Ly6g Rat Monoclonal Antibody [Clone ID: RB6-8C5]

CAT#: CL046

Ly6g rat monoclonal antibody, clone RB6-8C5, Ascites

Product Images

Cell Source: Bone Marrow Monocytes - Percentage of cells stained above control: 93.2%

Specifications

Product Data

• Clone Name: RB6-8C5
• Applications: CT, FC, WB
• Recommended Dilution: Flow Cytometry.
  Complement mediated depletion.
  Western blot.
  Immunohistochemistry on Frozen and Paraffin Sections.
• Reactivities: Mouse
• Host: Rat
• Isotype: IgG2b
• Clonality: Monoclonal
• Specificity: This antibody monoclonal antibody reacts with the myeloid differentiation antigen Gr-1. (1,2). This 25-30 kDa cell surface antigen is expressed on myeloid cells but not lymphoid or erythroid cells. The expression of the Gr-1 antigen increases with granulocyte maturation (3) as shown by the distinct populations of bone-marrow cells this monoclonal antibody labels: negative, low positive and high positive. Expression is transient on cells of monocytic lineage (3).
• Formulation: State: Ascites
  State: Lyophilized Ascites (non-sterile filtered to 0.45 µm).
• Reconstitution Method: Restore with 0.5 ml of cold distilled water.
• Gene Name: lymphocyte antigen 6 complex, locus G
• Database Link:
  Entrez Gene 546644 Mouse
• Synonyms: Gr-1 Granulocyte marker
• Note: Protocol: FLOW CYTOMETRY ANALYSIS:
  
  Method:
  1. Prepare a cell suspension in media A. For cell preparations, deplete the red blood cell population with Lympholyte®-M 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 50 µl of a 1:5000-1:10000 dilution* of this AB.
  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 (FITC Goat anti-rat IgG (H+L)) 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:
  Mouse Strain: BALB/c
  Cell Concentration: 1x10e6 cells per test
  Antibody Concentration Used: 50µl of a 1:5000 dilution/10e6 cells
  Isotypic Control: Rat IgG2b
  
  Cell Source Percentage of cells stained above control:
  Thymus: 9.7%
  Whole Blood - Granulocytes: 98.6%
  Whole Blood - Monocytes: 93.5%
  Bone Marrow - Granulocytes: 88.9%
  Bone Marrow - Monocytes: 93.2%
  
  Results - Strain Distribution:
  Cell Concentration: 1x10e6 cells per test
  Antibody Concentration Used: 1:5000 in 50µl/10e6 cells
  Strains Tested: BALB/c, C57BL/6, CBA, C3H/He, AKR
  Positive: BALB/c, C57BL/6, CBA, C3H/He, AKR
  Negative: none