Choice of medium affects PBMC quantification, cell size, and downstream respiratory analysis

Ida Bager Christensen; Lucas Ribas; Maria Mosshammer; Marie Louise Abrahamsen; Michael Kühl; Steen Larsen; Flemming Dela; Linn Gillberg

doi:10.1016/j.mito.2024.101890

Choice of medium affects PBMC quantification, cell size, and downstream respiratory analysis

Ida Bager Christensen
, Lucas Ribas
, Maria Mosshammer
, Marie Louise Abrahamsen
, Michael Kühl
, Steen Larsen
, Flemming Dela
, Linn Gillberg^* (Corresponding Author)

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

4 Citations (Scopus)

13 Downloads (Pure)

Abstract

High-resolution respirometry (HRR) can assess peripheral blood mononuclear cell (PBMC) bioenergetics, but no standardized medium for PBMC preparation and HRR analysis exist. Here, we study the effect of four different media (MiR05, PBS, RPMI, Plasmax) on the count, size, and HRR (Oxygraph-O2k) of intact PBMCs. Remarkably, the cell count was 21 % higher when PBMCs were resuspended in MiR05 than in PBS or Plasmax, causing O₂ flux underestimation during HRR due to inherent adjustments. Moreover, smaller cell size and cell aggregation was observed in MiR05. Based on our findings, we propose that Plasmax, PBS or RPMI is more suitable than MiR05 for HRR of intact PBMCs. We provide oxygen solubility factors for Plasmax and PBS and encourage further optimization of a standardized HRR protocol for intact PBMCs.

Original language	English
Article number	101890
Number of pages	7
Journal	Mitochondrion
Volume	77
DOIs	https://doi.org/10.1016/j.mito.2024.101890
Publication status	Published - Jul 2024
Externally published	Yes

Keywords*

Cell count
High-resolution respirometry
Medium
Oxygen solubility
Peripheral blood mononuclear cells

Field of Science*

1.6 Biological sciences
3.1 Basic medicine

Publication Type*

1.1. Scientific article indexed in Web of Science and/or Scopus database

Access to Document

10.1016/j.mito.2024.101890Licence: CC BY

Choice of medium affects PBMC quantificationFinal published version, 1.64 MBLicence: CC BY

Cite this

@article{46844adb92824384994ac1835bccbc17,

title = "Choice of medium affects PBMC quantification, cell size, and downstream respiratory analysis",

abstract = "High-resolution respirometry (HRR) can assess peripheral blood mononuclear cell (PBMC) bioenergetics, but no standardized medium for PBMC preparation and HRR analysis exist. Here, we study the effect of four different media (MiR05, PBS, RPMI, Plasmax) on the count, size, and HRR (Oxygraph-O2k) of intact PBMCs. Remarkably, the cell count was 21 \% higher when PBMCs were resuspended in MiR05 than in PBS or Plasmax, causing O2 flux underestimation during HRR due to inherent adjustments. Moreover, smaller cell size and cell aggregation was observed in MiR05. Based on our findings, we propose that Plasmax, PBS or RPMI is more suitable than MiR05 for HRR of intact PBMCs. We provide oxygen solubility factors for Plasmax and PBS and encourage further optimization of a standardized HRR protocol for intact PBMCs.",

keywords = "Cell count, High-resolution respirometry, Medium, Oxygen solubility, Peripheral blood mononuclear cells",

author = "\{Bager Christensen\}, Ida and Lucas Ribas and Maria Mosshammer and Abrahamsen, \{Marie Louise\} and Michael K{\"u}hl and Steen Larsen and Flemming Dela and Linn Gillberg",

note = "Publisher Copyright: {\textcopyright} 2024 The Author(s)",

year = "2024",

month = jul,

doi = "10.1016/j.mito.2024.101890",

language = "English",

volume = "77",

journal = "Mitochondrion",

issn = "1567-7249",

publisher = "Elsevier B.V.",

}

TY - JOUR

T1 - Choice of medium affects PBMC quantification, cell size, and downstream respiratory analysis

AU - Bager Christensen, Ida

AU - Ribas, Lucas

AU - Mosshammer, Maria

AU - Abrahamsen, Marie Louise

AU - Kühl, Michael

AU - Larsen, Steen

AU - Dela, Flemming

AU - Gillberg, Linn

PY - 2024/7

Y1 - 2024/7

N2 - High-resolution respirometry (HRR) can assess peripheral blood mononuclear cell (PBMC) bioenergetics, but no standardized medium for PBMC preparation and HRR analysis exist. Here, we study the effect of four different media (MiR05, PBS, RPMI, Plasmax) on the count, size, and HRR (Oxygraph-O2k) of intact PBMCs. Remarkably, the cell count was 21 % higher when PBMCs were resuspended in MiR05 than in PBS or Plasmax, causing O2 flux underestimation during HRR due to inherent adjustments. Moreover, smaller cell size and cell aggregation was observed in MiR05. Based on our findings, we propose that Plasmax, PBS or RPMI is more suitable than MiR05 for HRR of intact PBMCs. We provide oxygen solubility factors for Plasmax and PBS and encourage further optimization of a standardized HRR protocol for intact PBMCs.

AB - High-resolution respirometry (HRR) can assess peripheral blood mononuclear cell (PBMC) bioenergetics, but no standardized medium for PBMC preparation and HRR analysis exist. Here, we study the effect of four different media (MiR05, PBS, RPMI, Plasmax) on the count, size, and HRR (Oxygraph-O2k) of intact PBMCs. Remarkably, the cell count was 21 % higher when PBMCs were resuspended in MiR05 than in PBS or Plasmax, causing O2 flux underestimation during HRR due to inherent adjustments. Moreover, smaller cell size and cell aggregation was observed in MiR05. Based on our findings, we propose that Plasmax, PBS or RPMI is more suitable than MiR05 for HRR of intact PBMCs. We provide oxygen solubility factors for Plasmax and PBS and encourage further optimization of a standardized HRR protocol for intact PBMCs.

KW - Cell count

KW - High-resolution respirometry

KW - Medium

KW - Oxygen solubility

KW - Peripheral blood mononuclear cells

UR - https://www.scopus.com/pages/publications/85193636163

U2 - 10.1016/j.mito.2024.101890

DO - 10.1016/j.mito.2024.101890

M3 - Article

C2 - 38718898

AN - SCOPUS:85193636163

SN - 1567-7249

VL - 77

JO - Mitochondrion

JF - Mitochondrion

M1 - 101890

ER -

Choice of medium affects PBMC quantification, cell size, and downstream respiratory analysis

Abstract

Keywords*

Field of Science*

Publication Type*

Access to Document

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