MHC Class I H-2Ld Mouse Monoclonal Antibody [Clone ID: 30-5-7S]

Specifications

Product Data
Clone Name	30-5-7S
Applications	FC
Recommend Dilution	Flow Cytometry (See Protocol below).
Reactivity	Mouse
Host	Mouse
Clonality	Monoclonal
Specificity	This antibody antibody detects the public specificity H-2.65 of the H-2Ld antigen. It also recognizes H-2Dq and H-2Lq molecules.
Formulation	PBS containing 0.02% Sodium Azide as preservative and EIA grade BSA as a stabilizing protein to bring total protein concentration to 4-5 mg/ml. Label: Biotin State: Liquid purrified Ig fraction.
Concentration	lot specific
Purification	Protein G Chromatography.
Conjugation	Biotin
Storage Condition	Store the antibody undiluted at 2-8°C for one month or (in aliquots) at -20°C for longer. Avoid repeated freezing and thawing.
Database Link	UniProt ID P01897
Background	The classical MHC Class I molecules are histocompatibility antigens encoded by the H-2 gene complex and consist of heterodimers of highly polymorphic alpha chains noncovalently associated with the invariant Beta2-microglobulin. These antigens are expressed on most nucleated cells but expression varies on different cell types. MHC Class I molecules present endogenously synthesized peptides to CD8+ T lymphocytes, which are usually cytotoxic T cells. MHC Class I antigens expressed on thymic epithelial cells regulate the positive and negative selection of CD8+ T cells during T cell ontogeny.
Synonyms	H2-L
Note	Protocol: FLOW CYTOMETRY ANALYSIS: Method: 1. Prepare a cell suspension in media A. For cell preparations, deplete the red blood cell population with cell separation medium. 2. Wash 2 times. 3. Resuspend the cells to a concentration of 2x10e7cells/ml in media A. Add 50 µl of this suspension to each tube (each tube will then contain 1 x 10e6 cells, representing 1 test). 4. To each tube, add 1.0 µg of antibody per 10e6 cells. 5. Vortex the tubes to ensure thorough mixing of antibody and cells. 6. Incubate the tubes for 30 minutes at 4°C. 7. Wash 2 times at 4°C. 8. Add 100 µl of secondary antibody (Streptavidin-FITC) at a 1:500 dilution. 9. Incubate tubes at 4°C for 30-60 minutes (It is recommended that tubes are protected from light since most fluorochromes are light sensitive). 10. Wash 2 times at 4°C. 11. Resuspend the cell pellet in 50 µl ice cold media B. 12. Transfer to suitable tubes for flow cytometric analysis containing 15 µl of propidium iodide at 0.5 mg/ml in PBS. This stains dead cells by intercalating in DNA. Media: A. Phosphate buffered saline (pH 7.2) + 5% normal serum of host species + sodium azide (100 µl of 2M sodium azide in 100 mls). B. Phosphate buffered saline (pH 7.2) + 0.5% Bovine serum albumin + sodium azide (100 µl of 2M sodium azide in 100 mls). Results: Tissue Distribution by Flow Cytometry Analysis: Mouse Strain: Balb/c Cell Concentration : 1x10e6 cells per test Antibody Concentration Used: 1.0 µg/10e6 cells Isotypic Control: Biotin Mouse IgG2a Cell Source Percentage of cells stained above control: (See Figure 1.) Spleen 91.1% Thymus 99.4% Lymph Node 98.1% Strain Distribution: Strains Tested: Strain Haplotype + / - BALB/c H-2d + A.TH H-2KsDd + A.TL H-2KsDd + B.10A(3R) H-2KbDd + C3H/He H-2k - C57BL/6 H-2b -
Reference Data

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