MHC Class I (RT1Ac) Mouse Monoclonal Antibody [Clone ID: OX-27]

Specifications

Product Data
Clone Name	OX-27
Applications	FC
Recommend Dilution	Flow cytometry: use 0.5 µg of neat antibody to label 10e6 cells.
Reactivity	Rat
Host	Mouse
Clonality	Monoclonal
Immunogen	Phytohaemagglutinin Blasts.
Formulation	PBS buffer with 0.02% sodium azide as preservative and 0.5 % EIA grade BSA as stabilizer. Label: FITC State: Liquid purified IgG fraction. Label: conjugated
Concentration	lot specific
Purification	Protein G Chromatography.
Conjugation	FITC
Storage Condition	Store the antibody at 2-8°C for one month or (in aliquots) at -20°C for longer. Avoid repeated freezing and thawing.
Database Link	UniProt ID Q31255
Background	MHC Class I molecules play a central role in the immune system by presenting peptides derived from the endoplasmic reticulum lumen. MHC class I antigens are heterodimers consisting of one alpha chain (44kDa) with beta 2 microglobulin (11.5 kDa). The antigen is expressed by all somatic cells at varying levels. MHC Class I molecules are expressed on most nucleated cells where they present endogenously synthesized antigenic peptides to CD8+ T lymphocytes, which are usually cytotoxic T cells. Fibroblasts or neurons however only show a low level of antigen.
Synonyms	RT1-A3
Note	Recognizes a polymorphic determinant of the MHC class I antigen in the rat. This Antibody can be used for labelling cells of donor or host origin in bone marrow chimeras. (1,2) Protocol: FLOW CYTOMETRY ANALYSIS: Method: 1. Prepare a cell suspension in media A. For cell preparations, deplete the red blood cell population with Rat cell separation medium 2. Wash 2 times. 3. Resuspend the cells to a concentration of 2x10e7 cells/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 0.5-0.2 µg 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. (protect tubes from light) 7. Wash 2 times at 4°C. 8. Resuspend the cell pellet in 50 µl ice cold media B. 9. 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: Rat Strain: Brown Norway Cell Concentration: 1x10e6 cells per test. Antibody Concentration Used: 0.2 µg/10e6 cells Isotypic Control: FITC Mouse IgG2a. Cell Source-Percentage of cells stained above control: Thymus: 73.9% Spleen: 99.0% Lymph Node: 100%
Reference Data

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