Mouse cells derived from peritoneal lavage (exudate) were collected after 12 h from zymosan intraperitoneal injection. Cells were then plated on fibronectin-coated (catalog no. F1141; Sigma-Aldrich) glass coverslips in Dulbecco's Modified Eagle Medium growth media (catalog no. 10565018; ThermoFisher) supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin; after attachment for ∼3 h, cells were fixed with 4% paraformaldehyde. Cells were then washed with 0.05% tris-buffered saline Triton (TBS-T). Blocking was performed with 3% bovine serum albumin and 3% normal horse serum in 0.05% TBS-T. Cells were then stained with a rat anti-mouse Ly-6G (clone 1A8; catalog no. 551459; BD Biosciences) and a rabbit polyclonal anti-mouse F4/80 (D4C8V; catalog no. 30325; Cell Signaling). Secondary antibodies were conjugated with DyLight 488 and DyLight 594 (Vector Laboratories). Nuclei were stained with 4′,6-diamidino-2-phenylindole (DAPI), and slides were mounted with Dako mounting media (Agilent). Rabbit polyclonal immunoglobulin G (catalog no. ab171870; Abcam) was used as a negative control (SI Appendix, Fig. S2C ). Images were acquired with a Nikon Eclipse T12 confocal microscope and processed with Fiji open-source software.
Rat anti mouse ly 6g clone 1a8
Manufactured by BD
Sourced in United States
The Rat anti-mouse Ly-6G (clone 1A8) is a laboratory reagent used for the detection and analysis of Ly-6G, a cell surface marker expressed on mouse granulocytes. This antibody can be used in various immunological techniques, such as flow cytometry, to identify and study these cell populations.
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Lab products found in correlation
2 protocols using rat anti mouse ly 6g clone 1a8
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Immunostaining of Mouse Peritoneal Macrophages
2
Cryosectioning and Immunostaining of Muscle Tissue
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Isolated TA muscles were snap-frozen in liquefied nitrogen-chilled isopentane (Sigma-Aldrich, United States), cryoembedded and cryosectioned at 7-μm thickness. All sections were fixed with paraformaldehyde solution (4% in PBS) for 10 min, followed by either hematoxylin and eosin (H&E) staining or immunohistochemistry, as previously described (Zhang et al., 2014 (link); Xia et al., 2016 (link)).
To evaluate muscle histology, images covering entire H&E-stained muscle sections were captured for analyses of centrally nucleated myofibers and myofiber cross-sectional area. For immunohistochemistry, primary antibodies, including rat anti-mouse Ly6G (Clone 1A8; 5 μg/ml; BD Biosciences, United States) and rat anti-mouse F4/80 (CI:A3-1; 10 μg/ml; Bio-Rad, United States); and secondary antibodies, including Alexa Fluor 488- or 568-conjugated goat anti-rat secondary antibodies (5 μg/ml; Thermo Fisher Scientific, United States) were used in the present study. The injury loci were identified and the fluorescent images were captured for qualitative analyses. All images were captured using an Eclipse 80i microscope (Nikon, Japan) equipped with a SPOT-camera (SPOT Imaging, United States) and analyzed with ImageJ image analysis software (National Institutes of Health, United States).
To evaluate muscle histology, images covering entire H&E-stained muscle sections were captured for analyses of centrally nucleated myofibers and myofiber cross-sectional area. For immunohistochemistry, primary antibodies, including rat anti-mouse Ly6G (Clone 1A8; 5 μg/ml; BD Biosciences, United States) and rat anti-mouse F4/80 (CI:A3-1; 10 μg/ml; Bio-Rad, United States); and secondary antibodies, including Alexa Fluor 488- or 568-conjugated goat anti-rat secondary antibodies (5 μg/ml; Thermo Fisher Scientific, United States) were used in the present study. The injury loci were identified and the fluorescent images were captured for qualitative analyses. All images were captured using an Eclipse 80i microscope (Nikon, Japan) equipped with a SPOT-camera (SPOT Imaging, United States) and analyzed with ImageJ image analysis software (National Institutes of Health, United States).
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