Sample Stability and Variability of B-Cell Subsets in Blood from Healthy Subjects and Patients with Systemic Lupus Erythematosus

Objective: Characterization of peripheral leukocytes is an important aspect of monitoring the effect of immunotherapeutic interventions in systemic lupus erythematosus (SLE). We analyzed cell surface markers commonly used to assess patients with SLE, focusing on the effect of holding blood prior to processing/analysis and the relative reliability of the measurements that were conducted. Methods: Healthy volunteers (HV; n = 20) and patients with SLE (n = 42) were studied. Whole blood was collected for flow cytometric analysis on days 1, 8, 15, 105, 195, 285, and 375 and held overnight for analysis. A subset of samples was additionally analyzed on the day of collection. Results: Variability arising from overnight storage of whole blood was found to be within 20% for most lymphocyte subsets. There was greater between rather than within subject variability over a 1-year period. As anticipated, the data showed higher CD38 and lower CD19 densities on B cells from patients with SLE compared to HV. Although a higher percentage of cells with markers of plasmablasts/cells were observed in the blood of patients with SLE relative to HV, these measurements were found to be among the least reliable (i.e., most variable). Conclusions: This study provides technical perspectives for those conducting immunophenotypic analyses of B-cells in patients with SLE. We envision that our data, which addresses sample stability issues and presents a method to describe the relative reliability of one measure over another, holds value for clinical assessments of B-cells in SLE and the evaluation of investigational agents designed to modify the B-cell compartment. (C) 2009 Clinical Cytometry Society