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Background: The IL‐3‐pSTAT5 assay, a new, rapid, and standardized flow‐cytometry‐based assay may compensate for several limitations of the colony‐forming unit (CFU) assay typically used for stem cell potency assessments of cord blood units (CBU). We performed an inter‐laboratory evaluation of the performance of this new assay. Study design and methods: This Biomedical Excellence for Safer Transfusion (BEST) Collaborative multicenter, international study included 15 participants from public cord blood banks (CBBs), CBB‐supporting research laboratories, and stem cell laboratories. To perform the IL‐3‐pSTAT5 assay, participating centers received reagents, instructions, and 10 blind CBU samples, including eight normal samples and two samples exposed to a transient warming event. We measured inter‐laboratory agreement qualitatively (proportion of correctly classified samples) and quantitatively (coefficient of variation [CV], correlation coefficients, receiver operating characteristics (ROC) curve, and intraclass correlation coefficient [ICC]). Results: The qualitative agreement was 97.3% (i.e., 107/110; Fleiss' kappa = 0.835). The average CV on a per‐sample basis was 11.57% among all samples, 8.99% among normal samples, and on a per‐center basis was 9.42% among normal samples. In a correlation matrix that compared results across centers, the mean Pearson's correlation coefficient was 0.88 (standard deviation = 0.04). The ICC was 0.83 (95% confidence interval = 0.68–0.95). The area under the curve (AUC) from the ROC curve was 0.9974. Discussion: Excellent qualitative and quantitative agreement was exhibited across laboratories. The IL‐3‐pSTAT5 assay may therefore be implemented in flow cytometry laboratories to rapidly and reliably provide standardized measures of stem cell potency in CBUs. [ABSTRACT FROM AUTHOR]

Hollow fiber modules are commonly used to conveniently and efficiently remove cryoprotective agents (CPAs) from cryopreserved cell suspensions. In this paper, a steady-state model coupling mass transfers across cell and hollow fiber membranes is theoretically developed to evaluate the removal of CPAs from cryopreserved blood using hollow fiber modules. This steady-state model complements the unsteady-state model, which was presented in our previous study. The steady-state model, unlike the unsteady-state model, can be used to evaluate the effect of ultrafiltration flow rates on the clearance of CPAs. The steady-state model is validated by experimental results, and then is compared with the unsteady-state model. Using the steady-state model, the effects of ultrafiltration flow rates, NaCl concentrations in dialysate, blood flow rates and dialysate flow rates on CPA concentration variation and cell volume response are investigated in detail. According to the simulative results, the osmotic damage of red blood cells can easily be reduced by increasing ultrafiltration flow rates, increasing NaCl concentrations in dialysate, increasing blood flow rates, or decreasing dialysate flow rates. [DOI: 10.1115/1.4000110] Keywords: mass transfer, cryoprotective agent, ultrafiltration, cell volume, hollow fiber

Trials to explore the feasibility and reproducibility of islet transplantation by using Edmonton protocol so as to improve glycemic control in type 1 diabetes mellitus persons who are disabled by refractory hypoglycemia is conducted. This procedure successfully restores long-term endogenous insulin production and glycolic stability in cases of type 1 diabetes mellitus and unstable control but insulin independence is usually not sustainable and persistent islet function even without insulin independence provides protection from severe hypoglycemia and improved levels of glycated hemoglobin.

Amniotic products are potent immunomodulators used clinically to repair tissue injury. Little information exists regarding the potential of cell-free human amniotic fluid (hAF) to treat cardiovascular disease. Herein, we sought to determine the influence and efficacy of acellular hAF on myocardial ischemia-reperfusion injury. Processed hAF was obtained from volunteer donors at the time of elective caesarean section and manufactured using proprietary methods. Left anterior descending coronary artery ligation was performed on rats for 60 min. Thirty minutes after release and reperfusion, either saline or hAF was injected intramyocardially. Serial echocardiography revealed that compared with saline-injected rats, hAF animals maintained their ejection fraction and did not adversely remodel through the 4-wk period. This preserved ventricular function correlated with decreased infarct size, less fibrosis, and reduced expression of cytokines and infiltrating inflammatory cells. Comparative arrays of different donor hAF lots confirmed the presence of a wide array of immunomodulatory and host-defense proteins. The observed functional cardioprotection was furthermore evident when given intravenously and across multiple hAF donors. In conclusion, our data demonstrate, for the first time, the cardioprotective effect of acellular hAF on myocardial injury. These observations spanned across diverse donors and likely result from the mixture of a plethora of naturally produced cytokines, chemokines, and immune-modulating proteins rather than a single, defined mechanistic culprit. The ubiquitous availability of hAF as a cell-free solution further suggests its potential for widespread adoption as a therapy for myocardial ischemia-reperfusion injury. [ABSTRACT FROM AUTHOR]

Background: Growth in size and complexity of clinical hematopoietic progenitor cell (HPC) transplant programs necessitates parallel increases in cellular therapy laboratory (CTL) workload. Typically individually developed, HPC product processing is labor and time intensive. Variation in procedure type and numbers across CTLs complicates direct comparisons, and benchmark data are not readily available.Study Design and Methods: Studies were undertaken at seven CTLs. Transplant volume and staff numbers were determined. Staff recorded time performing tasks broken down into steps: paperwork, product acceptance, transport/infusion, processing, and cryopreservation. Times were added to obtain total times for 15 common CTL procedures.Results: Annual transplant volume ranged from 53.4 to 463.2, with products processed by a range of 2 to 10 dedicated CTL staff. Paperwork time constituted 23.7% to 62.3% total time; product processing time accounted for 1.8 (for National Marrow Donor Program product receipt) to 62.6% (for red blood cell reduction of allogeneic HPC products from bone marrow) of total processing time. Mean time for 15 procedures ranged from 1.27 to 8.28 hours (standard deviation range, 0.35-2.71 hr). Mean time for products from bone marrow versus peripheral blood was 6.6 ± 2.0 versus 5.5 ± 1.1 hours (p = 0.02). Cryopreservation (6.5 ± 1.6 vs. 4.4 ± 0.85 hr; p < 0.01) and manipulation (6.4 ± 1.5 vs. 4.4 ± 0.85 hr; p < 0.01) added time.Conclusion: CTL procedures are time intensive, with wide intra- and inter-CTL variation. Paperwork accounted for substantial portion of total time across procedures. Bone marrow source, cryopreservation, and manipulation contributed to longer times. These findings provide concrete data on which to build regarding CTL workload capacity. [ABSTRACT FROM AUTHOR]

Background: Viability testing is a common practice in laboratories. The goal of this study was to ascertain current laboratory practices internationally for performing viability testing for cryopreserved cord blood (CB) products and glean information about how to standardize the method to improve interlaboratory reproducibility.Study Design and Methods: A survey to evaluate current laboratory practices for viability testing was designed and distributed internationally. The question topics included sampling and testing methods, responses to unexpected results, and the rating of the reliability of the CB quality tests, together with expectations for standardization.Results: There were 32 respondents to the survey, of whom 28 responded to the more detailed questionnaire about viability methods. Overall, responses indicated that various stains were used among the laboratories, and when multiple sites used the same viability stain the methods differed. The majority of the respondents were in favor of standardizing the viability testing methods. A wide variety of preferences were communicated about how to standardize the method, but a majority did advocate the use of 7-aminoactinomycin D (7-AAD) with flow cytometry.Conclusions: The survey results revealed a variety of tests and inconsistent interlaboratory practices for performing the viability assay. Flow cytometry with a 7-AAD dye was suggested as a first step toward standardization. [ABSTRACT FROM AUTHOR]

Background: Cryopreserved donor lymphocyte infusion (DLI) products are manufactured and administered to treat relapse after allogeneic hematopoietic stem cell transplantation. Reported clinical responses to DLIs vary broadly, even within the same group of patients. While there is an implicit recognition of the fact that different manufacturing protocols may have specific effects on different cell types, cryopreservation protocols are frequently derived from our experience in the cryopreservation of stem cell products and do not account for the heterogeneous functional nature of DLI T-cell populations. Here, we report the results of a prospective, multicenter trial on the effect of four different cryopreservation solutions that were used to freeze DLIs compared to control DLIs that were refrigerated overnight.Study Design and Methods: Cryopreserved postthawed and refrigerated specimens were analyzed side by side for their T-cell subpopulation content and viability, as well as T-cell proliferation, cytokine secretion, and cytotoxic activities.Results: This study indicates that "homemade" 10% dimethyl sulfoxide (DMSO) results in reduced viability of different CD4+ T-cell populations, including T-helper, T-cytotoxic, and T-regulatory populations, and a decrease in their proliferative and cytotoxic response to immunologically relevant stimuli, while the use of solutions containing 5% DMSO with intracellular-like cryoprotectant stabilizers maintains T-cell function at levels similar to refrigerated control samples.Conclusion: This study has important implications in determining the best cryoprotectant solution for specific clinical applications in allogeneic immunotherapy. [ABSTRACT FROM AUTHOR]