Proteins were separated using 10% gel with an electrophoresis cells (GE Life Technologies). To separate α- and β-tubulins, 10% gel was prepared using a stock solution of 40% acrylamide solution (Meryer) supplemented with 0.54% bisacrylamide (w/v) powder (Yuanye) according to a protocol reported previously (39 (link)). After electrophoresis, proteins were transferred onto a nitrocellulose membrane using a semidry blotter (GE Healthcare). Membranes were incubated with anti-His tag (1:5000 dilution; Immuno Way), anti-glutamylation (GT335, 1:4000 dilution; Adipogen), anti-polyglutamylation (polyE, 1:4000 dilution; Adipogen), anti-α-tubulin (EP1332Y, 1:4000 dilution; Abcam or DM1A, 1:4000 dilution; Sigma), or anti-β-tubulin (anti-β-tubulin, 1:50,000 dilution; Proteintech) antibody. Immunoreactivity of proteins was visualized with WesternBright ECL (Advansta) using Western Blot Imager (Vilber) after incubation with horseradish peroxidase–labeled goat antimouse (1:5000 dilution, Bioss) or donkey anti-rabbit (1:5000 dilution, Bioss) antisera.
Horseradish peroxidase labeled goat antimouse
Manufactured by Bioss Antibodies
Sourced in China
Horseradish peroxidase–labeled goat antimouse is a laboratory reagent that contains goat-derived antibodies specific to mouse immunoglobulins, conjugated with the enzyme horseradish peroxidase. It is used as a detection agent in various immunoassay techniques, such as Western blotting and enzyme-linked immunosorbent assay (ELISA).
Automatically generated - may contain errors
2 protocols using horseradish peroxidase labeled goat antimouse
1
Separation and Detection of Tubulin Isoforms
2
Western Blot Analysis of Recombinant CCP6
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Recombinant CCP6 were separated by 10% polyacrylamide hydrogel. After electrophoresis, proteins were transferred onto a nitrocellulose membrane using a Semi-dry transfer device (100 mA, 1 h). Membranes were blocked with blocking buffer (5% milk/TBST), then incubated with mouse anti-His (1:5000; ImmunoWay, USA) antibody overnight at 4℃. After three times wash with TBST, the membrane was incubated with horseradish peroxidase-labeled goat anti-mouse (1:5000; Bioss, China) antisera at room temperature for 2 h. Immunoreactive protein bands were visualized with Western Bright ECL (Advansta, USA) reagents following three times wash with TBST.
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