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Comparative Study
. 2008 Jul 1;48(3):297-303.
doi: 10.1097/QAI.0b013e31817bbc3a.

Evaluation of Dynabeads and Cytospheres compared with flow cytometry to enumerate CD4+ T cells in HIV-infected Ugandans on antiretroviral therapy

Fred Lutwama  1 , Ronnie Serwadda, Harriet Mayanja-Kizza, Hasan M Shihab, Allan Ronald, Moses R Kamya, David Thomas, Elizabeth Johnson, Thomas C Quinn, Richard D Moore, Lisa A Spacek
Affiliations
Comparative Study

Evaluation of Dynabeads and Cytospheres compared with flow cytometry to enumerate CD4+ T cells in HIV-infected Ugandans on antiretroviral therapy

Fred Lutwama et al. J Acquir Immune Defic Syndr. .

Abstract

Background: Laboratory-based monitoring of antiretroviral therapy is essential but adds a significant cost to HIV care. The World Health Organization 2006 guidelines support the use of CD4 lymphocyte count (CD4) to define treatment failure in resource-limited settings.

Methods: We compared CD4 obtained on replicate samples from 497 HIV-positive Ugandans (before and during ART) followed for 18 months by 2 manual bead-based assays, Dynabeads (Dynal Biotech), and Cytospheres (Beckman Coulter) with those generated by flow cytometry at the Infectious Diseases Institute in Kampala, Uganda.

Results: We tested 1671 samples (123 before ART) with Dynabeads and 1444 samples (91 before ART) with Cytospheres. Mean CD4 was 231 cells/mm (SD, 139) and 239 cells/mm (SD, 140) by Dynabeads and flow cytometry, respectively. Mean CD4 was 186 cells/mm (SD, 101) and 242 cells/mm (SD, 136) by Cytospheres and flow cytometry, respectively. The mean difference in CD4 count by flow cytometry versus Dynabeads were 8.8 cells/mm (SD, 76.0) and versus Cytospheres were 56.8 cells/mm (SD, 85.8). The limits of agreement were -140.9 to 158.4 cells/mm for Dynabeads and -112.2 to 225.8 cells/mm for Cytospheres. Linear regression analysis showed higher correlation between flow cytometry and Dynabeads (r=0.85, r=0.73, slope=0.85, intercept=28) compared with the correlation between flow cytometry and Cytospheres (r=0.78, r=0.60, slope=0.58, intercept=45). Area under the receiver operating characteristics curve to predict CD4<200 cells/mm was 0.928 for Dynabeads and 0.886 for Cytospheres.

Conclusion: Although Dynabeads and Cytospheres both underestimated CD4 lymphocyte count compared with flow cytometry, in resource-limited settings with low daily throughput, manual bead-based assays may provide a less expensive alternative to flow cytometry.

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Figures

FIGURE 1
FIGURE 1
A, Correlation analysis of Dynabeads and flow cytometry for CD4 cell counts (cells/mm3). Solid line represents regression line. Dynabeads = 28 + 0.85 × flow cytometry; r2 = 0.73; Pearson correlation coefficient = 0.85. B, Bland-Altman plot comparing the difference between flow cytometry (FACS) and Dynabeads (Dyna) versus the mean of the 2 methods for CD4 cell count (cells/mm3).
FIGURE 2
FIGURE 2
A, Correlation analysis of Cytospheres and flow cytometry for CD4 cell counts (cells/mm3). Solid line represents regression line. Cytospheres = 45 + 0.58 × flow cytometry; r2 = 0.60; Pearson correlation coefficient = 0.78. B, Bland-Altman plot comparing the difference between flow cytometry (FACS) and Cytospheres (Cyto) versus the mean of the 2 methods for CD4 cell count (cells/mm3).
FIGURE 3
FIGURE 3
A, ROC curves for Dynabeads predicting CD4 lymphocyte count by flow cytometry. □, 100; ●, 200; *350 cells/mm3 by flow cytometry. B, ROC curves for Cytospheres predicting CD4 lymphocyte count by flow cytometry. □, 100; ●, 200; *, 350 cells/mm3 by flow cytometry.
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