Low cost CD4 enumeration using generic monoclonal antibody reagents and a two‐color user‐defined MultiSET™ protocol

Background: The standard three‐tube, three‐color flow cytometric method utilizing the TriTEST™ reagents in conjunction with the MultiSET™ software commonly used in most laboratories in Thailand for CD4 enumeration is expensive and thus unavailable to most HIV‐infected patients. A more affordable method, i.e., the Pan Leuco gating protocol using only two monoclonal antibody reagents, has been described but requires the use of the CellQUEST™ software that does not have automatic gating and reporting facilities. We describe a simple protocol that utilizes a two‐color user‐defined protocol with the automated MultiSET™ software for the acquisition, analysis, and reporting of CD4 results. Methods: A two‐color user‐defined protocol was set up following instructions in the Becton Dickinson Biosciences MultiSET™ manual, adhering strictly to the information regarding the Gate and Attractor Hierarchy for analyzing various reagent combinations. This simple two‐color user‐defined MultiSET™ software was evaluated using generic monoclonal reagents in comparison with the standard TriTEST™/MultiSET™ protocol. Results: The two‐color user‐defined MultiSET™ software is easy to use. It requires only modification of the original MultiSET™ program and the results obtained are comparable with those derived from the standard TriTEST™/MultiSET™ protocol. Conclusion: The use of this easy and reliable two‐color user‐defined MultiSET™ protocol represents an affordable alternative to CD4 testing in resource‐poor settings. © 2006 International Society for Analytical Cytology