Accurate and Affordable HIV Monitoring by CD4 Counting

[Guest guest]

Dear Colleagues,

I would like to inform you on a recently published international CD4 counting

multicenter study which evaluates new alternatives for accurate and affordable

HIV monitoring, especially in resource-poor settings:

Cassens, U. et al: " Simplified volumetric flow cytometry allows feasible and

accurate determination of CD4 T lymphocytes in immunodeficient patients

worldwide " , Antiviral Therapy 9:395-405 (2004) (ISI Impact Factor 6.57),

International Medical Press 1359-6535/02

http://www.intmedpress.com/Journals/AVT/journals_avt_Abstract.cfm?

AVTAT=365 & AVTVN=9 & AVTDT=2004 & AVTIN=3

The abstract of this study is given as follows:

The determination of CD4 cells is of crucial clinical importance for patients

with AIDS. However, the high costs involved represent limitations for CD4 cell

counting in developing countries. In order to provide an affordable technique,

we introduced a simplified volumetric counting (SVC) technique without sample

manipulations and investigated it in a multicentre study. Blood samples from 434

healthy donors and immunodeficient patients were tested in eight hospital

laboratories in Europe, Africa and Asia.

CD4 cell counts were compared using in-house flow cytometric methods and the SVC

technique. The SVC method was performed on a low-cost flow cytometer (CyFlow SL,

Partec, Münster, Germany) after 15 min antibody incubation without pre-analytic

manipulations, such as washing or erythrocyte lysing procedures. Linear

regression analysis demonstrated a correlation of r=0.942 (Europe), r=0.952

(Africa) and r=0.989 (Asia) between the SVC technique and the in-house methods.

Bland Altman plot analysis of all patient data showed a mean bias between the

two methods of +26 CD4 cells in favour of the SVC technique (measured range:

6–1905 cells/µl; median CD4 cell count: 388/µl).

Three centres used the FACS-count technique (Becton-Dickinson, San José, Calif.,

USA) as an inhouse method dispensing with pre-analytic manipulations. The

comparison of SVC and FACS-count method revealed a mean bias of +32 CD4 cells/µl

(median CD4 cell count: 349/µl). The accuracy of the SVC was tested on standards

with known CD4 cell counts (n=6) and was shown to be 95.2%. The low-cost device

and the simplified no-lyse, no-wash test procedure reduces the costs per

determination and facilitates the use of flow cytometry in developing countries.

Best regards

Roland Göhde

HIV/AIDS Project Coordinator

E-mail: <r.goehde@...>

[Guest guest]

Thanks for sharing this! A few quick questions,

What is the cost of the SVC flow cytometer? What is the per test cost? What

is the throughput? How does it compare in these terms as well as in

specificity/sensitivity to other low-cost techniques such as Dynabeads,

Capellia, Guava, Partec, FlowCare (PointCare Techno) and FLO CD4 kit (BD)?

Any help would be greatly appreciated!

M.

At 10:33 PM 6/18/2004 +0000, you wrote:

>Dear Colleagues,

>

>I would like to inform you on a recently published international CD4

>counting multicenter

>study which evaluates new alternatives for accurate and affordable HIV

>monitoring,

>especially for resource-poor settings:

>

>Cassens, U. et al: " Simplified volumetric flow cytometry allows feasible

>and accurate

>determination of CD4 T lymphocytes in immunodeficient patients

>worldwide " , Antiviral

>Therapy 9:395-405 (2004) (ISI Impact Factor 6.57), International Medical

>Press 1359

>-6535/02

>

>http://www.intmedpress.com/Journals/AVT/journals_avt_Abstract.cfm?AVTAT=365 & AVT\

VN=9 & AVTDT=2004 & AVTIN=3

[Guest guest]

Dear ,

these are the information of the manufacturer:

Cost of the instrument: EUR 19.825

Cost per CD4 count : EUR 2

including mAbs

Annual Service Costs : < EUR 500

Sensitivity : down to 5 cells/ul (microliter)

Reproducibility : CV < 2,4%

Correlation : 0.998 (national proficiency test)

Accuracy : 95.2% (national proficiency test)

You will see many more data on several hundreds of blood samples shown

in the international multicenter study.

No lyse Protocol : 3-step procedure with 12 minutes (including 10

minutes incubation)

System Throughput : > 300 tests / day (related to instrument)

Protocol Capacity : approx. 250 tests / day (related to protocol)

Considering initiatives like the " 3x5 " WHO programme or national

programmes against HIV/AIDS, it becomes quite obvious that even more

than the costs the capacity plays a major role for monitoring.

This can be easily evaluated by a calculation for an examplary country:

Population : 20 Million

Infected Persons : 5% = 1 Million

In order to (theoretically) cover the infected persons and patients

with CD4 counting in order to decide on the start of ART and for

targeted treatment by four CD4 counts a year (therefore max. of 4

million tests), the different monitoring techniques can contribute as

follows:

Technique Costs per Test I Capacity test per day) I Required Labs

------------------------------------------------------------------------

------------------------------------

Magnetic Beads USD 3-12 I 10 I 2000

Conventional FCM USD 5-30 I 50 I 400

Volumetric FCM USD 2-3 I 250 I 80

Considering the costs and efforts for qualified lab personnel,

infrastructure, service, maintenance, etc. it becomes very clear that

only high capacity techniques will have a chance to significantly

improve the situation of the patients.

Interestingly enough, the volumetric FCM technique already is in use in

more than 20 countries in Africa.

Among the institutes and research groups which also successfully

evaluated this technique are renown sites like:

Université de Ougadougou, UFR/SVT, Burkina Faso

Hopital General de Douala, Cameroon

Hopital Laquintinie de Douala, Douala, Cameroon

Programme des Recherche Santé, Faculté des Sciences, University of

Douala, Cameroon

Hopital La Pette, Marua, Cameroon and MSF Younde, Cameroon

MSF Chiradzulo, Malawi, MSF Bruxelles, Belgium, MSF Paris, France

Centre Hospitalier Kigali (CHK) / Projet RWA21, Lux Development,

Kigali, Rwanda

Laboratory of Bacteriology-Virology, CHU-Le Dantec, Dakar, Senegal

Laboratory of Immunology, Department of Microbiology, Institute of

Tropical Medicine, Antwerp, Belgium

Calmette Hospital, Phnom Penh, Cambodia

Pasteur Institute, Phnom Penh, Cambodia

Croix-Rouge Française, Paris, France

Laboratoire d’Immunologie Hopital Pitiè Salpetriere, Paris, France

Institute of Immunology, Dept. of Biomedical Sciences, University of

Modena and Reggio Emilia, Italy

Spallanzani Hospital, Rome, Italy

Servizio d’Immunologia Istituto Pascale Hospital, Naples, Italy

Infectious Diseases Clinic San Raffaele Hospital, Milano, Italy

Institute of Immunology, -Rössle-Klinik Berlin, Germany

Institute of Laboratory and Transfusion Medicine, University Hospital

Bad Oeynhausen, Germany

Institute of Immunology and Transfusion Medicine, University Hospital

Lübeck, Germany

Department of Internal Medicine, University Hospital Münster, Germany

Institute of Transfusion Medicine, University Hospital Münster, Germany

Institute of Radiation Biology, University Hospital Münster, Germany

California Department of Health Services, Richmond (CA), USA

Also MSF has independently evaluated the volumetric FCM approach very

successfully. The conclusion of MSF resulting from the field trial is:

" After being trained, local district laboratory staff found the

technique easy to manipulate and robust under routine field conditions.

The technique produces excellent results in a field setting and can be

installed in a district hospital setting. It could provide HIV/AIDS

programmes with a new option for CD4+ monitoring in the context of

scaling-up ART in resource-limited settings. "

If your are interested in more details which show the feasibility of

the volumetric technique in comparison to conventional protocols and

the problematic use of standard protocols using lysing steos please see

also:

Cassens, U. et al: " A novel true volumetric method for the

determination of residual leucocytes in blood components " ,Vox

Sanguinis (2002) 82, 198–206

Greve, B. et al: " A New No-Lyse, No-Wash Flow-Cytometric Method for the

Determination of CD4 T Cells in Blood Samples " , Transfusion Medicine

and Hemotherapy (2003), 30:8-13

Göhde, W. et al: " Individual Patient-Dependent Influence of Erythrocyte

Lysing Procedures on Flow-Cytometric Analysis of Leukocyte

Subpopulations " , Transfusion Medicine and Hemotherapy (2003),

30:165-170

Greve, B. et al: " High-Grade Loss of Leukocytes and Hematopoietic

Progenitor Cells Caused by Erythrocyte-Lysing Procedures for Flow

Cytometric Analyses " , JOURNAL OF HEMATOTHERAPY & STEM CELL RESEARCH

12:321–330 (2003)

At the AIDS Conference in Bangkok in July there will be some scientific

talks and poster presentations concerning the successful implementation

of volumetric FCM CD4+ T-cell counting in Asia and Africa.

Concerning more information on the different CD4 counting techniques it

would be best to directly contact the manufacturers themselves and -

more important - the users who are working with these techniques in the

field settings in Africa.

Hopefully, I could answer your questions sufficiently.

Best regards

Roland Göhde

HIV/AIDS Project Coordinator

Am 19.06.2004 um 10:43 schrieb M. :

> Thanks for sharing this! A few quick questions,

>

> What is the cost of the SVC flow cytometer? What is the per test

> cost? What

> is the throughput? How does it compare in these terms as well as in

> specificity/sensitivity to other low-cost techniques such as

> Dynabeads,

> Capellia, Guava, Partec, FlowCare (PointCare Techno) and FLO CD4 kit

> (BD)?

>

> Any help would be greatly appreciated!

> M.

>

>

> At 10:33 PM 6/18/2004 +0000, you wrote:

> >Dear Colleagues,

> >

> >I would like to inform you on a recently published international CD4

> >counting multicenter

> >study which evaluates new alternatives for accurate and affordable

> HIV

> >monitoring,

> >especially for resource-poor settings:

> >

> >Cassens, U. et al: " Simplified volumetric flow cytometry allows

> feasible

> >and accurate

> >determination of CD4 T lymphocytes  in immunodeficient patients

> >worldwide " , Antiviral

> >Therapy 9:395-405 (2004) (ISI Impact Factor 6.57), International

> Medical

> >Press 1359

> >-6535/02

> >

>

> >http://www.intmedpress.com/Journals/AVT/journals_avt_Abstract.cfm?

> AVTAT=365 & AVTVN=9 & AVTDT=2004 & AVTIN=3