In vitro immunity: an overview of immunocompetent organ-on-chip models.

Impressive advances have been made to replicate human physiology in vitro over the last few years due to the growth of the organ-on-chip (OoC) field in both industrial and academic settings. OoCs are a type of microphysiological system (MPS) that imitates functional and dynamic aspects of native human organ biology on a microfluidic device. Organoids and organotypic models, ranging in their complexity from simple single-cell to complex multi-cell type constructs, are being incorporated into OoC microfluidic devices to better mimic human physiology. OoC technology has now progressed to the stage at which it has received official recognition by the Food and Drug Administration (FDA) for use as an alternative to standard procedures in drug development, such as animal studies and traditional in vitro assays. However, an area that is still lagging behind is the incorporation of the immune system, which is a critical element required to investigate human health and disease. In this review, we summarise the progress made to integrate human immunology into various OoC systems, specifically focusing on models related to organ barriers and lymphoid organs. These models utilise microfluidic devices that are either commercially available or custom-made. This review explores the difference between the use of innate and adaptive immune cells and their role for modelling organ-specific diseases in OoCs. Immunocompetent multi-OoC models are also highlighted and the extent to which they recapitulate systemic physiology is discussed. Together, the aim of this review is to describe the current state of immune-OoCs, the limitations and the future perspectives needed to improve the field.

Immunogenicity of COVID-19 vaccines in patients with follicular lymphoma receiving frontline chemoimmunotherapy.

Immune responses to primary COVID-19 vaccination were investigated in 58 patients with follicular lymphoma (FL) as part of the PETReA trial of frontline therapy (EudraCT 2016-004010-10). COVID-19 vaccines (BNT162b2 or ChAdOx1) were administered before, during or after cytoreductive treatment comprising rituximab (depletes B cells) and either bendamustine (depletes CD4+ T cells) or cyclophosphamide-based chemotherapy. Blood samples obtained after vaccine doses 1 and 2 (V1, V2) were analysed for antibodies and T cells reactive to the SARS-CoV-2 spike protein using the Abbott Architect and interferon-gamma ELISpot assays respectively. Compared to 149 healthy controls, patients with FL exhibited lower antibody but preserved T-cell responses. Within the FL cohort, multivariable analysis identified low pre-treatment serum IgA levels and V2 administration during induction or maintenance treatment as independent determinants of lower antibody and higher T-cell responses, and bendamustine and high/intermediate FLIPI-2 score as additional determinants of a lower antibody response. Several clinical scenarios were identified where dichotomous immune responses were estimated with >95% confidence based on combinations of predictive variables. In conclusion, the immunogenicity of COVID-19 vaccines in FL patients is influenced by multiple disease- and treatment-related factors, among which B-cell depletion showed differential effects on antibody and T-cell responses.

Small molecule induced STING degradation facilitated by the HECT ligase HERC4.

Stimulator of interferon genes (STING) is a central component of the cytosolic nucleic acids sensing pathway and as such master regulator of the type I interferon response. Due to its critical role in physiology and its' involvement in a variety of diseases, STING has been a focus for drug discovery. Targeted protein degradation (TPD) has emerged as a promising pharmacology for targeting previously considered undruggable proteins by hijacking the cellular ubiquitin proteasome system (UPS) with small molecules. Here, we identify AK59 as a STING degrader leveraging HERC4, a HECT-domain E3 ligase. Additionally, our data reveals that AK59 is effective on the common pathological STING mutations, suggesting a potential clinical application of this mechanism. Thus, these findings introduce HERC4 to the fields of TPD and of compound-induced degradation of STING, suggesting potential therapeutic applications.

Evaluation of shaft angle to ball-to-target line as a predictor of horizontal delivery plane angle in the golf swing.

The shaft angle to the ball-to-target line at various points in the golf swing is used by coaches as an indication of the horizontal delivery plane angle (HPA). The aim of the current study was to understand to what degree this simplified method of using the shaft orientation can predict the orientation of the HPA. Fifty-two male golfers hit 40 drives each in an indoor biomechanics laboratory. Between-subject regression models were created for the relationship between the HPA and the shaft angle to the ball-to-target line at three different swing positions. Additionally, single subject regression models were created for each subject for the small variables. The only significant between-subjects regression model was for mid-downswing (Adjusted R2 = 89.5%, RMSE = 2.41°); however, this was deemed not accurate enough to distinguish differences between typical driver and wedge HPA. The only shaft position to have significant single-subject regression models for all participants was mid-downswing. The mean RMSE for those models was determined to be low enough to distinguish typical driver and wedge swing planes within individuals. Overall, the shaft angle was only deemed accurate enough to predict the HPA within individual subjects, and only for mid-downswing.

An Organotypic Human Lymph Node Model Reveals the Importance of Fibroblastic Reticular Cells for Dendritic Cell Function.

BACKGROUND: Human lymph node (HuLN) models have emerged with invaluable potential for immunological research and therapeutic application given their fundamental role in human health and disease. While fibroblastic reticular cells (FRCs) are instrumental to HuLN functioning, their inclusion and recognition of importance for organotypic in vitro lymphoid models remain limited. METHODS: Here, we established an in vitro three-dimensional (3D) model in a collagen-fibrin hydrogel with primary FRCs and a dendritic cell (DC) cell line (MUTZ-3 DC). To study and characterise the cellular interactions seen in this 3D FRC-DC organotypic model compared to the native HuLN; flow cytometry, immunohistochemistry, immunofluorescence and cytokine/chemokine analysis were performed. RESULTS: FRCs were pivotal for survival, proliferation and localisation of MUTZ-3 DCs. Additionally, we found that CD1a expression was absent on MUTZ-3 DCs that developed in the presence of FRCs during cytokine-induced MUTZ-3 DC differentiation, which was also seen with primary monocyte-derived DCs (moDCs). This phenotype resembled HuLN-resident DCs, which we detected in primary HuLNs, and these CD1a- MUTZ-3 DCs induced T cell proliferation within a mixed leukocyte reaction (MLR), indicating a functional DC status. FRCs expressed podoplanin (PDPN), CD90 (Thy-1), CD146 (MCAM) and Gremlin-1, thereby resembling the DC supporting stromal cell subset identified in HuLNs. CONCLUSION: This 3D FRC-DC organotypic model highlights the influence and importance of FRCs for DC functioning in a more realistic HuLN microenvironment. As such, this work provides a starting point for the development of an in vitro HuLN.

Evaluating consistency of physical activity and exercise prescription in the UK for people with diabetes - a Delphi study.

Introduction: Increased physical activity is recommended as a cost-effective measure to tackle long-term management of people with diabetes, but research on interventions lacks consistency in terms of effective duration and modality. Methods: The aim of this study was to evaluate expert consensus on exercise and physical activity prescription via a three-round Delphi study conducted with 45 UK-based health and fitness professionals experienced in prescribing exercise or physical activity to people with diabetes. Results: The majority of items put forward to the panel reached consensus with 70% or above voting these items as important, but the details of the type, duration and/or modality of exercise or physical activity prescription within these items often contradicted each other, suggesting that patients are receiving inconsistent advice. The range of different exercise prescription found in this study suggests that patients are being given inconsistent and potentially confusing advice, which may affect their participation in exercise and long-term lifestyle change. Conclusion: More consistent promotion of advice from healthcare and fitness professionals may help with increasing physical activity in this participant group and achieving long term behavior change, reducing patient symptoms as well as reducing the cost to the National Health Service (NHS).

A Scoping Review of the Validity and Reliability of Smartphone Accelerometers When Collecting Kinematic Gait Data.

The aim of this scoping review is to evaluate and summarize the existing literature that considers the validity and/or reliability of smartphone accelerometer applications when compared to 'gold standard' kinematic data collection (for example, motion capture). An electronic keyword search was performed on three databases to identify appropriate research. This research was then examined for details of measures and methodology and general study characteristics to identify related themes. No restrictions were placed on the date of publication, type of smartphone, or participant demographics. In total, 21 papers were reviewed to synthesize themes and approaches used and to identify future research priorities. The validity and reliability of smartphone-based accelerometry data have been assessed against motion capture, pressure walkways, and IMUs as 'gold standard' technology and they have been found to be accurate and reliable. This suggests that smartphone accelerometers can provide a cheap and accurate alternative to gather kinematic data, which can be used in ecologically valid environments to potentially increase diversity in research participation. However, some studies suggest that body placement may affect the accuracy of the result, and that position data correlate better than actual acceleration values, which should be considered in any future implementation of smartphone technology. Future research comparing different capture frequencies and resulting noise, and different walking surfaces, would be useful.

Unclean Cooking Fuel Use and Slow Gait Speed Among Older Adults From 6 Countries.

BACKGROUND: Outdoor air pollution has been reported to be associated with frailty (including slow gait speed) in older adults. However, to date, no literature exists on the association between indoor air pollution (eg, unclean cooking fuel use) and gait speed. Therefore, we aimed to examine the cross-sectional association between unclean cooking fuel use and gait speed in a sample of older adults from 6 low- and middle-income countries (China, Ghana, India, Mexico, Russia, and South Africa). METHODS: Cross-sectional, nationally representative data from the World Health Organization Study on global AGEing and adult health were analyzed. Unclean cooking fuel use referred to the use of kerosene/paraffin, coal/charcoal, wood, agriculture/crop, animal dung, and shrubs/grass based on self-report. Slow gait speed referred to the slowest quintile based on height, age, and sex-stratified values. Multivariable logistic regression and meta-analysis were done to assess associations. RESULTS: Data on 14 585 individuals aged ≥65 years were analyzed (mean [standard deviation] age 72.6 [11.4] years; 45.0% males). Unclean cooking fuel use (vs clean cooking fuel use) was significantly associated with higher odds for slow gait speed (odds ratio = 1.45; 95% confidence interval: 1.14-1.85) based on a meta-analysis using country-wise estimates. The level of between-country heterogeneity was very low (I2 = 0%). CONCLUSIONS: Unclean cooking fuel use was associated with slower gait speed among older adults. Future studies of longitudinal design are warranted to provide insight into the underlying mechanisms and possible causality.

The global state of play: A study of the demographic characteristics of disability golfers.

BACKGROUND: Golf is a moderate-intensity physical activity that provides positive physical and mental health benefits. However, the inclusiveness of the sport for individuals with disabilities globally is unknown. OBJECTIVE: To characterize the demographics and disability characteristics of individuals engaging in disability golf globally. It was hypothesized that the majority of participants would be middle-aged, male, and from countries with higher gross domestic product, similar to the nondisabled population. DESIGN: Descriptive, cross-sectional analysis using European Disability Golf Association (EDGA) database. SETTING: Various international golf tournaments. PARTICIPANTS: Golfers (n = 1734) with disability from 44 countries registered with the EDGA (2017-2021). INTERVENTIONS: Not applicable. MAIN OUTCOMES: Descriptive analyses of frequencies, distributions, and means differences of demographic characteristics (age, gender, type of disability, level of handicap, golf cart use, and country of origin) of golfers with disability were performed. Data provided analysis of the association between number of participants and a country's gross domestic product (GDP). RESULTS: Individuals had a mean age of 52.5 (±15.6) years: 1589 (91.6%) male and 145 (8.4%) female. Twenty-three countries had female participation. The most commonly reported primary disability diagnoses were neurologic (24.8%), orthopedic (21.4%), and amputation below the knee (14.4%). Neurologic impairments (24.7%) were most common in men and orthopedic impairments (29.7%) were most common in women. Individuals with neurological impairments (27.4%) most frequently required golf carts to play. The GDP of a country had a positive correlation (r = 0.68) with the number of registered golfers with disability. CONCLUSION: Golf is played by individuals with a variety of disabilities and provides numerous benefits. However, there is an underrepresentation of youth, women, and individuals with certain impairments and from lower-income countries. These are the potential areas of opportunity to improve engagement and inclusiveness of golf.

Tracking transients in steelpan strikes using surveillance technology.

This paper presents advancements in tracking features in high-speed videos of Caribbean steelpans illuminated by electronic speckle pattern interferometry, made possible by incorporating robust computer vision libraries for object detection and image segmentation, and cleaning of the training dataset. Besides increasing the accuracy of fringe counts by 10% or more compared to previous work, this paper introduces a segmentation-regression map for the entire drum surface yielding interference fringe counts comparable to those obtained via object detection. Once trained, this model can count fringes for musical instruments not part of the training set, including those with non-elliptical antinode shapes.

Surface Coverage as an Important Parameter for Predicting Selectivity Trends in Electrochemical CO2 Reduction.

The electrochemical CO2 reduction reaction (CO2RR) is important for a sustainable future. Key insights into the reaction pathways have been obtained by density functional theory (DFT) analysis, but so far, DFT has been unable to give an overall understanding of selectivity trends without important caveats. We show that an unconsidered parameter in DFT models of electrocatalysts-the surface coverage of reacting species-is crucial for understanding the CO2RR selectivities for different surfaces. Surface coverage is a parameter that must be assumed in most DFT studies of CO2RR electrocatalysts, but so far, only the coverage of nonreacting adsorbates has been treated. Explicitly treating the surface coverage of reacting adsorbates allows for an investigation that can more closely mimic operating conditions. Furthermore, and of more immediate importance, the use of surface coverage-dependent adsorption energies allows for the extraction of ratios of adsorption energies of CO2RR intermediates (COOHads and HCOOads) that are shown to be predictive of selectivity and are not susceptible to systematic errors. This approach allows for categorization of the selectivity of several monometallic catalysts (Pt, Pd, Au, Ag, Zn, Cu, Rh, W, Pb, Sn, In, Cd, and Tl), even problematic ones such as Ag or Zn, and does so by only considering the adsorption energies of known intermediates. The selectivity of the further reduction of COOHads can now be explained by a preference for Tafel or Heyrovsky reactions, recontextualizing the nature of selectivity of some catalysts. In summary, this work resolves differences between DFT and experimental studies of the CO2RR and underlines the importance of surface coverage.

Calculating the acoustic input impedance of a simplified brass instrument as an educational laboratory activity.

The concept of acoustic impedance is often difficult for students in introductory acoustics courses to make sense of, especially students without advanced mathematics backgrounds. This work summarizes a laboratory activity for students in a general education musical acoustics class where a simplified brass musical instrument is examined, focusing on how the geometry of the air column affects the input impedance of the instrument. Students are guided through making bore profile measurements for use in a computation of the input impedance. Options for making experimental measurements of the simplified instrument are explained. The laboratory activity was successfully used with students who reported their increased understanding of the acoustics of brass musical instruments.

Validating the Cambridge Protocol: Reliability of Hip Muscle Strength Measurements Using a Motorized Dynamometer and Electromyography.

BACKGROUND: Muscle weakness is common after injury in athletes and in the presence of hip pathology. It will cause abnormal hip biomechanics and can predict future injury. However, objective measurement of hip muscle strength is difficult to perform accurately and reliably. Therefore, it is challenging to determine when an athlete has returned to preinjury levels of strength. In addition, there is currently no standardized method of obtaining measurements, which prevents the data being compared or shared between research centers. PURPOSE: The purpose of this study is to comprehensively assess the inter- and intraobserver reliability of our standardized muscle strength measurement protocol. STUDY DESIGN: Descriptive laboratory study. LEVEL OF EVIDENCE: Level 3, inception cohort study. METHODS: A total of 16 healthy male volunteers (age = 28.3 ± 7.9 years) were recruited. Those with a previous history of hip injuries or disorders were excluded. These volunteers underwent strength testing according to the Cambridge Protocol on 4 separate occasions, performed by 2 independent assessors. Maximal voluntary contractions, fatigue torque fluctuations, and electromyography measurements were recorded. Intra- and interobserver reliability was assessed using intraclass correlation coefficient (ICC). RESULTS: Good-to-excellent correlation was seen for both intra- and interobserver reliability across almost all hip movements for maximal contractions: ICC ranged 0.78 to 0.93 and 0.78 to 0.96, respectively. The standard error of the mean for all hip movements was also extremely low at 2% to 3%. CONCLUSION: The Cambridge Protocol is a highly reliable method for objective measurement of hip muscle strength. We recommend future studies use this protocol, or the principles underpinning it, to enable data sharing and comparison across different studies. CLINICAL RELEVANCE: This is a description and analysis of hip muscle strength measurement. If widely used, it will allow for accurate and objective strength assessment and closer monitoring of hip injuries and pathology.

Development of a qPCR-based method for counting overwintering spruce budworm (Choristoneura fumiferana) larvae collected during fall surveys and for assessing their natural enemy load: a proof-of-concept study.

BACKGROUND: In eastern Canada, surveys of overwintering 2nd instar spruce budworm (Choristoneura fumiferana) larvae ('L2s') are carried out each fall to guide insecticide application decisions in the following spring. These surveys involve the collection of fir and spruce branches in selected stands, followed by the mechanical/chemical removal of larvae. The latter then are counted manually on filter papers, using a stereomicroscope. Considering the significant effort and difficulties which this manual counting entails, we developed a quantitative (q)PCR-based 'molecular counting' approach designed to make this step less tedious. RESULTS: Using the C. fumiferana mitochondrial cytochrome c oxidase 1 (COI) gene as a target for qPCR DNA quantification, we show that the amount of DNA in a larval extract is strongly correlated with the number of larvae used to generate that extract, and that molecular estimates of L2 counts are comparable to those generated using the manual approach. In addition, we used the same DNA extracts to monitor the microsporidian pathogen Nosema fumiferanae, and the hymenopteran parasitoids Glypta fumiferanae and Apanteles fumiferanae in overwintering L2s employing a subset of a TaqMan assay developed by Nisole et al. (2020) for the identification of budworm natural enemies. We show that the proportion of individuals affected by each natural enemy in samples containing a known number of larvae can be estimated from presence/absence data through the binomial probability distribution. CONCLUSION: The present proof-of-principle study shows that a molecular approach for counting L2s and assessing their natural enemy load is clearly possible and is expected to generate reliable results. © 2021 Her Majesty the Queen in Right of Canada. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry. Reproduced with the permission of the Minister of Natural Resources Canada.

Electroreduction of CO2/CO to C2 Products: Process Modeling, Downstream Separation, System Integration, and Economic Analysis.

Direct electrochemical reduction of CO2 to C2 products such as ethylene is more efficient in alkaline media, but it suffers from parasitic loss of reactants due to (bi)carbonate formation. A two-step process where the CO2 is first electrochemically reduced to CO and subsequently converted to desired C2 products has the potential to overcome the limitations posed by direct CO2 electroreduction. In this study, we investigated the technical and economic feasibility of the direct and indirect CO2 conversion routes to C2 products. For the indirect route, CO2 to CO conversion in a high temperature solid oxide electrolysis cell (SOEC) or a low temperature electrolyzer has been considered. The product distribution, conversion, selectivities, current densities, and cell potentials are different for both CO2 conversion routes, which affects the downstream processing and the economics. A detailed process design and techno-economic analysis of both CO2 conversion pathways are presented, which includes CO2 capture, CO2 (and CO) conversion, CO2 (and CO) recycling, and product separation. Our economic analysis shows that both conversion routes are not profitable under the base case scenario, but the economics can be improved significantly by reducing the cell voltage, the capital cost of the electrolyzers, and the electricity price. For both routes, a cell voltage of 2.5 V, a capital cost of $10,000/m2, and an electricity price of <$20/MWh will yield a positive net present value and payback times of less than 15 years. Overall, the high temperature (SOEC-based) two-step conversion process has a greater potential for scale-up than the direct electrochemical conversion route. Strategies for integrating the electrochemical CO2/CO conversion process into the existing gas and oil infrastructure are outlined. Current barriers for industrialization of CO2 electrolyzers and possible solutions are discussed as well.

ConvNets for counting: Object detection of transient phenomena in steelpan drums.

We train an object detector built from convolutional neural networks to count interference fringes in elliptical antinode regions in frames of high-speed video recordings of transient oscillations in Caribbean steelpan drums, illuminated by electronic speckle pattern interferometry (ESPI). The annotations provided by our model aim to contribute to the understanding of time-dependent behavior in such drums by tracking the development of sympathetic vibration modes. The system is trained on a dataset of crowdsourced human-annotated images obtained from the Zooniverse Steelpan Vibrations Project. Due to the small number of human-annotated images and the ambiguity of the annotation task, we also evaluate the model on a large corpus of synthetic images whereby the properties have been matched to the real images by style transfer using a Generative Adversarial Network. Applying the model to thousands of unlabeled video frames, we measure oscillations consistent with audio recordings of these drum strikes. One unanticipated result is that sympathetic oscillations of higher-octave notes significantly precede the rise in sound intensity of the corresponding second harmonic tones; the mechanism responsible for this remains unidentified. This paper primarily concerns the development of the predictive model; further exploration of the steelpan images and deeper physical insights await its further application.

How Is Jump Performance Affected in Male Athletes when Completed with a Visual Impairment?

SIGNIFICANCE: High-, long-, and triple-jump athletic events may need to consider whether it is appropriate to group vision-impaired athletes in the same classification with loss of different visual functions, and a greater emphasis may need to be placed on the visual field (VF) within the current classification system used. PURPOSE: Athletes with vision impairment are grouped, based on their visual function, into one of three different classes (B1, B2, and B3, with B1 being the most severe). Athletes in class B2 have loss in visual acuity (VA; range, 1.50 to 2.60 logMAR) or VF (constricted to a diameter of <10°). The current study investigated how loss of different visual function (VA or VF) within the same class impacts jumping performance, a fundamental component in long-, triple-, and high-jump athletic events. METHODS: Ten subelite male athletes (age, 21.6 ± 0.96 years; height, 178.8 ± 2.97 cm; mass, 82.2 ± 10.58 kg) with normal vision who participate in athletics were recruited. Participants completed drop jumps in four vision conditions: habitual vision condition (Full), VA no better than 1.60 logMAR (B2-VA), VF restricted to <10° (B2-VF), and VA no better than 1.30 logMAR (B3-VA). RESULTS: Meaningful differences were observed between Full and B2-VF conditions. After rebound, vertical velocity at take-off was highest in Full condition (2.84 ± 0.35 m · s-1; 95% confidence interval [CI], 2.68 to 2.99 m · s-1) and was lowest in B2-VF condition (20% reduction; 2.32 ± 0.29 m · s-1; 95% CI, 2.16 to 2.48 m · s-1). Peak vertical jump height was highest in Full (0.42 ± 0.10 m; 95% CI, 0.38 to 0.46 m) and reduced by 40% in B2-VF (0.28 ± 0.07 m; 95% CI, 0.24 to 0.32 m). Minimal differences were found between Full and B2-VA, or B3-VA conditions. CONCLUSIONS: Jump performance is compromised in athletes with simulated vision impairment. However, decrements in performance seem specific to those with severely constricted VF. Those with reduced VA (in B2-VA and B3-VA classes) seem to produce performance comparable to those with normal vision.

Marker location and knee joint constraint affect the reporting of overhead squat kinematics in elite youth football players.

Motion capture systems are used in the analysis and interpretation of athlete movement patterns for a variety of reasons, but data integrity remains critical regardless. The extent to which marker location or constraining degrees of freedom (DOF) in the biomechanical model impacts on this integrity lacks consensus. Ten elite academy footballers performed bilateral overhead squats using a marker-based motion capture system. Kinematic data were calculated using four different marker sets with 3DOF and 6DOF configurations for the three joint rotations of the right knee. Root mean squared error differences between marker sets ranged in the sagittal plane between 1.02 and 4.19 degrees to larger values in the frontal (1.30-6.39 degrees) and transverse planes (1.33 and 7.97 degrees). The cross-correlation function of the knee kinematic time series for all eight marker-sets ranged from excellent for sagittal plane motion (>0.99) but reduced for both coronal and transverse planes (<0.9). Two-way ANOVA repeated measures calculated at peak knee flexion revealed significant differences between marker sets for frontal and transverse planes (p < 0.05). Pairwise comparisons showed significant differences between some marker sets. Marker location and constraining DOF while measuring relatively large ranges of motion in this population are important considerations for data integrity.

Evaluating dynamic similarity of fixed, self-selected and anatomically scaled speeds in non-linear analysis of gait during treadmill running.

OBJECTIVES: The aim of this study is to evaluate how speed affects non-linear measures of variability. Fixed and self-selected speeds were compared to an anatomically scaled speed calculated based on leg length to evaluate which provided a more reproducible result between subjects. METHODS: Sixteen subjects ran on a treadmill at a fixed, scaled and self-selected speed and at ±10% in each case. Kinematic data were collected for two minutes at 250 Hz for each trial. Sample entropy (SaEn) and maximum Lyapunov exponents (LyE) were calculated from the sagittal knee and hip joint angles to evaluate regularity of gait and local stability. These nonlinear measures were compared to evaluate the dynamic similarity of the movement in each case, and to evaluate speed as a confounding variable in non-linear analysis. RESULTS: An anatomically scaled speed shows more dynamic similarity than a fixed or self-selected speed with the lowest observed coefficient of variation for each measure. This was found to be statistically significant for both nonlinear measures of the hip (SaEn p = 0.038; LyE p = 0.040). Speed was not found to be a confounding variable in non-linear analysis of running gait of a healthy population (η2 < 0.05). CONCLUSIONS: Changes in speed by ±10% do not significantly affect stability and variability of gait for healthy participants, suggesting that they make adaptations to ensure optimal gait variability. Anatomically scaled speeds provide a more reliable methodology for both linear and non-linear analysis by providing a definitive protocol, suggesting it could replace self-selected or fixed speeds in future research.