Western blotting was performed as described previously [13 (link)]. Briefly, calluses were ground in liquid nitrogen and incubated with RIPA lysis buffer (P0013B, Beyotime Institute of Biotechnology, Shanghai) at 4 °C for 30 min, with shocking every 5 min. Samples were then centrifuged for 10 min at 14,000 rpm at 4 °C. Denatured proteins were separated by SDS-PAGE and transferred onto a PVDF membrane, which was blocked with 5% skim milk in 1× Tris-buffered saline Tween-20 (TBST, Applygen Technologies, Beijing). The membrane was then incubated with a primary antibody (anti-BMP2, anti-TGF-β1, anti-VEGF, or anti-β-actin (all from Abcam, MA, USA)) followed by a horseradish peroxidase-conjugated goat anti-mouse IgG secondary antibody (Beyotime Institute of Biotechnology, Shanghai). The membrane was then exposed to Kodak Biomax-Ml film (Eastman Kodak Co., USA). Signals were quantified by densitometry.
Horseradish peroxidase conjugated goat anti mouse igg secondary antibody
Manufactured by Beyotime
Sourced in United States, China
Horseradish peroxidase-conjugated goat anti-mouse IgG secondary antibody is a laboratory reagent used in immunoassays. It consists of a goat-derived antibody that binds to mouse immunoglobulin G (IgG), conjugated to the enzyme horseradish peroxidase.
Automatically generated - may contain errors
Lab products found in correlation
Anti β actin, by Abcam (1 mentions)
Anti bmp2, by Abcam (1 mentions)
Ripa lysis buffer p0013b, by Beyotime (1 mentions)
Anti vegf, by Abcam (1 mentions)
Anti tgf β1, by Abcam (1 mentions)
Ecl western blot kit, by CWBIO (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
Bicinchoninic acid protein kit, by Beyotime (1 mentions)
2 protocols using horseradish peroxidase conjugated goat anti mouse igg secondary antibody
1
Western Blotting Analysis of Growth Factors
2
NFIC Expression in Developing Dental Follicle Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total protein in DFCs at different developmental stages (PN days 1-7) was extracted using RIPA lysis buffer, and the protein concentration was determined using a Bicinchoninic Acid protein kit (Beyotime, Shanghai, China) according to the manufacturer's protocol. Protein lysates (40 µg) were separated using 12% SDS-PAGE gels and transferred to polyvinylidene fluoride (PVDF) membranes (Millipore, Bedford, MA, USA). The membranes were blocked with 1 3
5% skim milk at room temperature for 2 h and incubated with monoclonal mouse anti-rat NFIC antibody (1:1000, Abcam, Burlingame, CA, USA) and monoclonal mouse anti-rat β-actin (1:1000, Beyotime, Shanghai, China) overnight at 4 °C. β-actin was used as an internal control. The secondary antibody was horseradish peroxidase-conjugated goat anti-mouse IgG secondary antibody (1:1000, Beyotime, Shanghai, China). Immunoreactive bands were visualized using an ECL-Westernblot kit (CWBiotech, China). The densitometry of the bands was quantified with ImageJ 1.36b (NIH Freeware, USA). Immunocytochemical staining for NFIC in DFCs at PN day 5 was performed according to the protocol described for immunocytochemical staining for cytokeratin and vimentin.
5% skim milk at room temperature for 2 h and incubated with monoclonal mouse anti-rat NFIC antibody (1:1000, Abcam, Burlingame, CA, USA) and monoclonal mouse anti-rat β-actin (1:1000, Beyotime, Shanghai, China) overnight at 4 °C. β-actin was used as an internal control. The secondary antibody was horseradish peroxidase-conjugated goat anti-mouse IgG secondary antibody (1:1000, Beyotime, Shanghai, China). Immunoreactive bands were visualized using an ECL-Westernblot kit (CWBiotech, China). The densitometry of the bands was quantified with ImageJ 1.36b (NIH Freeware, USA). Immunocytochemical staining for NFIC in DFCs at PN day 5 was performed according to the protocol described for immunocytochemical staining for cytokeratin and vimentin.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!