The World Health Organization (WHO) "3 × 5 program" has spurred efforts to place 3 million people on combination antiretroviral therapy (ART) for treatment of AIDS in resource-limited countries [1, 2]. This has been made possible by reduction in price of proprietary ARVs and the introduction of generic versions of these drugs. Paradoxically, the cost of CD4+ T-lymphocyte count essential for decision-making to commence ART for HIV positive adults and to monitor responses to ART remains unaffordable in most resource-limited countries. Although, some resource-limited countries that cannot access affordable CD4+ T-lymphocyte enumeration may use clinical criteria in accordance with WHO recommendations for commencing HIV seropositive individuals on ART , CD4+ T-lymphocyte count remains an important measure of immune response, and provide laboratory criteria for stopping co-trimoxazole prophylaxis.
Laboratory-based WHO recommendations for decisions to commence adult HIV-infected individuals on ART are largely based on absolute CD4+ T-lymphocyte counts. Flow cytometry is the gold standard for accurate and automated measurements of absolute T-lymphocyte subset profiles. Flow cytometric techniques are not only expensive, but require sophisticated equipment and reagents as well as highly trained personnel. Furthermore, in resource-limited countries ready access to technical support and quality assurance programs for flow cytometry are often not readily available. Non-flow cytometric methods for counting absolute CD4+ T-lymphocytes such as the manual magnetic bead Dynabead technique have also been evaluated [4, 5] but are also considered expensive, labor intensive and can only accommodate low sample throughput .
Current clinical flow cytometric methods use either dual or single platforms for enumeration of absolute T-lymphocyte subpopulations. Dual platforms rely on a combination of flow cytometric percentages and white blood cells (WBC) obtained from haematological analyzers to obtain absolute numbers of T-lymphocytes. On the other hand, single platform cytometers are dedicated flow cytometers with direct absolute counting facility. Previous studies have shown that dual platforms have a higher inter-laboratory percent coefficient of variation (%CV) of absolute CD4+ T-lymphocytes compared to single platforms . The higher %CV was attributed to use of different haematological analyzers, with no external quality assurance and internal quality control for WBC used in dual platforms and references therein.] Single platform flow cytometers may allow the efficient and accurate enumeration of T-lymphocyte subpopulations in clinical settings where large numbersof patients are receiving care .
Recently, great strides have been made in developing easy to use affordable single platform flow cytometric methods for enumerating absolute CD4+ T-lymphocytes. For example, Janossy and colleagues introduced primary CD4 gating on volumetric flow cytometry for absolute CD4+ T-lymphocyte counts without microbeads or haematology . However, this method was developed using the 'state of the art' flow cytometers (Cytoron Ortho and FACSCalibur) with the attendant capacity and costs requirements.
Recently, an increasing number of point of care less expensive methods to measure absolute CD4+ T-lymphocyte counts have been developed. These include the development and evaluation of simplified single platform flow cytometric methods using a low cost flow cytometer that can be powered from a car battery or by solar panels (Cyflow SL, Partec, Munster, Germany) by Cassens and colleagues [11, 12], modification of a commercially available 4-parameter flow cytometer, Luminex 100 (Luminex, Austin Texas, USA) to a compact portable prototype instrument that can operate with a 12-volt rechargeable battery  and Guava PCA System (Guava Technologies, Hayward, CA, USA).
The objective of the current study was to evaluate single platform Cyflow cytometry (Cyflow SL, Partec, Munster, Germany) for enumeration of absolute CD4+ T-lymphocytes among subtype C HIV-1 seropositive subjects using single platform FACSCount (Becton and Dickinson, Immunocytometry Systems, San Jose, CA, USA) as the predicate method.