CD4 (Leu-3a), clone SK3,7 and CD4 (Leu-3b), clone SK4, are derived from
hybridization of mouse NS-1 myeloma cells with spleen cells from BALB/c mice ...
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Monoclonal Antibodies Detecting Human Antigens
TM
Multi-Clone CD4 (Leu -3a+3b) FITC
Catalog No. 347413 (7413)
100 Tests
DESCRIPTION Specificity
CD4 (Leu-3a and Leu-3b) recognizes an antigen, Mr 59 kdaltons (kDa), present on T-helper/ inducer lymphocytes and monocytes.1-3
Antigen Distribution
Leu-3a and Leu-3b recognize non–cross-blocking epitopes on the CD4 molecule. The CD4 antigen is present on the helper/inducer T-lymphocyte subset (ie, CD4+CD3+) that comprises 28% to 58% of normal peripheral blood lymphocytes.4 It is also present on 78% to 89% of normal thymocytes.1 The CD4 antigen is present in low density on the cell surface of monocytes and in the cytoplasm of monocytes and macrophages (CD3–CD4+).3 The CD4 antigen is the receptor for the human immunodeficiency virus (HIV).5 Subjects infected with HIV have been found to exhibit a continuous loss of CD4+ lymphocytes and a relative increase in the CD8+ lymphocyte subset.6
Clone
CD4 (Leu-3a), clone SK3,7 and CD4 (Leu-3b), clone SK4, are derived from hybridization of mouse NS-1 myeloma cells with spleen cells from BALB/c mice immunized with human peripheral blood T lymphocytes.
Ig Chain Composition
The CD4 antibodies are composed of mouse IgG1 heavy chains and kappa light chains.
RESEARCH APPLICATIONS
• Enumeration of the T-helper/inducer subset in peripheral blood • Analysis of immunoregulatory T-lymphocyte subsets involved in helper/inducer functions8,9* • Studies characterizing T-cell leukemias and lymphomas10*
Product/Amount for Staining
Multi-Clone CD4 (Leu-3a+3b) FITC Cat. No. 347413 (7413) 20 μL/test
Method for Direct Immunofluorescence
Add 20 μL of reagent to 106 peripheral blood mononuclear cells (PBMCs) in 50 μL of medium containing 0.1% azide. Mix thoroughly and incubate for 15 to 30 minutes in the dark at 2° to 8°C. Wash with 1X PBS with 0.1% azide, add 0.5 mL of 1% paraformaldehyde, mix thoroughly, and analyze. If samples are not to be analyzed immediately, mix thoroughly just prior to analysis. See Becton Dickinson Procedure for Direct Immunofluorescence Staining of Cell Surfaces, Source Book Section 2.4.
Flow Cytometric Analysis
Single-Parameter Display of Peripheral Blood Lymphocytes Analyzed with a FACS® Brand Flow Cytometer
Relative Number of Cells
SINGLE-COLOR DIRECT IMMUNOFLUORESCENCE
Mouse IgG1 FITC Control
CD4 (Leu-3a+3b)FITC + 36%
1 2 3 4 Logarithmic Fluorescence Intensity
Performed on PBMCs with scatter gates set on the lymphocyte fraction. Laser excitation is at 488 nm.
* The published methods in the cited references have not been developed or tested by Becton Dickinson.
For research use only. Not for use in diagnostic or therapeutic procedures. Becton Dickinson Immunocytometry Systems 2350 Qume Drive San Jose, CA 95131-1807 Tel 877-232-8995 Ordering Information (800) 223-8226; Customer Support Center (800) 448-BDIS Source Book Section 4.16.1
HANDLING AND STORAGE
CHARACTERIZATION
The FITC conjugate is supplied as 50 μg of Leu-3a and 100 μg of Leu-3b in 2.0 mL (75 μg of antibody/mL). Buffered saline contains gelatin and 0.1% sodium azide. The vials should be stored at 2° to 8°C. Conjugated forms should not be frozen and should be protected from prolonged exposure to light. Each reagent is stable for the period shown on the bottle label when stored as directed.
To ensure consistently high-quality reagents, each lot of monoclonal antibody is tested for conformance with characteristics of a standard reagent using fluorescence analysis with a FACS brand flow cytometer. Representative data are included in this data sheet. Standard procedures are presented in detail in the Becton Dickinson Monoclonal Antibodies Source Book.
WARRANTY
WARNING
The products sold hereunder are warranted only to conform to the quantity and contents stated on the label at the time of delivery to the customer. There are no warranties, expressed or implied, which extend beyond the description on the label of the product. Becton Dickinson’s sole liability is limited to either replacement of the products or refund of the purchase price. Becton Dickinson is not liable for property damage, personal injury, or economic loss caused by the product.
Reagents contain sodium azide. Under aqueous acidic conditions, sodium azide yields hydrazoic acid, an extremely toxic compound. Azide compounds should be diluted with large volumes of water during disposal to avoid deposits in lead or copper plumbing where explosive conditions may develop.
REFERENCES 1. Evans RL, Wall DW, Platsoucas CD, et al. Thymus-dependent membrane antigens in man: Inhibition of cell-mediated lympholysis by monoclonal antibodies to the TH2 antigen. Proc Natl Acad Sci USA. 1981;78(1):544-548. 2. Ledbetter JA, Evans RL, Lipinski M, Cunningham-Rundles C, Good RA, Herzenberg LA. Evolutionary conservation of surface molecules that distinguish T-lymphocyte helper/inducer and T cytotoxic/suppressor subpopulations in mouse and man. J Exp Med. 1981;153:310-323. 3. Wood GS, Warner N, Warnke R. Anti-Leu-3/T4 antibodies react with cells of monocyte/macrophage and Langerhans lineage. J Immunol. 1983;131(1):212-216. 4. Reichert T, DeBruyère M, Denys V, et al. Lymphocyte subset reference ranges in adult Caucasians. Clin Immunol Immunopathol. 1991;60:190-208. 5. Dalgleish A, Beverley PC, Clapham PR, Crawford DH, Greaves MF, Weiss RA. The CD4 (T4) antigen is an essential component of the receptor for the AIDS retrovirus. Nature. 1984;312(December):763-767. 6. Giorgi J, Hultin L. Lymphocyte subset alterations and immunophenotyping by flow cytometry in HIV disease. Clin Immunol Newslett. 1990;10(4):55-62. 7. Bernard A, Boumsell L, Hill C. Joint report of the First International Workshop on Human Leucocyte Differentiation Antigens by the investigators of the participating laboratories: T2 protocol. In: Bernard A, Boumsell L, Dausset J, Milstein C, Schlossman S, eds. Leucocyte Typing. Berlin: Springer-Verlag; 1984:25-60. 8. Engleman EG, Benike CJ, Glickman E, Evans RL. Antibodies to membrane structures that distinguish suppressor/cytotoxic and helper T lymphocyte subpopulations block the mixed leukocyte reaction in man. J Exp Med. 1981;154:193-198. 9. Kotzin BL, Benike CJ, Engleman EG. Induction of immunoglobulin secreting cells in the allogeneic mixed leukocyte reaction: Regulation by helper and suppressor lymphocyte subsets in man. J Immunol. 1981;127:931-935. 10. Picker LF, Weiss LM, Medeiros LJ, Wood GS, Warnke RA. Immunophenotypic criteria for the diagnosis of non-Hodgkin’s lymphoma. Am J Pathol. 1987;128:181.
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