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

CAT#: CL060B

MHC Class I H-2Ld mouse monoclonal antibody, clone 30-5-7S, Biotin

Product Images

Figure 1. Cell Source: Spleen. Percentage of cells stained above control: 91.1%

Specifications

Product Data

• Clone Name: 30-5-7S
• Applications: FC
• Recommended Dilution: Flow Cytometry (See Protocol below).
• Reactivities: Mouse
• Host: Mouse
• Isotype: IgG2a
• 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: 0.1 mg/ml
• Purification: Protein G Chromatography.
• Conjugation: Biotin
• 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 -