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05-8404 - CD4% Easy Count Kit - 10-03-24 - Rev009
05-8404 - CD4% Easy Count Kit - 10-03-24 - Rev009
General Information: Patients with AIDS exhibit T-cell Clone CD4 mAb PE MEM-241
lymphopenia, a loss of CD4+ lymphocytes and a relative CD45 mAb FITC MEM-28
increase in the CD8+ subtype and in the CD3+CD4-CD8-
subtype. The generally used threshold of 200 CD4+ Isotype CD4 mAb PE Mouse monoclonal IgG1
T-cells/µl blood to start antiretroviral therapy (ART) for adult CD45 mAb FITC Mouse monoclonal IgG1
patients may not be appropriate for children since the CD4
counts in infants are significantly higher than for adults.
Therefore a CD4% measurement should be performed in Applications:
pediatric patients for reliable and controlled ART and • Identification and enumeration of the CD4+ helper/inducer
patient monitoring. T-lymphocyte subset as well as CD45+ cells of
haemopoietic lineage
Specificity and Antigen Distribution: The mouse • Determination of percentage of CD4+ T-cells in relation to
monoclonal antibody MEM-241 recognizes the human CD4 all lymphocytes
antigen, a transmembrane glycoprotein (55 kDa) of the • Studies of HIV infection. Patients infected with HIV loose
immunoglobulin supergene family, present on a subset of CD4+ helper/inducer lymphocytes and contemporarily
T-lymphocytes (“helper/inducer” T-cells) and also increase CD8+ T-cells
expressed at a lower level on monocytes, tissue • Characterization of subtypes of T-cell leukemias and
macrophages and granulocytes. Approximately 20-60% of lymphomas
human peripheral blood mononuclear cells as well as a • For in vitro diagnostic use
subpopulation of monocytes but with a weaker signal are
stained. The antibody has been studied at the 8th Method:
International Workshop on Human Cell Differentiation • Add 20 µl whole blood (EDTA as anticoagulant) to a
Molecules HCDM (former HLDA VIII), May 2006, Québec, Partec test tube
Canada. CD4 is the primary cellular receptor for the human • Add 10 µl of CD4 mAb PE and 10 µl of CD45 mAb FITC.
immunodeficiency virus (HIV). Mix gently and incubate for 15 minutes at room temperature
The mouse monoclonal antibody HI-28 reacts with all protected from light
alternative forms of the human CD45 antigen (Leukocyte • Add 400 µl of Buffer 1 and shake gently
Common Antigen), a 180-220 kDa single chain type I • Directly prior to the measurement add 400 µl of Buffer 2
transmembrane protein expressed at high level on all cells and analyze immediately (within 10 minutes)
of hematopoietic origin, except erythrocytes and platelets. • Analyze blood sample on a Partec CyFlow device
The antibody has been clustered to CD45 at the 3rd
International Workshop on Human Leukocyte
Differentiation Antigens (Ref.: Leucocyte Typing III,
McMichael A. J. et al. (Eds.), Oxford University Press,
Oxford 1987).
Contact Information:
Flow Cytometric Analysis: CD4-PE and CD45-FITC Warnings and Precautions: Reagents contain 0.09%
fluorescence can be analysed on a Partec Flow Cytometer sodium azide that under acidic conditions yields hydrazoic
with an excitation light source of 488 nm (blue solid state acid, an explosive compound. Sodium azide may react with
laser). To count CD4+ T-cells and CD45+ cells connect the lead and copper plumbing to form potentially explosive metal
test tube with 840 µl of the ready prepared blood sample (see azides. When disposing of sodium azide, flush plumbing with
Method) to your Partec Flow Cytometer and carry out a count a large volume of water to prevent build-up. Very toxic if
analysis. For detailed information on the handling and swallowed. After contact with skin, wash immediately with
calibration of the instrument please refer to the operating plenty of water.
manuals and application notes.
The blood preparation allows one count procedure on the Harmful if swallowed (R22). Contact with acids liberates very
same sample. Following the above procedure the dilution toxic gas (R32). Keep out of reach of children (S2). Keep
factor is 42. In the CyFlow® SL this dilution factor can be away from food, drink and animal feeding stuffs (S13). Wear
directly set by the instrument operator in the instrument suitable protective clothing (S36). If swallowed, seek medical
software. Counting results will be displayed automatically as advice immediately and show this container or label (S46).
CD4+ T-cells per µl whole blood and as percentage of CD4+
cells in relation to all lymphocytes. Handle all specimens and materials as potentially infectious
materials. Decontaminate with 0.5% sodium hypochlorite. The
Recommended and Required Materials: product should be handled by trained staff only. Reagents are
• Partec Flow Cytometry instrument (e.g. CyFlow® SL) not sterile. Not for in vivo use in human. For in vitro use only.
• Partec test tubes (Code No. 04-2000)
• Micropipettes and pipette tips (e.g. Eppendorf, Code No. Literature:
3112 000.029 and 3111 000.0165) Cassens, U. et al.: Simplified volumetric flow cytometry allows feasible and
accurate determination of CD4 T lymphocytes in immunodeficient patients
• Powder-free latex gloves (e.g. Safeskin, Code No. 545-950- worldwide”, Antiviral Therapy 9: 395-405 (2004)
06)
• Venous Blood Collection System with EDTA as Göhde, W. et al.: A New Protocol to Follow-up Immune Impairment in HIV/AIDS-
Children, Scientific Presentation - Abstract No. MoPeLB151C02 at the 3rd IAS
anticoagulant (e.g. Greiner Bio-One: Vacuette® EDTA Conference on HIV Pathogenesis and Treatment in Rio de Janeiro, July 2005
Tubes, K3E/EDTA K3, 3 ml, Code No. 454217, Vacuette®
Blood Collection Needles 38 x 0.8 mm, Code No. 450076, Greve, B. et al.: ”A New No-Lyse, No-Wash Flow-Cytometric Method for the
Determination of CD4 T Cells in Blood Samples”, Transfus Med Hemoth (2003),
Vacuette® Tube Holders, Code No. 450201) 30: 8-13
Handling and Storage: CD4 mAb PE and CD45 mAb FITC Imade, G. et al.: “Comparison of a New, Affordable Flow Cytometric Method and
each are supplied in 1 ml of phosphate-buffered saline (PBS) the Manual Magnetic Bead Technique for CD4 T-Lymphocyte Counting in a
Northern Nigerian Setting”, Clinical and Diagnostic Laboratory Immunology, Jan.
pH 7.4, containing BSA and 0.09% sodium azide, sufficient 2005, p. 224-227
for 100 tests. 10 µl of CD4 mAb PE and 10 µl of CD45 mAb
FITC are sufficient for labelling 1x106 cells. When stored at 2- Dieye, T.N. et al.: Absolute CD4 T-Cell Counting in Resource-Poor Settings: Direct
Volumetric Measurements Versus Bead-Based Clinical Flow Cytometry
8°C in the dark, the CD4% easy count kit is stable until the Instruments, Journal of Acquired Immune Deficiency Syndromes, Volume 39,
expiration date printed on the kit label. Do not freeze or Number 1, May 1 2005
expose to elevated temperatures.
Fryland, M. et al.: The PARTEC CyFlow counter for CD4+ T-cell counting
produces high quality results and is robust when evaluated under routine field
conditions in Malawi, Scientific Oral Presentation - Abstract No. TuOrB1149 at the
XV. International AIDS Conference in Bangkok, July 2004
Contact Information: