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CD CLUSTERED ANTIBODIES
(anti-Human and others as indicated)
Mouse anti-human CD80 antibodies
CATALOG# CLONE# Workshop Host Form Price
RDI-CBL518 BB1 V mIgM purified $375.00
RDI-CD80-BB1FT " " " " FITC $469.00
RDI-CD80BB1PE "" " " PE $531.00
RDI-M1708clb Dal-1 -- mIgG1 purified $375.00
Product Specification: mouse monoclonal anti-human CD80
This clone has been derived from hybridization of SP2/0 cells with spleen cells of BALB/c mouse immunized with 3T3, which was transfected with the FCRII + B7 gen.. This antibody has been clustered to CD80 in one of the international Workshop on Human White Cell differentiation Antigens
Isotype Mouse IgG1.
Source Hybridoma culture supernatant.
Purification Ammoniumsulphate precipitation and ionexchange chromat0graphy.
Packing Each vial contains 1 ml with approximately 0.2 mg/ml monoclonal antibody and 10 mg BSA in 20 mM TRIS and 150 mM NaCl, pH 8.0.
Preservative Mertiolate (0.001%).
Storage and stability Monoclonal antibodies should be stored in the dark at 2-8°C. The reagent is stable until the expiry date stated on the vial label.
Major reactivity The monoclonal antibody is directed against the CD80- antigen, which is expressed on activated B lymphocytes.
Molecular mass 60 kDa.
Application Detection of activated B-cells.
Methods Indirect immunofluorescence staining with analysis by flowcytometry or fluorescence microscopy.
For research only! Not for human use!
APPLICATION FOR AN INDIRECT IMMUNOFLUORESCENCE TECHNIQUE
- FITC-conjugated goat anti-mouse immunoglobulin antiserum (M1204).
Remove aggregates by centrifugation at 1000 x g for 10 minutes.
- Buffer: Phosphate Buffered Saline (PBS), containing 0.2% Bovine Serum Albumin (BSA).
- PFA: Para-Formaldehyde 1%, pH 7.2, containing 0.2% BSA.
- Microtiter plates (96 wells, V bottom) or tubes.
1 Fixate the cells with PFA 1% and wash them with buffer.
2 Prepare a mononuclear cell suspension with a concentration of 1 x 10 7 cells/ml in buffer.
3 Add 45 µl of cell suspension to the microtiter wells or tubes.
4 Add 5 µl monoclonal antibody to the microtiter wells or tubes and mix gently.
5 Incubate for 30 minutes at 2-8 C.
6 Add 100 µl buffer to the microtiter wells or 2 ml buffer to the tubes and centrifuge at 500 x g for 5 minutes.
7 Aspirate the supernatant from the cell pellet and resuspend the cells.
8 Add 200 µl buffer to the microtiter wells or 2 ml buffer to the tubes and centrifuge at 500 x g for 5 minutes.
9 Aspirate the supernatant from the cell pellet and resuspend the cells.
10 Add 50 µl FITC-conjugated goat anti-mouse immunoglobulin serum, solid phase adsorbed with human immunoglobulins, diluted 1:80.
11 Incubate for 30 minutes at 2-8 C.
12 Repeat step 6-9.
13 Add 200 µl buffer.
14 The number of positive cells can be determined by flowcytometry analysis or by means of fluorescence microscopy.
NOTE: Care should be taken when drawing blood to avoid activation of platelets.Platelets activation markers are weakly expressed on platelets prepared from directly fixed normal whole blood.
The expression is maximal on thrombin (1 U/ml) stimulated washed platelets.
Mouse ANTI-HUMAN CD80
PRODUCT CODE RDI-CBL518 $438.00/vial 200ug
SOURCE: Tissue culture supernatant
Antigen: 594-S-F9 cell line
Fusion Partner: B3/NS1/1-Ag4.1 myeloma cell line
PURIFICATION METHOD: Gel filtration
SPECIFICITY: This antibody reacts with a 37 Kda protein expressed on activated B cells, macrophages and dendritic cells. The antigen is the cell surface ligand for CD28 which is a key T cell surface molecule.
Antigen-specific T cell activation depends on T cell receptor-ligand interaction and co-stimulatory signals generated when accessory molecules bind to their ligands such as CD28 to the BB1 accessory molecule. The BB1 ligand is also expressed by dendritic cells and this cell expression increases after binding to allogenic T cells. Recently it has been found that the BB1 molecule is also expressed on some other melanomas and perhaps on some other cancers. Two groups have shown recently that tumour cells transfected with BB1 become immunogenic and are rejected by the immune system.
*Flow cytometric analysis of CD28 binding
-B cell studies
-separation and isolation of dendritic cells
-Leucocyte Typing V. Oxford University Press (in press)
-Young, J W. et al, J. Clin. Invest. 90, 229-237 (1992)
-Koulova L. et al, J. Exp. Med. 173 , 759-762 (1991)
-Linsley, P.S. et al, Proc. Natl. Acad Sci, USA 87 5031-5035 (1990)
PRESENTATION: Each monoclonal is delivered in a sealed vial at a concentration of not less than 200µg/2ml in phosphate buffered saline containing 10mM sodium azide and 1mg/ml BSA . We recommend that each laboratory determine an optimum working titre for use in its particular application.
STORAGE: Purified monoclonal antibodies are generally stable for up to one month at 4oC. However repeated warming to room temperature and re-cooling may result in a loss of activity and hence effective working titre. It is recommended that monoclonal antibodies are aliquotted upon receipt and then stored at -20C. After thawing they should then be used as a stock solution for up to 30 days. Dilute working solutions are best prepared on the day of use.
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