Interlaboratory assessment of a novel colony-forming unit assay: A multicenter study by the cellular team of Biomedical Excellence for Safer Transfusion (BEST) collaborative

Background: Interlaboratory scoring performances were determined using a traditional 14-day colony-forming unit (CFU) assay and a new 7-day CFU assay. Study Design and Methods: Digital images of colonies were utilized to train personnel at each site. A central laboratory inoculated methylcellulose with progenitors and sent the samples by overnight courier to participating labs for plating. Results: Colony counts from two digital images showed greater variability by novice counters (coefficients of variation [CV], 18.5 and 23.0%; n = 8) than for experienced staff (CV, 7.3 and 4.8%; n = 5). CFU assays plated immediately, 24 and 48 hours after methylcellulose inoculation displayed 39.5 CFU, 37.1 ± 10.6 (CV, 28%) and 34.8 ± 8.5 (CV, 24%) colonies for the 7-day assay and 39.5 CFU, 39.1 ± 9.9 (CV, 25%) and 37.1 ± 10.6 (CV, 28%) colonies for the 14-day assay, respectively. Overall, no significant differences in colony counts were noted between assays (p = 0.68). Also, no differences in CFU counts were seen when assays were set up immediately, 24 and 48 hours after methylcellulose inoculation (14-day p = 0.695; 7-day p = 0.632). CONCLUSION: Total CFUs obtained in 7- and 14-day CFU assays are comparable and show similar levels of interlaboratory variability. The major source of this variability is due to differences in how CFU plates are scored by individuals at different sites. UCB progenitor cells can be maintained in methylcellulose- based media at room temperature for up to 48 hours prior to transport without a significant loss in CFUs.