1. Immune phenotyping in a pediatric multicenter transplant study: Suitability of a preformulated dry-antibody panel system.
- Author
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Ionescu LI, Blydt-Hansen T, Foster BJ, Allen U, Birk PE, Hamiwka L, Phan V, Min S, Ivison S, Levings M, West LJ, Mital S, and Urschel S
- Subjects
- Humans, Child, Female, Male, Child, Preschool, Adolescent, Infant, Reproducibility of Results, Canada, Cryopreservation, Antibodies immunology, Antibodies blood, Phenotype, Immunophenotyping methods, Organ Transplantation, Flow Cytometry methods
- Abstract
Flow-cytometric immune phenotyping is influenced by cryopreservation and inter-laboratory variability limiting comparability in multicenter studies. We assessed a system of optimized, pre-mixed dry-antibody panel tubes requiring small amounts of whole blood for validity, reliability and challenges in a Canadian multicenter study (POSITIVE) with long-distance sample shipping, using standardized protocols. Thirty-seven children awaiting solid-organ transplant were enrolled for parallel immune-phenotyping with both validated, optimized in-house panels and the dry-antibody system. Samples were collected before, 3 and 12 months post-transplant. Quality-assurance measures and congruence of phenotypes were compared using Bland-Altman comparisons, linear regression and group comparisons. Samples showed excellent lymphocyte viability (mean 94.8 %) and recovery when processed within 30 h. Comparing staining methods, significant correlations (Spearman correlation coefficient >0.6, p < 0.05), mean difference <5 % and variation 2SD <25 % were found for natural-killer, T and B cells, including many immunologically important cell subsets (CD8+, naïve, memory CD4+ T; switched-memory, transitional B). Some subgroups (plasmablasts, CD1d+CD5hi B cells) showed weak correlations, limiting interpretation reliability. The dry-antibody system provides a reliable method for standardized analysis of many immune phenotypes after long-distance shipping when processed within 30 h, rendering the system attractive for pediatric studies due to small blood amounts required and highly standardized processing and analysis., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Seema Mital reports a relationship with Bristol Myers Squibb that includes: consulting or advisory. Seema Mital reports a relationship with Tenaya Therapeutics that includes: consulting or advisory. Lavinia Ionescu reports a relationship with Beckman Coulter Canada that includes: employment. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.)
- Published
- 2024
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