BACKGROUND:Blood transfusion is the most common hospital procedure performed in the United States. While inadequate physician transfusion medicine knowledge may lead to inappropriate practice, such an educational deficit has not been investigated on an international scale using a validated assessment tool. Identifying specific deficiencies is critical for developing curricula to improve patient care. STUDY DESIGN AND METHODS:Rasch analysis, a method used in high-stakes testing, was used to validate an assessment tool consisting of a 23-question survey and a 20-question examination. The assessment tool was administered to internal medicine residents to determine prior training, attitudes, perceived ability, and actual knowledge related to transfusion medicine. RESULTS:A total of 474 residents at 23 programs in nine countries completed the examination. The overall mean score of correct responses was 45.7% (site range, 32%-56%). The mean score for Postgraduate Year (PGY)1 (43.9%) was significantly lower than for PGY3 (47.1%) and PGY4 (50.6%) residents. Although 89% of residents had participated in obtaining informed consent from a patient for transfusion, residents scored poorly (<25% correct) on questions related to transfusion reactions. The majority of residents (65%) would find additional transfusion medicine training "very" or "extremely" helpful. CONCLUSION:Internationally, internal medicine residents have poor transfusion medicine knowledge and would welcome additional training. The especially limited knowledge of transfusion reactions suggests an initial area for focused training. This study not only represents the largest international assessment of transfusion medicine knowledge, but also serves as a model for rigorous, collaborative research in medical education.

BACKGROUND AIMS:Wide acceptance of the colony-forming unit (CFU) assay as a reliable potency test for stem cell products is hindered by poor inter-laboratory reproducibility. The goal of this study was to ascertain current laboratory practices for performing the CFU assay with an eye towards identifying practices that could be standardized to improve overall reproducibility. METHODS:A survey to evaluate current laboratory practices for performing CFU assays was designed and internationally distributed. RESULTS:There were 105 respondents to the survey, of whom 68% performed CFU assays. Most survey recipients specified that an automated rather than a manual cell count was performed on pre-diluted aliquots of stem cell products. Viability testing methods employed various stains, and when multiple sites used the same viability stain, the methods differed. Cell phenotype used to prepare working cell suspensions for inoculating the CFU assay differed among sites. Most respondents scored CFU assays at 14-16 days of incubation, but culture plates were read with various microscopes. Of 57 respondents, 42% had not performed a validation study or established assay linearity. Only 63% of laboratories had criteria for determining if a plate was overgrown with colonies. CONCLUSIONS:Survey results revealed inconsistent inter-laboratory practices for performing the CFU assay. The relatively low number of centers with validated CFU assays raises concerns about assay accuracy and emphasizes a need to establish central standards. The survey results shed light on numerous steps of the methodology that could be targeted for standardization across laboratories.

Objective. The aim of the present study was to correlate new periprocedural diffusion-weighted imaging (DWI) lesions during stenting of supra-aortal arteries with the level of platelet inhibition using point-of-care analysis. Background. Cardiological studies have shown that patients undergoing coronary PTA have a significantly elevated risk of severe thrombotic complications if patients show insufficient inhibition of platelet function. Methods. From August 2008 to June 2009, 44 patients with an indication of supra-aortal angioplasty and/or stenting were prospectively enrolled. Platelet reactivity was tested using a Multiplate device (Dynabyte). These patients underwent MRI before and after the intervention to determine the prevalence of new DWI lesions. The primary endpoint was the prevalence of DWI lesions; the secondary endpoint was clinical status until discharge from hospital. Results. There was no significant relationship between the primary endpoint and the degree of platelet function. Patients with high platelet reactivity showed the same amount of periprocedural complications as patients with sufficient inhibition of platelets. Conclusions. Clopidogrel did not have a protective effect on periprocedural complications, nor did it decrease the number of silent DWI lesions after the procedure. The predescribed strong relationship between high platelet reactivity and early post-procedural adverse events was not observed in our cohort.

Delayed-type hypersensitivity reactions (DTHRs) after subcutaneous application of unfractionated heparins or low-molecular-weight heparins are not uncommon. Standard allergological testing usually includes intracutaneous skin tests and patch testing of different heparins, heparinoids, and thrombin inhibitors followed by subcutaneous and/or intravenous challenge with skin test-negative drugs. We present data from a single-center case series of 15 patients with DTHR after low-molecular-weight heparin administration. Intracutaneous testing that can be considered as gold standard identified the suspicious elicitor in 11 (73.4%) of 15 of the patients. Patch testing was positive in 5 (33.4%) of 15 of the patients and was only positive in patients who were also reacting in the intradermal testing. Intravenous challenge with heparin sodium was performed in 10 of 15 patients and was well tolerated in all cases, despite prior positive intracutaneous tests with the same substance. Intracutaneous documentation of DTHR was not an adequate predictor of intravenous challenge.

BACKGROUND: Clinical transplant outcome with umbilical cord blood (UCB) as source of hematopoietic progenitor cells (HPCs) is, among other factors, determined by the total number of viable nucleated cells and/or CD34+ cells in the unit. Quantitative and qualitative losses by processing and cryopreservation and by thawing and washing before transfusion may occur, however. Another reason for a discrepancy between the number of cells in the unit released by the cord blood bank and found in the transplant center may be technical differences in cell counting methods between the two sites.STUDY DESIGN AND METHODS: With the collaborative group for Biomedical Excellence for Safer Transfusion (BEST), an interlaboratory exercise was conducted among nine sites for thawed UCB variables: total nucleated cells, CD34+ cells, viability, and HPC cultures. Three frozen UCB samples were shipped, with instructions for thawing, counting, and HPC plating.RESULTS: Unexpectedly samples arrived at all nine receiving centers without detectable hematopoietic progenitor colony-forming cells. Nevertheless, wide interlaboratory ranges for viability were obtained. The proportion of viable cells was found higher with manual methods, but all viability assays used in the study overestimated functional progenitor cells.CONCLUSIONS: The results underscore the complexity of evaluation of frozen-thawed cord blood cells and the need for standardization of assessment.

Mesenchymal stem cells (MSCs) represent a promising tool for new clinical concepts in supporting cellular therapy. Bone marrow (BM) was the first source reported to contain MSCs. However, for clinical use, BM may be detrimental due to the highly invasive donation procedure and the decline in MSC number and differentiation potential with increasing age. More recently, umbilical cord blood (UCB), attainable by a less invasive method, was introduced as an alternative source for MSCs. Another promising source is adipose tissue (AT). We compared MSCs derived from these sources regarding morphology, the success rate of isolating MSCs, colony frequency, expansion potential, multiple differentiation capacity, and immune phenotype. No significant differences concerning the morphology and immune phenotype of the MSCs derived from these sources were obvious. Differences could be observed concerning the success rate of isolating MSCs, which was 100% for BM and AT, but only 63% for UCB. The colony frequency was lowest in UCB, whereas it was highest in AT. However, UCB-MSCs could be cultured longest and showed the highest proliferation capacity, whereas BM-MSCs possessed the shortest culture period and the lowest proliferation capacity. Most strikingly, UCB-MSCs showed no adipogenic differentiation capacity, in contrast to BM- and AT-MSCs. Both UCB and AT are attractive alternatives to BM in isolating MSC: AT as it contains MSCs at the highest frequency and UCB as it seems to be expandable to higher numbers.

Multipotent mesenchymal stem cells (MSCs) derived from human umbilical cord blood (UCB) represent promising candidates for the development of future strategies in cellular therapy. To create a comprehensive protein expression profile for UCB-MSCs, one UCB unit from a full-term delivery was isolated from the unborn placenta, transferred into culture, and their whole-cell protein fraction was subjected to two-dimensional electrophoresis (2-DE). Unambiguous protein identification was achieved with peptide mass fingerprinting matrix-assisted laser desorption/ionization - time of flight - mass spectrometry (MALDI-TOF-MS), peptide sequencing (MALDI LIFT-TOF/TOF MS), as well as gel-matching with previously identified databases. In overall five replicate 2-DE runs, a total of 2037 +/- 437 protein spots were detected of which 205 were identified representing 145 different proteins and 60 isoforms or post-translational modifications. The identified proteins could be grouped into several functional categories, such as metabolism, folding, cytoskeleton, transcription, signal transduction, protein degradation, detoxification, vesicle/protein transport, cell cycle regulation, apoptosis, and calcium homeostasis. The acquired proteome map of nondifferentiated UCB-MSCs is a useful inventory which facilitates the identification of the normal proteomic pattern as well as its changes due to activated or suppressed pathways of cytosolic signal transduction which occur during proliferation, differentiation, or other experimental conditions.

Background and Objectives The INTERCEPT Blood System for Platelets utilizes amotosalen-HCl (S-59) in combination with ultraviolet A (UVA) light to inactivate viruses, bacteria, protozoa and leucocytes that may contaminate platelet concentrates (PCs). To facilitate implementation of this technique into routine blood bank manufacturing procedures, this study evaluated the impact of different time settings of photochemical treatment on in vitro platelet function.Materials and Methods Platelets derived from apheresis (6.5-7.0 x 10(11) platelets) were resuspended in 240 ml of autologous plasma and 360 ml of platelet additive solution (PAS III) and split into two equal-sized PC units. Whereas one unit was not treated, the other was treated with 150 muM amotosalen and 3 J/cm(2) UVA light followed by a compound adsorption device (CAD) step for reduction of residual amotosalen and photoproducts. In a first series of experiments (arm A, n = 7), PC units were photochemically treated after an overnight storage period of 16-23 h followed by a CAD step of 4 h. In a second series (arm B, n = 8), photochemical treatment occurred after a short storage time of 4 h with a subsequent CAD step of 16 h. Platelet function was evaluated by assaying blood gas analysis, glucose and lactate concentration, lactate dehydrogenase (LDH), hypotonic shock response (HSR) and the expression of CD62p, over a period of 7 days.Results Neither of the photochemical treatment procedures showed differences for pH, pCO(2), pO(2), HCO3, glucose consumption or platelet activation until the end of day 7. Increased lactate values detected for the treated units of arm A at the end of the storage period were independent from the PCT time setting.Conclusions Photochemical pathogen inactivation with different initial resting periods between 4 and 23 h, and different CAD steps of 4 and 16 h, had no influence on the platelet in vitro function during 7 days of storage.

BACKGROUND: The OrbiSac device, which was developed to automate the manufacture of buffy-coat PLT concentrates (BC-PCs), was evaluated.STUDY DESIGN AND METHODS: In-vitro characteristics of BC-PC preparations using the OrbiSac device were compared with manually prepared BC-PCs. For standard processing (Std-PC, n = 20), four BC-PCs were pooled using 300 mL of PLT AS (PAS) followed by soft-spin centrifugation and WBC filtration. The OrbiSac pre-paration (OS-PC, n = 20) was performed by automated pooling of four BC-PCs with 300 mL PAS followed by centrifugation and inline WBC filtration. All PCs were stored at 22degreesC. Samples were withdrawn on Day 1, 5, and 7 evaluating PTL count, blood gas analysis, glucose, lactate, LDH, beta-thromboglobulin, hypotonic shock response, and CD62p expression.RESULTS: A PLT content of 3.1 +/- 0.4 x 10(11) (OS-PCs) versus 2.7 +/- 0.5 x 10(11) (Std-PCs, p < 0.05) was found. A CV of 19 percent (Std-PC) versus 14 percent (OS-PC) suggests more standardization in the OS group. At Day 7, the Std-PCs versus OS-PCs showed a glucose consumption of 1.03 +/- 0.32 mumol per 10(9) PLT versus 0.75 +/- 0.25 mumol per 10(9) PLT (p < 0.001), and a lactate production of 1.50 +/- 0.86 mumol per 10(9) versus 1.11 +/- 0.61 mumol per 10(9) (p < 0.001). The pH (7.00 +/- 0.19 vs. 7.23 +/- 0.06; p < 0.001), pO(2) (45.3 +/- 18 vs. 31.3 +/- 10.4 mmHg; p < 0.01), and HCO3 levels (4.91 +/- 1.49 vs. 7.14 +/- 0.95 mmol/L; p < 0.001) suggest a slightly better aerobic metabolism within the OS group. Only small differences in CD62p expression was observed (37.3 +/- 12.9% Std-PC vs. 44.8 +/- 6.6% OS-PC; p < 0.05).CONCLUSION: The OrbiSac device allows an improved PLT yield without affecting PLT in-vitro characteristics and may enable an improved consistency in product volume and yield.

Platelets are rich in growth factors and cytokines that are responsible for the induction of wound healing mechanisms and tissue regeneration. Therefore, the potential osteogenic effect of local platelet administration isused to improve future implant sites in oral surgery. We established and evaluated an autologous procedure according to national regulation and guidelines to prepare concentrated platelet-rich plasma (cPRP) for clinical application in bone regeneration. Overall, a 17-fold increase in platelet concentrations was achieved in the cPRP compared to the patients' whole bloods. Platelet counts correlated with increased levels of TGF-β1 within the cPRP. The method has proven to be suitable for blood services to support the needs for treatments with platelets in oral surgery.

Background: By inhibiting nucleic acids, photochemical treatment (PCT) using amotosalen hydrochloride (S-59) in combination with UVA light provides a new method for the inactivation of pathogens and leukocytes, potential contaminants of platelet concentrates (PCs). Material and Methods: We evaluated the effect of PCT on function and viability of PCs derived from apheresis over a storage period of 5 days. Double-dose PCs containing 6.5–7.0 × 1011 platelets (n = 8) were collected in 600 ml of approximately 35% autologous plasma and 65% platelet additive solution. Each collection was split into two equal-sized portions for paired analysis. One day after collection, test PCs were mixed with 15 ml of 3 mmol/l S-59 and illuminated with UVA light for 3 min, followed by a 6- to 8-hour incubation in a compound adsorption device (CAD) for the reduction of residual S-59 and unbound photoproducts. In vitro platelet function measurements included platelet count, pH, lactate dehydrogenase (LDH), β-thromboglobulin (β-TG), hypotonic shock response (HSR), extent of shape change (ESC) and the expression of p-selectin. Platelet morphology and ultrastructure were analyzed by light and electron microscopy (EM). Results: We found no statistically significant differences on day 5 of the storage between the control and test platelets for pH, platelet count, p-selectin, ESC, HSR, morphology, and the percentage of lysed platelets analyzed by EM. The level of β-TG was reduced in the test units by the CAD step. Significant differences (p < 0.05) were observed for LDH and HSR immediately after PCT, indicating a slight platelet activation. Until the end of the storage period, LDH increased further in control and test platelets, but the quantitative difference between both products did not vary upon further storage. The effect for HSR was transient as no differences between treated and untreated PCs were detected at the end of the storage period. Conclusion: These in vitro results indicate that PCT with amotosalen and UVA light is applicable for split double-dose platelets derived from apheresis as it leads only to a transient activation of platelets immediately after the procedure, without affecting the function and structure of PCs over the 5-day storage period.

s 4 228 Advances in Transfusion Safety – 2001, June 7 to 8, 2001 Guest Editor: Seitz, R., Langen