Optimization of Blood Handling and Peripheral Blood Mononuclear Cell Cryopreservation of Low Cell Number Samples
- Author(s)
- Hope, CM; Huynh, D; Wong, YY; Oakey, H; Perkins, GB; Nguyen, T; Binkowski, S; Bui, M; Choo, AYL; Gibson, E; Huang, D; Kim, KW; Ngui, K; Rawlinson, WD; Sadlon, T; Couper, JJ; Penno, MAS; Barry, SC; On Behalf Of The Endia Study Group,;
- Details
- Publication Year 2021-08-24,Volume 22,Issue #17,Page 9129
- Journal Title
- International Journal of Molecular Sciences
- Abstract
- BACKGROUND: Rural/remote blood collection can cause delays in processing, reducing PBMC number, viability, cell composition and function. To mitigate these impacts, blood was stored at 4 °C prior to processing. Viable cell number, viability, immune phenotype, and Interferon-γ (IFN-γ) release were measured. Furthermore, the lowest protective volume of cryopreservation media and cell concentration was investigated. METHODS: Blood from 10 individuals was stored for up to 10 days. Flow cytometry and IFN-γ ELISPOT were used to measure immune phenotype and function on thawed PBMC. Additionally, PBMC were cryopreserved in volumes ranging from 500 µL to 25 µL and concentration from 10 × 10(6) cells/mL to 1.67 × 10(6) cells/mL. RESULTS: PBMC viability and viable cell number significantly reduced over time compared with samples processed immediately, except when stored for 24 h at RT. Monocytes and NK cells significantly reduced over time regardless of storage temperature. Samples with >24 h of RT storage had an increased proportion in Low-Density Neutrophils and T cells compared with samples stored at 4 °C. IFN-γ release was reduced after 24 h of storage, however not in samples stored at 4 °C for >24 h. The lowest protective volume identified was 150 µL with the lowest density of 6.67 × 10(6) cells/mL. CONCLUSION: A sample delay of 24 h at RT does not impact the viability and total viable cell numbers. When long-term delays exist (>4 d) total viable cell number and cell viability losses are reduced in samples stored at 4 °C. Immune phenotype and function are slightly altered after 24 h of storage, further impacts of storage are reduced in samples stored at 4 °C.
- Publisher
- MDPI
- Keywords
- PBMC; blood handling; delay in processing; cryopreservation; cell concentration
- Research Division(s)
- Population Health And Immunity
- PubMed ID
- 34502038
- Publisher's Version
- https://doi.org/10.3390/ijms22179129
- Open Access at Publisher's Site
https://doi.org/10.3390/ijms22179129- Terms of Use/Rights Notice
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Creation Date: 2021-09-17 11:23:12
Last Modified: 2021-09-17 11:43:42