Evaluation of a CD4-aptamer for Flow Cytometry-based Enumeration of T-cell Subsets

Abstract

Introduction: Aptamers are engineered oligonucleotides that can be selected to bind with high specificity and affinity. As aptamers are cost-effective (estimated to be ~1,000-fold cheaper than antibodies) and also temperature-stable, aptamers may be useful in resource-limited settings for T-cell subset quantification. Here, we evaluated the performance characteristics of a CD4 aptamer for T-cell subset quantification. Methods: A CD4-biotinylated-streptavidin-PE aptamer provided by SomaLogic, Boulder, CO was combined with CD3-PC5, CD8-ECD and CD45-FITC antibodies in the multicolor flow cytometric assay; a CD4-A594 antibody was also added as an internal control for the CD4 aptamer. Residual patient samples were evaluated using the aptamer versus antibody reagents. Both the proportion of CD4 T cell and absolute CD4 T cell count were analyzed to evaluate the clinical performance of the aptamer reagent. Results and Conclusion: The CD4 aptamer showed comparable results to the established single platform method, which uses CYTO-STAT reagent (CD45F/CD4RD1/CD8ECD/CD3PC5) and an FC-500 flow cytometry system, with r2 >0.92 of CD4 T-cell% measurement,. Further, both the percentage of CD4 T cells and absolute T-cell count showed excellent agreement over the range of T cell proportions (8.6% to 50%) and absolute T-cell counts (150 cells/ul to 1342 cells/ul) as assessed by the CD4 aptamer as compared to an internal CD4-A594 antibody control, with r2 >0.99 and 9 cells /ul in mean difference. Our results suggest that a CD4 aptamer can perform comparably to an established CD4 antibody for CD4 T-cell subset quantitation and supports further development of aptamers as potential cost-effective reagents in resource-limited regions.