To detect Notch1- and GFAP-positive cells in the right cerebral cortex of the ischemic hemisphere, brain sections were incubated for 30 minutes in 2.0 M HCl to denature DNA, and the reaction was neutralized in 0.1 M boric acid for 10 minutes. Thereafter, brain sections were rinsed in phosphate-buffered saline (PBS) containing 0.3% Triton for 30 minutes, preincubated in 10% normal goat serum for 2 hours at room temperature, and incubated with polyclonal rabbit anti-Notch1 (1:150; Biosynthesis Biotechnology, Beijing, China) and monoclonal rabbit anti-GFAP (astrocyte marker) antibody (1:200; Boster Biotechnology, Wuhan, China) at 4°C overnight, and then incubated with Cy3-conjugated affinity-purified goat anti-rabbit IgG (1:100; Sigma, St. Louis, MO, USA) in a humidified chamber for 1 hour at 37°C. Anti-Notch1 and anti-GFAP were used as cell-type specific markers in each brain section. Numbers of Notch1- and GFAP-positive cells were counted by laser-scanning confocal microscopy (Olympus).
Cy3 conjugated affinity purified goat anti rabbit igg
Manufactured by Merck Group
Sourced in United States
Cy3-conjugated affinity-purified goat anti-rabbit IgG is a secondary antibody used in various immunoassays and imaging techniques. It is designed to detect and bind to rabbit primary antibodies, allowing for the visualization and detection of target proteins or molecules in a sample.
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2 protocols using cy3 conjugated affinity purified goat anti rabbit igg
1
Immunohistochemical Analysis of Notch1 and GFAP
2
Wnt Signaling in Ischemic Hippocampus
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In order to investigate the regulatory role of Wnt signalling pathway, β-catenin or Wnt-3a positive cell in the ischaemic hippocampus was detected by immunofluorescence staining. Brain section was incubated for half-hour in 2.0 M HCl to denature DNA, and the reaction was neutralised in 0.1 M boric acid for 15 min. Thereafter, brain section was rinsed in PBS containing 0.3% Triton for half an hour, preincubated in 10% normal goat serum for 2 h at indoor temperature, and incubated with monoclonal rabbit anti-Wnt-3a or anti-β-catenin (1:150; Boster Biotechnology, Wuhan, China) at 4°C overnight, and then incubated with Cy3-conjugated affinity purified goat anti-rabbit IgG (1:100; Sigma, St. Louis, MO, USA) in a humidified chamber for 1 h at 37°C. Anti-Wnt-3a or anti-β-catenin was used as cell-type specific markers in each brain section. The number of β-catenin or Wnt-3a positive cells was analysed by laser scanning confocal microscope analysis system (Olympus, Japan).
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