Cells were washed with PBS and harvested in RIPA buffer containing protease and phosphatase inhibitors. Lysates were quantified by the Bradford method using BSA as a standard. The proteins were separated on 10% SDS-PAGE and transferred onto polyvinylidene difluoride membranes. Membranes were blocked in 5% BSA for 2h at room temperature and incubated overnight at 4°C with the following primary antibodies: anti-FGFR1 (1:500, ab76464), anti-pFGFR1 (1:750, ab59194), and anti-V5 (1:1500, ab27671). Beta-tubulin (1:2000, ab6046) was used as loading control. Membranes were washed with TBST and then incubated with HRP-conjugated anti-rabbit IgG secondary antibody (1:5000, NA934V) or HRP-conjugated anti-mouse IgG secondary antibody (1:5000; NB7539) for 1h at room temperature and revealed utilizing Amersham ECL detection (Amersham Biosciences).
Hrp conjugated anti rabbit igg secondary antibody
Manufactured by Cytiva
Sourced in United States
The HRP-conjugated anti-rabbit IgG secondary antibody is a laboratory reagent used to detect and visualize rabbit primary antibodies in various immunoassays and immunochemistry techniques. The antibody is conjugated with horseradish peroxidase (HRP), an enzyme that can catalyze a colorimetric or chemiluminescent reaction, enabling the detection of the target rabbit primary antibody.
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Lab products found in correlation
2 protocols using hrp conjugated anti rabbit igg secondary antibody
1
Immunoblot Analysis of FGFR1 Signaling
2
Immunohistochemical Analysis of Kidney Tissue
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The kidney tissue was fixed in 4% paraformaldehyde and embedded in paraffin wax. Tissue sections were cut at 5 μm, dewaxed, and then washed three times with PBS for 5 minutes. The tissue sections were incubated in 1% Triton X-100 and 3% hydrogen peroxide solution for 10 minutes [10 ]. The tissue sections were blocked with 5% goat serum in PBS for 30 min and incubated overnight at 4°C with primary antibodies against type IV collagen (1: 200), fibronectin (1: 200), HIF-1α (1: 100), and VEGF (1: 100). After washing three times with TBS for 5 min, the tissue sections were incubated with HRP-conjugated anti-rabbit IgG secondary antibody (Amersham, Arlington Heights, IL, USA) for 30 min at 37°C. Then, 50 μl of 3,3’-diaminobenzidine (DAB) substrate solution (Sigma-Aldrich, St. Louis, MO, USA) was added to the tissue sections and incubated for 10 min at room temperature. The tissue sections were rinsed three times with PBS, and counterstained with Mayer’s hematoxylin for 6–10 min, dehydrated, and examined using a light microscope.
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