Neutrophils Rat Monoclonal Antibody [Clone ID: 7/4]

CAT#: CL050B

Neutrophils rat monoclonal antibody, clone 7/4, Biotin

Product Images

Cell Source: Blood - Percentage of cells stained above control: 7.6%

Specifications

Product Data

• Clone Name: 7/4
• Applications: FC
• Recommended Dilution: Flow Cytometry.
  (Reported to be useful in Immunohistochemistry on frozen and paraffin sections. Strains reported to be positive for the 7/4 clone are: AKR, C57BL/6, C57BL/10, C58, DBA/2, MF1, NZB, NZW, SJL, Swiss (PO) and 129J. Strains reported to be negative/weak for the 7/4 clone are: A2G, A/Sn, ASW, BALB/c, C3H/HEH and CBA.T6T6.)
• Reactivities: Mouse
• Host: Rat
• Isotype: IgG2a
• Clonality: Monoclonal
• Immunogen: Cultured bone marrow cells
• Specificity: This monoclonal antibody is specific for mouse neutrophils. It reacts with the 7/4 antigen that is a polymorphic 40 kD molecule expressed by polymorphonuclear cells, but absent on resident tissue macrophages.
• Formulation: PBS
  Label: Biotin
  State: Liquid purified Ig
• Concentration: 0.1 mg/ml
• Purification: Protein A affinity purified
• Conjugation: Biotin
• Note: Protocol: FLOW CYTOMETRY ANALYSIS:
  
  Method:
  1. Prepare a cell suspension in media A. For cell preparations, deplete the red blood cell population using a NH4Cl lysing buffer.
  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 1x10e6 cells, representing 1 test).
  4. To each tube, add ~1.0 µg* of this Ab 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 detection reagent (Streptavidin-FITC) at 1:50 dilution.
  9. Incubate the tubes at 4°C for 30-60 minutes.
  (It is recommended that the tubes are protected from light since most fluorochromes are light sensitive).
  10. Wash 2 times at 4°C in media B.
  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: C57BL/6
  Cell Concentration: 1x10e6 cells per test
  Antibody Concentration Used: 1.0 ug/10e6 cells
  Isotypic Control: Biotin Rat IgG2a