Delayed isolation and cryopreservation have significant effects on the yield and proliferation of feline peripheral blood mononuclear cells.

Abstract

Objective: To determine the temporal effects of feline peripheral blood mononuclear cell (PBMC) isolation delay and cryopreservation over multiple time points on cell yields and PBMC proliferation.

Methods: PBMCs from whole feline blood were isolated immediately upon blood collection or stored at 4 °C for 72 hours before isolation. Peripheral blood mononuclear cell proliferation was assessed on freshly isolated PBMCs and after 2-week, 2-month, 6-month, and 12-month cryopreservation time points. Peripheral blood mononuclear cell recovery yield was recorded at every time point.

Results: Delaying PBMC isolation by 72 hours leads to an approximately 50% reduction in PBMC count and a significant increase in PBMC proliferation (43.1%; n = 6 donors) compared to freshly isolated PBMCs (25.4%; n = 13 donors; 17.7 Effect size; 95% CI, 0.9827 to 34.42 Range). Cryopreserving freshly isolated PBMCs results in approximately 50% cell loss upon thawing. The 2-month cryopreservation time point exhibited significantly higher viable cell recovery compared to the 12-month time point (20.6 Effect size; 95% CI, 3.98 to 37.2 Range). The proliferation capacity of PBMCs was, on average, 16.6% across all cryopreservation time points, with no significant difference between fresh and cryopreserved samples.

Conclusions: Delayed PBMC isolation reduces PBMC yield and enhances proliferation. Cryopreservation did not significantly impact PBMC proliferation across time points, whereas viable cell yields were reduced by an additional 50% after extended cryopreservation.

Clinical relevance: This study provides critical insights for clinical studies involving feline PBMCs, highlighting the importance of timely isolation and the possibility of performing PBMC proliferation studies following extended cryopreservation, albeit on account of reduced recovery yields.