Performance of the Panleucogating protocol for CD4+ T cell enumeration in an HIV dedicated laboratory facility in Barbados

Abstract

Objective: To compare the Panleucogating (PLG) protocol with the routinely used four-color protocol for CD4+ T cell count enumeration.

Design and methods: One hundred fifty-three blood samples were randomly selected from samples received at the National HIV Laboratory for routine immunological monitoring. Samples were prepared using Coulter CYTO-STAT tetraCHROME monoclonal antibodies and FlowCARE PLG CD4 reagent for four-color and PLG, respectively, and analyzed on the Beckman Coulter EPICS XL flow cytometer. The PLG protocol used a sequential gating strategy where CD4+ T cells were identified using side scatter properties of cells and CD45 staining. The four-color protocol used CD45 and CD3 to identify CD4+ T cells.

Results: Absolute CD4+ T cell counts and percentages ranged from 4 to 1,285 cells/microL and 0.9 to 46.7%, respectively. Linear regression analyses revealed good correlation of PLG with the four-color protocol (absolute counts, R2 = 0.95; percentages, R2 = 0.98) over the entire range including the clinically relevant range. Bland Altman statistics revealed no bias for CD4 counts <500 cells/microL and a slight underestimation by PLG for counts >500 cells/microL (Bias = -32.7 cells/microL; 95% agreement limits = -151.3- +86.0). CD4+ T cell percentages were the similar over the entire range (Bias = 0.6%; 95% agreement limits = -1.97 +/- 3.18).

Conclusions: PLG is an accurate method for enumerating CD4+ T cells and has resulted in major cost savings to the Government of Barbados. This has implications for the sustainability of the National HIV containment program in Barbados and the other resource limited Caribbean countries. The PLG technique is now being routinely used in Barbados.