Abstract
BackgroundThe National Health Laboratory Service (NHLS) offers wide-scale CD4 testing through a network of laboratories in South Africa. A new “load and go” cytometer (Aquios CL, Beckman Coulter), developed with a PLG protocol, was validated against the predicate PLG method on the Beckman Coulter FC500 MPL/CellMek platform.MethodsRemnant routine EDTA blood CD4 reference results were compared to results from two Aquios/PLG instruments (n = 205) and a further n = 1885 samples tested to assess daily testing capacity. Reproducibility was assessed using ImmunotrolTM and patient samples with low, medium, high CD4 counts. Data was analyzed using GraphPad software for general statistics and Bland-Altman (BA) analyses. The percentage similarity (%Sim) was used to measure the level of agreement (accuracy) of the new platform versus the predicate and variance (%SimCV) reported to indicate precision of difference to predicate.Results205 samples were tested with a CD4 count range of 2–1228 cells/μl (median 365cells/μl). BA analysis revealed an overall -40.5±44.0cells/μl bias (LOA of 126.8 to 45.8cells/μl) and %Sim showing good agreement and tight precision to predicate results (94.83±5.39% with %SimCV = 5.69%). Workflow analysis (n = 1885) showed similar outcomes 94.9±8.9% (CV of 9.4%) and 120 samples/day capacity. Excellent intra-instrument reproducibility was noted (%Sim 98.7±2.8% and %SimCV of 2.8%). 5-day reproducibility using internal quality control material (Immunotrol™) showed tight precision (reported %CV of 4.69 and 7.62 for Normal and Low material respectively) and instrument stability.ConclusionThe Aquios/PLG CD4 testing platform showed clinically acceptable result reporting to existing predicate results, with good system stability and reproducibility with a slight negative but precise bias. This system can replace the faded XL cytometers in low- to medium volume CD4 testing laboratories, using the standardized testing protocol, with better staff utilization especially where technical skills are lacking. Central monitoring of on-board quality assessment data facilitates proactive maintenance and networked instrument performance monitoring.
Highlights
Flow Cytometry (FC) has been the testing platform of choice for CD4 T-lymphocyte enumeration in HIV infected patients [1,2,3] since the correlation between CD4 loss and HIV disease progression was first described [4,5,6]
The Aquios/PLG CD4 testing platform showed clinically acceptable result reporting to existing predicate results, with good system stability and reproducibility with a slight negative but precise bias
South Africa has a high burden of disease with 11.2% (6.2 million) of the local populace living with HIV in 2015 (Statistics South Africa, 2015) and an average of ~ 9.7% of HIV+ patients noted to have CD4 counts less than 100cells/μl [7]
Summary
Flow Cytometry (FC) has been the testing platform of choice for CD4 T-lymphocyte enumeration in HIV infected patients [1,2,3] since the correlation between CD4 loss and HIV disease progression was first described [4,5,6]. With the onset of the national HIV program in South Africa and the development of strategic HIV and AIDS Plans for South Africa [8,9,10], the challenges of providing CD4 enumeration as a routine test across multiple laboratories, led to the local development of a low-cost, easy-to-use, single platform, 2-color, lyse-no-wash cytometry assay, i.e. the PanLeucogate (PLG) CD4 [11,12,13] This protocol incorporated CD45 FITC and CD4 PE labeled antibodies with a gating strategy based on total white cells as reference rather than using the total lymphocyte population as reference as recommended previously [14, 15].
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