Abstracts (complete list) - Wissenschaft Online

Rainer Wurth, Angelika Bold, Thomas Keller, Ulrike Trahorsch, Peter Voigt, Stefan
Schubert, Ulrich Sack
Evaluation and validation of a manual low-cost assay for the
monitoring of CD4 counts in HIV-infected individuals in non-
OECD countries
Background: The CD4+ T-cell count is considered to be the best surrogate marker for
monitoring the clinical course of infection with HIV. Flow cytometry, as the standard
reference method for the enumeration of CD4+ T-cells, requires expensive equipment
and well-trained technicians, thus preventing its widespread use in developing
countries.
Objective: In order to make this important surrogate marker available to more patients
in non-OECD countries, we have modified a commercially available density-based cell
preparation assay to make it applicable for low-cost cell enumeration.
Methods: 1.) For evaluation (step 1), whole venous blood taken from 25 HIV-patients
as well as 29 healthy blood donors was incubated with a cocktail of bi-specific
tetrameric antibody complexes which crosslink unwanted nucleated cells (NC) to red
blood cells (RBC) by forming RBC rosettes around targeted NC. After the following
gradient centrifugation over a density medium, the enriched CD4+ T-cells were
harvested and then counted on a haemocytometre using a light optical microscope. In
parallel, the CD4 count of each sample was assessed by flow cytometry. 2.) For
validation (step 2), this method was performed in blind quintuplicates on 12 HIV+ blood
samples according to the FDA guidelines ICH-Q2A and ICH-Q2B.
Results: Association of the introduced method (modified negative selection, MNS) with
the reference method (flow cytometry, FCM) is given by regression models for both
steps:
Step 1: slope = 1.091, intercept = -46.5; (95% confidence intervall [CI] = 0.948 to
1.240). Step 2: slope = 1.074, intercept = -38.3 (CI = 0.971 to 1.200 (step 2). The
imprecision of MNS assessed during step 2 was 21.2% (intra-serial) and 18.8% (interserial).
The cost of the examination of one blood sample is $0.30 - $0.50 per sample.
Conclusion: The results suggest that the MNS-method, at is capable of providing an
approximate CD4 count. This would enable a physician attending to an HIV+ patient to
decide whether or not to start HAART or OI prophylaxis. At a cost of $0.30 - $0.50, it is
affordable to patients living in resource-restrained areas. The technique has the
potential to deliver an accurate, precise, low-cost test to monitor the status of HIV+
patients, especially in small laboratories in non-OECD countries.
Keywords: CD4 count; HIV-infection; HIV-monitoring; RosetteSepTM; low-cost
assay; non-OECD countries