A multi-laboratory evaluation of in vitro platelet assays: the tests for extent of shape change and response to hypotonic shock

BACKGROUND: There is no consensus regarding the use of specific in vitro tests for the assessment of the quality of platelet components. A literature review found that the platelet discoid shape as measured photometrically by the extent of shape change (ESC) and hypotonic shock response (HSR) correlated well with in vivo viability. The purpose of this study was to determine whether multiple research laboratories can perform the ESC and HSR assays in an accurate, reproducible manner, with acceptable sensitivity and comparable results. STUDY DESIGN AND METHODS: Eleven laboratories conducted five identical experiments, each with a different unit of platelet-rich plasma (PRP). For each experiment, 2 half-units of PRP were prepared and stored overnight: 1 half-unit at 20 to 24 degrees C in CPD (CPD-PRP) and the other at 1 to 6 degrees C with 2 mg per mL of EDTA (cold EDTA-PRP) added to produce spherical platelets with reduced HSR. Platelet suspensions having different proportions of the two PRPs were prepared and evaluated in duplicate by ESC and HSR assays, and morophologically scored by microscopy. One-way ANOVA and Duncan multiple-range tests were performed to determine significant differences in assay results for suspensions having different proportions of CPD-PRP. RESULTS: Comparable ESC (mean range: 20-28% for CPD-PRP and 1-6% for cold EDTA-PRP) and HSR (mean range: 58-81% for CPD-PRP and 12-31% for cold EDTA-PRP) measurements were obtained by nine laboratories. Duplicate testing showed high reproductibility of ESR and HSR results /in all laboratories. A 25-percent difference in the proportion of CPD-PRP (indicative of a difference of approximately 25% in the proportions of discoid and spherical platelets) was detected with a sample size of five (p<0.05) for both the ESC and HSR assays. A high correlation was found for the ESC assay and morphology score (r = 0.93, n = 345). CONCLUSION: Multiple laboratories were able to obtain comparable results with the ESC and HSR tests. They were able to show that the tests can be performed in an accurate, reproducible manner and with acceptable sensitivity.