Resected tumors were frozen on dry ice in a cryo-embedding compound (Ted Pella, Inc; 27300), sectioned, and fixed in a 3:1 mixture of acetone:ethanol. Non-specific binding was blocked with 2% BSA in PBS, followed by staining with an antibody mix containing rabbit anti-mouse Ki67 antibody (Cell Signaling Technologies; 12202) and rat anti-mouse CD31 antibody (Dianova; DIA-310). Detection of primary antibodies was achieved using FITC conjugated goat anti-rabbit IgG (BD Pharmingen; 554020) and Cy3 conjugated goat anti-rat IgG (Invitrogen; A10522). Samples were counterstained with DAPI (Vector; H-1500) and mounted with a hard-set mounting medium for fluorescence. Random images from each section were obtained with a Zeiss AxioImager A2 epifluorescence microscope at 200x magnification, and analyzed with ImageJ. CD31+ cells (% area) and Ki67+ cells (% cells) were quantified automatically using macro functions, whereas Ki67+/CD31+ cells (proliferating endothelial cells) were quantified manually.
Cy3 conjugated goat anti rat igg
Manufactured by Thermo Fisher Scientific
Sourced in United States
Cy3 conjugated goat anti-rat IgG is a secondary antibody used for detection and visualization of rat immunoglobulin G (IgG) in various immunoassays and imaging applications. The antibody is conjugated with the Cyanine 3 (Cy3) fluorescent dye, which emits in the orange-red region of the visible spectrum upon excitation.
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Lab products found in correlation
4 6 diamidino 2 phenylindole (dapi), by Vector Laboratories (1 mentions)
Rat anti mouse cd31 antibody, by Dianova (1 mentions)
Fitc conjugated goat anti rabbit igg, by BD (1 mentions)
Axio imager a2 epifluorescence microscope, by Zeiss (1 mentions)
Fitc conjugated goat anti rat igg, by Thermo Fisher Scientific (1 mentions)
Cy3 conjugated goat anti rabbit igg, by Thermo Fisher Scientific (1 mentions)
2 protocols using cy3 conjugated goat anti rat igg
1
Quantitative Tumor Angiogenesis Imaging
2
Immunofluorescent Labeling of Brain Sections
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Individual brain sections were incubated with PBS containing 0.3% Triton X-100 and 3% normal donkey serum for 2 hours, and then with the appropriate primary antibodies overnight for 4°C as follows: rabbit anti-Oligo2 polyclonal antibody (1:500; Millipore, Billerica, MA, USA), rat anti-MBP polyclonal antibody (1:250; Millipore) and rat anti-PLP polyclonal antibody (1:250; Millipore). The sections were washed three times in PBS and incubated with Cy3-conjugated goat anti-rabbit IgG (1:400; Invitrogen, Carlsbad, CA, USA), Cy3-conjugated goat anti-rat IgG (1:400; Invitrogen) or fluorescein isothiocyanate (FITC)-conjugated goat anti-rat IgG (1:400; Invitrogen) for 2 hours at room temperature. Finally, the sections were observed under a fluorescence microscope.
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