Reliable and accurate CD4+ T cell count and percent by the portable flow cytometer CyFlow MiniPOC and "CD4 Easy Count Kit-Dry", as revealed by the comparison with the gold standard dual platform technology

Abstract

Background: An accurate and affordable CD4+ T cells count is an essential tool in the fight against HIV/AIDS. Flow cytometry (FCM) is the "gold standard" for counting such cells, but this technique is expensive and requires sophisticated equipment, temperature-sensitive monoclonal antibodies (mAbs) and trained personnel. The lack of access to technical support and quality assurance programs thus limits the use of FCM in resource-constrained countries. We have tested the accuracy, the precision and the carry-over contamination of Partec CyFlow MiniPOC, a portable and economically affordable flow cytometer designed for CD4+ count and percentage, used along with the "CD4% Count Kit-Dry".

Materials and methods: Venous blood from 59 adult HIV+ patients (age: 25-58 years; 43 males and 16 females) was collected and stained with the "MiniPOC CD4% Count Kit-Dry". CD4+ count and percentage were then determined in triplicate by the CyFlow MiniPOC. In parallel, CD4 count was performed using mAbs and a CyFlow Counter, or by a dual platform system (from Beckman Coulter) based upon Cytomic FC500 ("Cytostat tetrachrome kit" for mAbs) and Coulter HmX Hematology Analyzer (for absolute cell count).

Results: The accuracy of CyFlow MiniPOC against Cytomic FC500 showed a correlation coefficient (CC) of 0.98 and 0.97 for CD4+ count and percentage, respectively. The accuracy of CyFlow MiniPOC against CyFlow Counter showed a CC of 0.99 and 0.99 for CD4 T cell count and percentage, respectively. CyFlow MiniPOC showed an excellent repeatability: CD4+ cell count and percentage were analyzed on two instruments, with an intra-assay precision below ± 5% deviation. Finally, there was no carry-over contamination for samples at all CD4 values, regardless of their position in the sequence of analysis.

Conclusion: The cost-effective CyFlow MiniPOC produces rapid, reliable and accurate results that are fully comparable with those from highly expensive dual platform systems.

Figure 1. Comparison of the CyFlow MiniPOC and CyFlow Counter.

Correlation and R² values for CD4+ T cell counts obtained by Cyflow MiniPOC and CyFlow Counter by using the CD4 easy count kit (A) and the CD4% easy count kit (B).

Figure 2. Comparison of the CyFlow MiniPOC and CyFlow Counter.

Bland-Altman plot comparing absolute CD4+ T cell counts obtained by Cyflow MiniPOC and CyFlow Counter by using the CD4 easy count kit (A) and the CD4% easy count kit (B).

Figure 3. Comparison of the CyFlow MiniPOC and CyFlow Counter.

Linear Regression(A) and Bland-Altman Plot (B) for CD4+ T cell percent obtained by Cyflow MiniPOC and CyFlow Counter by using the CD4% easy count kit.

Figure 4. Comparison of the CyFlow MiniPOC and BC FC 500.

Correlation and R² values for CD4+ T cell count (A) and percentage (B) obtained by Cyflow MiniPOC and BC FC 500 reference system.

Figure 5. Comparison of the CyFlow MiniPOC and BC FC 500.

Bland-Altman plot comparing absolute CD4+ T cell count (A) and percentage (B) obtained by Cyflow MiniPOC and BC FC 500 reference system

Figure 6. Precision assessment.

Bar charts with error bars indicating the standard deviation of CD4 T cell count (A) and CD4% (B) of a sample analyzed 10 times in single or in bulk. CV = Coefficient of variation.

Figure 7. Carry-over contamination analysis.

The CD4 absolute count (A) and CD4% (B) were plotted as the mean value of three analysis sequences. Error bars indicate the standard deviation of three measurements.

Figure 8. Partec CyFlow MiniPOC.

The Partec CyFlow MiniPOC counting device (A) and its display (B) in routine (left) or expert mode (right). Both images are used with permission of Partec/Sysmex GmbH, Germany.