled through harmonization at all. We wonder whether harmonization could unintentionally lead to an increase of inequality in postgraduate medical education if some countries evolve towards the harmonized curriculum easily, whereas other countries need more time and effort to change. The countries that need to invest more time and effort to adopt the harmonized curriculum need to initiate their development while being aware of the size of the effort. This awareness may affect a country’s perception of harmonization in a negative way and potentially lead to undermining its efforts even further. For countries that do not need to invest so much time and effort to change towards the harmonized curriculum, harmonization may seem a more achievable aim. In a harmonization process it is important to be aware of these different effects on countries. Negotiating room for flexibility within a harmonized curriculum allows for better alignment of the harmonized curriculum to different contexts and makes adaptation of current curricula more feasible.This study originated from discussions around tensions between standardization and contextualization in medical education in general. In their paper, Bates et al. recommended to embrace both standardization and contextualization as valuable concepts, accepting the tensions they cause, and we should aim to deliver medical education that is both globally responsible and locally engaged . RelatinLiterature regarding change processes in postgraduate medical education curricula describes how expectations of a novel curriculum are not always met during implementation in the context –32. WhatThe findings of this study may benefit the harmonization of postgraduate medical education in the future. We propose a strategy to allow flexibility of a harmonized curriculum. That is, to consider if a harmonized curriculum could be developed at two levels. For instance, a harmonized curriculum can describe educational outcomes and strategies for all the countries involved. In addition, all countries may describe how they intend to interpret and ensure implementation of the harmonized curriculum in the context of training. For a harmonized curriculum to serve different contexts, local interpretation may be necessary. This two-level approach of curriculum development may answer the need for local flexibility, while still allowing for harmonization of postgraduate medical education.The strength of this study is that it provides a novel insight into tensions between standardization and contextualization in the development of a harmonized curriculum for postgraduate medical education in Europe. As explained, three of the researchers as well as most participants in this research share a positive conception around harmonization. This may have affected the outcomes of the study. Future research could be aimed at further exploring more deeply seated tensions, by including participants with potentially more diverging conceptions around harmonization.The findings of this study open up possibilities for further research. More empirical evidence is required on the value of the suggested negotiation of flexibility through a two-level approach of curriculum development. Dealing with tensions between standardization and contextualization when harmonizing postgraduate medical education is an investment whose benefits are not clear yet. As previously described, harmonization of postgraduate medical education aims to enhance the international exchange of best practices in healthcare as well as the mobility of medical specialists . FurtherCurriculum developers recognize tensions between standardization and contextualization in the development of a harmonized curriculum. Tensions may surface due to varying ideas about what the harmonized curriculum means for the current curriculum, and due to inconsistencies between educational principles and the reality of training. Negotiating room for flexibility in anticipation of national implementation challenges seems to be the main strategy to deal with perceived tensions in the harmonization of postgraduate medical education.