Ly-6G Neutrophil Marker antibody

Antigen: Ly-6G Neutrophil Marker

Clonality: Monoclonal (RB6-8C5)

Host: Rat

Reactivity: Mouse (Murine)

Conjugate: Un-conjugated

Application: Cytotoxicity Test (CyTox), Western Blotting (WB), Flow Cytometry (FACS)

Catalog no.: ABIN114268

Additional Information

Alternative name: Ly6G / GR1 Neutrophil Marker

UniProt: P35461

Cross-Reactivity: Mouse (Murine)

Isotype: IgG2b

Clone: RB6-8C5

Characteristics: Synonyms: Gr-1 Granulocyte marker

Application Details

Protocol: FLOW CYTOMETRY ANALYSIS: Method: 1. Prepare a cell suspension in media A. For cell preparations, deplete the red blood cellpopulation 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 thissuspension 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 protectedfrom 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 propidiumiodide 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 (100µl of 2M sodium azide in 100 mls). Results - Tissue Distribution: Mouse Strain: BALB/cCell

Application Notes: Flow Cytometry. Complement mediated depletion. Western blot. Immunohistochemistry on Frozen and Paraffin Sections. Other applications not tested. Optimal dilutions are dependent on conditions and should be determined by the user.

Concentration: 1x10e6 cells per testAntibody Concentration Used: 50µl of a 1: 5000 dilution/10e6 cellsIsotypic Control: Rat IgG2bCell Source Percentage of cells stained above control: Thymus: 9.7%Whole Blood - Granulocytes: 98.6%Whole Blood - Monocytes: 93.5%Bone Marrow

Purification: Ascites

Storage: Prior to reconstitution and Following reconstitution store the antibody undiluted at 2-8°Cfor one month or (in aliquots) at -20°C for longer. Avoid repeated freezing and thawing. Shelf life: one year from despatch.

Restrictions: For Research Use only

Images

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

Publications

Hestdal, Ruscetti, Ihle et al.: "Characterization and regulation of RB6-8C5 antigen expression on murine bone marrow cells." in: Journal of immunology (Baltimore, Md. : 1950), Vol. 147, Issue 1, pp. 22-8, 1991 (PubMed).

Jutila, Kroese, Jutila et al.: "Ly-6C is a monocyte/macrophage and endothelial cell differentiation antigen regulated by interferon-gamma." in: European journal of immunology, Vol. 18, Issue 11, pp. 1819-26, 1989 (PubMed).

Spangrude, Heimfeld, Weissman: "Purification and characterization of mouse hematopoietic stem cells." in: Science (New York, N.Y.), Vol. 241, Issue 4861, pp. 58-62, 1988 (PubMed).

Brummer, Sugar, Stevens: "Immunological activation of polymorphonuclear neutrophils for fungal killing: studies with murine cells and blastomyces dermatitidis in vitro." in: Journal of leukocyte biology, Vol. 36, Issue 4, pp. 505-20, 1984 (PubMed).

Whiteland, Nicholls, Shimeld et al.: "Immunohistochemical detection of T-cell subsets and other leukocytes in paraffin-embedded rat and mouse tissues with monoclonal antibodies." in: The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society, Vol. 43, Issue 3, pp. 313-20, 1995 (PubMed).