Within- and between-subject biological variation estimates for the enumeration of lymphocyte deep immunophenotyping and monocyte subsets.

Objectives: This study aimed to deliver biological variation (BV) estimates for 25 types of lymphocyte subpopulations subjected to deep immunophenotyping (memory T/B cells, regulatory T cells, etc.) and classical, intermediate, and nonclassical monocyte subsets based on the full spectrum flow cytometry (FS-FCM) and a Biological Variation Data Critical Appraisal Checklist (BIVAC) design.

Methods: Samples were collected biweekly from 60 healthy Chinese adults over 10 consecutive two-week periods. Each sample was measured in duplicate within a single run for lymphocyte deep immunophenotyping and monocyte subset determination using FS-FCM, including the percentage (%) and absolute count (cells/μL). After trend adjustment, a Bayesian model was applied to deliver the within-subject BV (CV_I) and between-subject BV (CV_G) estimates with 95 % credibility intervals.

Results: Enumeration (% and cells/μL) for 25 types of lymphocyte deep immunophenotyping and three types of monocyte subset percentages showed considerable variability in terms of CV_I and CV_G. CV_I ranged from 4.23 to 47.47 %. Additionally, CV_G ranged between 10.32 and 101.30 %, except for CD4⁺ effector memory T cells re-expressing CD45RA. No significant differences were found between males and females for CV_I and CV_G estimates. Nevertheless, the CV_Gs of PD-1⁺ T cells (%) may be higher in females than males. Based on the desired analytical performance specification, the maximum allowable imprecision immune parameter was the CD8⁺PD-1⁺ T cell (cells/μL), with 23.7 %.

Conclusions: This is the first study delivering BV estimates for 25 types of lymphocyte subpopulations subjected to deep immunophenotyping, along with classical, intermediate, and nonclassical monocyte subsets, using FS-FCM and adhering to the BIVAC design.