Effect of delayed isolation of peripheral blood mononuclear cells on cell viability and functionality

Lehle S, Völkl S, Seitz K, Goossens C, Emons J, Rübner M, Uhrig S, Ziegler P, Theuser AK, Beckmann M, Fasching P, Hübner H (2025)

Publication Type: Journal article

Publication year: 2025

Journal

Bmc Immunology BioMed Central

Book Volume: 26

Article Number: 21

Journal Issue: 1

DOI: 10.1186/s12865-025-00701-y

Abstract

Background: Peripheral blood mononuclear cells (PBMCs) are valuable biomarkers, providing crucial insights into the patients’ immune system. Reliable biobanking of PBMCs is essential to minimize heterogeneity. In multicenter trials, blood sample transportation to central laboratories can increase the time between blood collection and PBMC isolation. This study evaluated the effect of prolonged blood hold time on PBMC viability and cytotoxicity. Methods: From July 2021 to May 2023, 104 patients with early HER2-positive breast cancer were enrolled in the NeoOn trial, of whom 49 patients were included in this subproject. PBMCs were isolated ≤ 6 hours (h) or ≥ 20 h after blood collection. PBMC yield and viability were determined using the LUNA-II Automated Cell Counter. Flow cytometry was used to quantify in vitro cytotoxicity, the percentage of natural killer (NK) and T cells, as well as apoptotic and necrotic cells. Results: Isolating PBMCs ≥ 20 h resulted in a higher cell yield, but lower NK cell viability compared to PBMCs ≤ 6 h. PBMCs ≥ 20 h were less robust to thawing and showed higher loss during recovery. Compared to PMBCS ≤ 6 h, PBMCs ≥ 20 h exhibited lower antibody-mediated cytotoxicity (p ≤ 0.0001) and antibody-dependent phagocytosis (p < 0.0051). While the percentage of T and NK cells and the T cell viability remained unaffected by hold time, the percentage of apoptotic NK cells was higher for PBMCs ≥ 20 h (41.0 ± 12.9% vs. 23.8 ± 13.4%; p = 0.0364). Conclusions: Extended blood storage time caused increased apoptosis and necrosis of NK cells, adversely affecting PBMC quality and reducing NK cell related functionality. Hence, blood hold time should be minimized to maintain PBMC integrity and NK cell functionality for in vitro biomarker assays. Trial registration: Trial registration number: EudraCT 2020-001943-21. Date of registration: December 29th 2020

Authors with CRIS profile

Sarah Lehle Department of Obstetrics and Gynaecology Simon Völkl Department of Medicine 5 – Haematology and Oncology Katharina Seitz Department of Obstetrics and Gynaecology Chloe Goossens Department of Obstetrics and Gynaecology Julius Emons Department of Obstetrics and Gynaecology Matthias Rübner Department of Obstetrics and Gynaecology Sabrina Uhrig Department of Obstetrics and Gynaecology Matthias Beckmann Lehrstuhl für Geburtshilfe und Frauenheilkunde Peter Fasching Professur für Translationale Frauenheilkunde und Geburtshilfe Hanna Hübner Department of Obstetrics and Gynaecology

Involved external institutions

Institut für Frauengesundheit GmbH DE Germany (DE)

How to cite

APA:

Lehle, S., Völkl, S., Seitz, K., Goossens, C., Emons, J., Rübner, M.,... Hübner, H. (2025). Effect of delayed isolation of peripheral blood mononuclear cells on cell viability and functionality. Bmc Immunology, 26(1). https://doi.org/10.1186/s12865-025-00701-y

MLA:

Lehle, Sarah, et al. "Effect of delayed isolation of peripheral blood mononuclear cells on cell viability and functionality." Bmc Immunology 26.1 (2025).

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