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Optimization of Blood Handling and Peripheral Blood Mononuclear Cell Cryopreservation of Low Cell Number Samples

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 me...

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Autores principales: Hope, Christopher M., Huynh, Dao, Wong, Ying Ying, Oakey, Helena, Perkins, Griffith Boord, Nguyen, Trung, Binkowski, Sabrina, Bui, Minh, Choo, Ace Y. L., Gibson, Emily, Huang, Dexing, Kim, Ki Wook, Ngui, Katrina, Rawlinson, William D., Sadlon, Timothy, Couper, Jennifer J., Penno, Megan A. S., Barry, Simon C.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8431655/
https://www.ncbi.nlm.nih.gov/pubmed/34502038
http://dx.doi.org/10.3390/ijms22179129
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author Hope, Christopher M.
Huynh, Dao
Wong, Ying Ying
Oakey, Helena
Perkins, Griffith Boord
Nguyen, Trung
Binkowski, Sabrina
Bui, Minh
Choo, Ace Y. L.
Gibson, Emily
Huang, Dexing
Kim, Ki Wook
Ngui, Katrina
Rawlinson, William D.
Sadlon, Timothy
Couper, Jennifer J.
Penno, Megan A. S.
Barry, Simon C.
author_facet Hope, Christopher M.
Huynh, Dao
Wong, Ying Ying
Oakey, Helena
Perkins, Griffith Boord
Nguyen, Trung
Binkowski, Sabrina
Bui, Minh
Choo, Ace Y. L.
Gibson, Emily
Huang, Dexing
Kim, Ki Wook
Ngui, Katrina
Rawlinson, William D.
Sadlon, Timothy
Couper, Jennifer J.
Penno, Megan A. S.
Barry, Simon C.
author_sort Hope, Christopher M.
collection PubMed
description 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.
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spelling pubmed-84316552021-09-11 Optimization of Blood Handling and Peripheral Blood Mononuclear Cell Cryopreservation of Low Cell Number Samples Hope, Christopher M. Huynh, Dao Wong, Ying Ying Oakey, Helena Perkins, Griffith Boord Nguyen, Trung Binkowski, Sabrina Bui, Minh Choo, Ace Y. L. Gibson, Emily Huang, Dexing Kim, Ki Wook Ngui, Katrina Rawlinson, William D. Sadlon, Timothy Couper, Jennifer J. Penno, Megan A. S. Barry, Simon C. Int J Mol Sci Article 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. MDPI 2021-08-24 /pmc/articles/PMC8431655/ /pubmed/34502038 http://dx.doi.org/10.3390/ijms22179129 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Hope, Christopher M.
Huynh, Dao
Wong, Ying Ying
Oakey, Helena
Perkins, Griffith Boord
Nguyen, Trung
Binkowski, Sabrina
Bui, Minh
Choo, Ace Y. L.
Gibson, Emily
Huang, Dexing
Kim, Ki Wook
Ngui, Katrina
Rawlinson, William D.
Sadlon, Timothy
Couper, Jennifer J.
Penno, Megan A. S.
Barry, Simon C.
Optimization of Blood Handling and Peripheral Blood Mononuclear Cell Cryopreservation of Low Cell Number Samples
title Optimization of Blood Handling and Peripheral Blood Mononuclear Cell Cryopreservation of Low Cell Number Samples
title_full Optimization of Blood Handling and Peripheral Blood Mononuclear Cell Cryopreservation of Low Cell Number Samples
title_fullStr Optimization of Blood Handling and Peripheral Blood Mononuclear Cell Cryopreservation of Low Cell Number Samples
title_full_unstemmed Optimization of Blood Handling and Peripheral Blood Mononuclear Cell Cryopreservation of Low Cell Number Samples
title_short Optimization of Blood Handling and Peripheral Blood Mononuclear Cell Cryopreservation of Low Cell Number Samples
title_sort optimization of blood handling and peripheral blood mononuclear cell cryopreservation of low cell number samples
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8431655/
https://www.ncbi.nlm.nih.gov/pubmed/34502038
http://dx.doi.org/10.3390/ijms22179129
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