FACSCount CD4 and Pima CD4 T-Cell Count Systems
Background: CD4 T-cell counts are used to screen and follow-up HIV-infected patients during treatment. As part of the World Health Organization prequalification program of diagnostics, we conducted an independent multicenter evaluation of the FACSCount CD4 and the Pima CD4, using the FACSCalibur as reference method.
Methods: A total of 440 paired capillary and venous blood samples were collected from HIV-infected patients attending the HIV outpatient clinic in Antwerp, Belgium, and the HIV care and treatment center in Dar es Salam, Tanzania. Capillary blood was run on Pima analyzer, whereas venous blood was analyzed on FACSCount, Pima, and FACSCalibur instruments. Precision and agreement between methods were assessed.
Results: The FACSCount CD4 results were in agreement with the FACSCalibur results with relative bias of 0.4% and 3.1% on absolute CD4 counts and an absolute bias of −0.6% and −1.1% on CD4% in Antwerp and Dar es Salam, respectively. The Pima CD4 results were in agreement with the FACSCalibur results with relative bias of −4.1% and −9.4% using venous blood and of −9.5% and −0.9% using capillary blood in Antwerp and Dar es Salam, respectively. At the threshold of 350 cells per microliter, the FACSCount CD4 and Pima CD4 using venous and capillary blood misclassified 7%, 9%, and 13% of patients, respectively.
Conclusions: The FACSCount CD4 provides reliable CD4 counts and CD4% and is suitable for monitoring adult and pediatric HIV patients in moderate-volume settings. The Pima CD4 is more suitable for screening eligible adult HIV patients for antiretroviral treatment initiation in low-volume laboratories.
Laboratory monitoring of HIV-infected patients receiving antiretroviral treatment (ART) is currently done by measuring HIV viral load and counting CD4 T cells. As many middle- and low-income countries do not have regular access to viral load testing, CD4 T-cell enumeration is the more common biological assay used to monitor ART. CD4 T-cell enumeration is used to start chemoprophylaxis against opportunistic infections and is also recommended as marker to identify patients in need of ART. Single-platform (SP) flow cytometry is the most preferred reference method for CD4 T-cell enumeration. However, classical and dedicated flow cytometers are still very expensive and operated by highly trained personnel. In addition, they require stable electricity supply, a cold chain to transport and store reagents, and regular instrument maintenance services, which are not readily available in most low-income countries. The FACSCount (BD Biosciences, Erembodegem, Belgium) is the first flow cytometer dedicated for absolute CD4 counting and is mainly used in resource-limited countries since 1996. Recently, new reagents and software have been developed to allow the additional measurements of CD4% essential for monitoring pediatric patients. Other small flow cytometers dedicated for CD4 counting have been introduced in the past decade and are also mainly used in resource-limited countries. These include CyFlow Counter (Partec, Munster, Germany), Apogee Auto40 (Apogee Flow System, Hemel Hempstead, United Kingdom), and Guava EasyCD4 (Merck Millipore, Billerica, MA). Most of these instruments can provide both CD4 T-cell counts and CD4%. More recently, alternative non–flow cytometry-based CD4 T-cell counting devices have been released in the market. Pima CD4 (Alere, Jena, Germany) has been introduced a few years ago as a point-of-care (POC) device, which uses either venous or capillary blood. However, it only provides absolute CD4 T-cell counts and thus is less suited for monitoring pediatric HIV-infected patients. Other CD4 POC technologies are in the pipeline; some are currently evaluated but are not yet available in the market. An overview of all existing and newly emerging CD4 technologies is provided in a recent UNITAID technical report on the diagnostic technology landscape and in recently published reviews.
The World Health Organization (WHO) recommended initiating ART in HIV patients with CD4 count up to 350 cells per microliter in 2010 and in 2013 raised the threshold up to 500 cells per microliter in specific patient populations. Misclassification of HIV patients at these thresholds may sometimes have grave outcomes. Very few studies have reported the ability of these assays to correctly classify patients at these critical thresholds. Laboratory and field evaluation of the CD4 systems is part of the prequalification of diagnostic program and is conducted after a successful review of the product dossier by the WHO. The assessment of the performance and operational characteristics of the product is one of the final phases and is carried out by WHO collaborating centers. As part of this program, we conducted an independent multicenter evaluation of the recently introduced FACSCount CD4 to measure absolute CD4 T-cell counts and CD4% and of the Pima CD4 using venous and capillary blood. The main objective was to assess their operational characteristics and performance, including patients' misclassification probabilities.
Abstract and Introduction
Abstract
Background: CD4 T-cell counts are used to screen and follow-up HIV-infected patients during treatment. As part of the World Health Organization prequalification program of diagnostics, we conducted an independent multicenter evaluation of the FACSCount CD4 and the Pima CD4, using the FACSCalibur as reference method.
Methods: A total of 440 paired capillary and venous blood samples were collected from HIV-infected patients attending the HIV outpatient clinic in Antwerp, Belgium, and the HIV care and treatment center in Dar es Salam, Tanzania. Capillary blood was run on Pima analyzer, whereas venous blood was analyzed on FACSCount, Pima, and FACSCalibur instruments. Precision and agreement between methods were assessed.
Results: The FACSCount CD4 results were in agreement with the FACSCalibur results with relative bias of 0.4% and 3.1% on absolute CD4 counts and an absolute bias of −0.6% and −1.1% on CD4% in Antwerp and Dar es Salam, respectively. The Pima CD4 results were in agreement with the FACSCalibur results with relative bias of −4.1% and −9.4% using venous blood and of −9.5% and −0.9% using capillary blood in Antwerp and Dar es Salam, respectively. At the threshold of 350 cells per microliter, the FACSCount CD4 and Pima CD4 using venous and capillary blood misclassified 7%, 9%, and 13% of patients, respectively.
Conclusions: The FACSCount CD4 provides reliable CD4 counts and CD4% and is suitable for monitoring adult and pediatric HIV patients in moderate-volume settings. The Pima CD4 is more suitable for screening eligible adult HIV patients for antiretroviral treatment initiation in low-volume laboratories.
Introduction
Laboratory monitoring of HIV-infected patients receiving antiretroviral treatment (ART) is currently done by measuring HIV viral load and counting CD4 T cells. As many middle- and low-income countries do not have regular access to viral load testing, CD4 T-cell enumeration is the more common biological assay used to monitor ART. CD4 T-cell enumeration is used to start chemoprophylaxis against opportunistic infections and is also recommended as marker to identify patients in need of ART. Single-platform (SP) flow cytometry is the most preferred reference method for CD4 T-cell enumeration. However, classical and dedicated flow cytometers are still very expensive and operated by highly trained personnel. In addition, they require stable electricity supply, a cold chain to transport and store reagents, and regular instrument maintenance services, which are not readily available in most low-income countries. The FACSCount (BD Biosciences, Erembodegem, Belgium) is the first flow cytometer dedicated for absolute CD4 counting and is mainly used in resource-limited countries since 1996. Recently, new reagents and software have been developed to allow the additional measurements of CD4% essential for monitoring pediatric patients. Other small flow cytometers dedicated for CD4 counting have been introduced in the past decade and are also mainly used in resource-limited countries. These include CyFlow Counter (Partec, Munster, Germany), Apogee Auto40 (Apogee Flow System, Hemel Hempstead, United Kingdom), and Guava EasyCD4 (Merck Millipore, Billerica, MA). Most of these instruments can provide both CD4 T-cell counts and CD4%. More recently, alternative non–flow cytometry-based CD4 T-cell counting devices have been released in the market. Pima CD4 (Alere, Jena, Germany) has been introduced a few years ago as a point-of-care (POC) device, which uses either venous or capillary blood. However, it only provides absolute CD4 T-cell counts and thus is less suited for monitoring pediatric HIV-infected patients. Other CD4 POC technologies are in the pipeline; some are currently evaluated but are not yet available in the market. An overview of all existing and newly emerging CD4 technologies is provided in a recent UNITAID technical report on the diagnostic technology landscape and in recently published reviews.
The World Health Organization (WHO) recommended initiating ART in HIV patients with CD4 count up to 350 cells per microliter in 2010 and in 2013 raised the threshold up to 500 cells per microliter in specific patient populations. Misclassification of HIV patients at these thresholds may sometimes have grave outcomes. Very few studies have reported the ability of these assays to correctly classify patients at these critical thresholds. Laboratory and field evaluation of the CD4 systems is part of the prequalification of diagnostic program and is conducted after a successful review of the product dossier by the WHO. The assessment of the performance and operational characteristics of the product is one of the final phases and is carried out by WHO collaborating centers. As part of this program, we conducted an independent multicenter evaluation of the recently introduced FACSCount CD4 to measure absolute CD4 T-cell counts and CD4% and of the Pima CD4 using venous and capillary blood. The main objective was to assess their operational characteristics and performance, including patients' misclassification probabilities.
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