Expression of Beclin-1 was detected by IF staining. Paraffin-embedded tissue fraction (5 mm thickness) and xylene dewaxed, then dehydrated using graded concentrations of alcohol, and incubated with 3% H2O2 inhibited endogenous peroxidase.17 (link) Afterwards, it was blocked in 10% goat serum for 10 min at room temperature, incubated with the primary antibody in blocking solution overnight at 4°C. The primary antibodies for Beclin-1 (1:200, WL02508, WanleiBio) were used. Afterwards, the slides were washed in phosphate buffer saline (PBS), applied with horseradish peroxidase (HRP)-labeled anti-rabbit IgG (1:100; Beyotime, China) for 30 min at 37°C, then washed with PBS again. Then, images were acquired with a Nikon EclipseNi inverted microscope (TE2000, Nikon, Tokyo, Japan).
Horseradish peroxidase hrp labeled anti rabbit igg
Manufactured by Beyotime
Sourced in China
Horseradish peroxidase (HRP)-labeled anti-rabbit IgG is a secondary antibody that binds to rabbit primary antibodies. It is used in various immunoassay techniques to detect and quantify target proteins.
Automatically generated - may contain errors
Lab products found in correlation
Eclipse ni inverted microscope, by Nikon (1 mentions)
Wl02508, by Wanlei (1 mentions)
Enhanced chemiluminescence reagent, by Boster Bio (1 mentions)
Pvdf membrane, by Biosharp (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
Pvdf membrane, by Beyotime (1 mentions)
β actin, by Biosharp (1 mentions)
2 protocols using horseradish peroxidase hrp labeled anti rabbit igg
1
Beclin-1 Expression by Immunofluorescence
2
Western Blot Analysis of STIM1 and ORAI1
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Western blot analysis was performed as previously described in detail [20 (link)]. Simply, cells were lysed and centrifugated at 12,000g for 10 min at 4 °C. The supernatants were loaded in each well and subjected to 10% SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to PVDF membrane (Millipore, USA). Next, 5% nonfat milk was used to block the PVDF membrane in washing buffer for 2 h at room temperature and then incubated with primary antibody including STIM1, ORAI1 and β-actin (Biosharp, China) with diverse diluted ratio (1:1500, 1:2000 and 1:10,000) at 4 °C overnight. On the following day, washing PVDF membrane with TBST-T and incubate with 1:10,000 dilution of horseradish peroxidase HRP-labeled anti-rabbit IgG (Beyotime, China) for 1 h at room temperature. Finally each protein bands were visualized using enhanced chemiluminescence reagents (BOSTER, Wuhan, China).
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