Improving Reliability of Immunological Assays by Defining Minimal Criteria for Cell Fitness
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Le résumé fourni par la source
Human PBMC-based assays are often used as biomarkers for the diagnosis and prognosis of disease, as well as for the prediction and tracking of response to biological therapeutics. However, the development and use of PBMC-based biomarker assays is often limited by poor reproducibility. Complex immunological assays can be further complicated by variation in cell handling before analysis, especially when using cryopreserved cells. Variation in postthaw viability is further increased if PBMC isolation and cryopreservation are done more than a few hours after collection. There is currently a lack of evidence-based standards for the minimal PBMC viability or "fitness" required to ensure the integrity and reproducibility of immune cell-based assays. In this study, we use an "induced fail" approach to examine the effect of thawed human PBMC fitness on four flow cytometry-based assays. We found that cell permeability-based viability stains at the time of thawing did not accurately quantify cell fitness, whereas a combined measurement of metabolic activity and early apoptosis markers did. Investigation of the impact of different types and levels of damage on PBMC-based assays revealed that only when cells were >60-70% live and apoptosis negative did biomarker values cease to be determined by cell fitness rather than the inherent biology of the cells. These data show that, to reproducibly measure immunological biomarkers using cryopreserved PBMCs, minimal acceptable standards for cell fitness should be incorporated into the assay protocol.
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Le contrôle bibliographique ouvert
DOI retrouvé dans Crossref DOI retrouvé ; titre concordant.
- Titre Crossref
- Improving Reliability of Immunological Assays by Defining Minimal Criteria for Cell Fitness
- Date Crossref
- 01/09/2024
- Éditeur
- Oxford University Press (OUP)
- Type
- journal-article
Ce recoupement confirme des métadonnées liées au DOI. Il ne confirme ni la méthode ni les conclusions de l’étude, et il ne compte pas comme une seconde source scientifique indépendante.
Où se fait cette recherche
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University of British Columbia Department of Surgery pays non établi dans la noticeUniversité ou école supérieure
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BC Children's Hospital pays non établi dans la noticeÉtablissement de santé
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BC Children's Hospital Research Institute pays non établi dans la noticeUniversité ou école supérieure
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Montreal Heart Institute pays non établi dans la noticeOrganisme public
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McGill University Health Centre pays non établi dans la noticeÉtablissement de santé
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Université de Montréal Department of Biochemistry and Molecular Medicine pays non établi dans la noticeUniversité ou école supérieure
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School of Biomedical Engineering pays non établi dans la noticeUniversité ou école supérieure
Department of Surgery — University of British Columbia, BC Children's Hospital et BC Children's Hospital Research Institute, avec 4 autres affiliations.
Une affiliation ne permet pas de déduire la nationalité d’un auteur.