Three pituitaries from each genotype were pooled and lysed using RIPA buffer. The total lysate (30 μg/lane) was separated on an SDS-PAGE gel, and then a Western blot (WB) was performed using a standard protocol. We used the following antibodies: anti-TSHβ monoclonal antibody at 1:500 dilution for WB (Life Span Biosciences, LS-C334949) and anti-chorionic gonadotropin α at 1:1000 dilution for WB (Thermo Fisher Scientific, PA5-88517). THRB protein was assessed in the pituitary. Pituitaries were lysed in RIPA buffer with protease inhibitors and 20 mM N-ethylmaleimide. Two 10% SDS gels were prepared, 1 for WB and another for Coomassie blue staining. Protein (20 μg) was loaded 2 each lane, and gels were run in the same tank. Membrane was first blotted with anti-THRB antibody at 1:250 dilution (Abcam, ab104417). THRB protein was assessed in the thyroid. Crude lysate (30 μg) of thyroid gland from each mouse was loaded on a 10% SDS-PAGE gel. Detection of THRB and sumoylated THRB in the thyroid gland was carried out using ab-THRB1 (Abcam, ab180612) at 1:500 dilution.
Ab180612
Manufactured by Abcam
Ab180612 is a laboratory reagent for use in scientific research. It is a purified monoclonal antibody that can be used to detect a specific target protein in biological samples. The core function of this product is to provide a tool for researchers to identify and quantify the target protein of interest.
Automatically generated - may contain errors
Lab products found in correlation
Ab3625, by Abcam (1 mentions)
Criterion tgx precast gel, by Bio-Rad (1 mentions)
Pa1 211a, by Thermo Fisher Scientific (1 mentions)
Ma3 922, by Thermo Fisher Scientific (1 mentions)
Trans blot turbo, by Bio-Rad (1 mentions)
Odyssey infrared imaging system, by LI COR (1 mentions)
Ab8245, by Abcam (1 mentions)
2 protocols using ab180612
1
Western Blot Analysis of Pituitary and Thyroid THRB
2
Immunoblotting Analysis of Cardiac Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Immunoblotting was performed as previously described.38 Briefly, protein extracts from the heart were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) using 4-20% gradient polyacrylamide gels (Criterion™ TGX™ Precast Gels, No. 5671095; Bio-Rad) and then electroblotted into nitrocellulose membranes (Trans-Blot® Turbo; Bio-Rad). Blots were blocked and incubated with primary antibodies to phospholambam (MA3-922; Thermo Fisher Scientific), phospho-phospholambam (8496; Cell Signaling) ATPase sarcoplasmic/endoplasmic reticulum Ca2+ transporting 2 (SERCA2a; PA1-211A/IRDye 800; ab3625; Abcam), and thyroid hormone receptors a (PA1-211A; Thermo Fisher Scientific) and b (ab180612; Abcam) overnight at 4ºC. The immunoblots were subsequently washed and incubated with 700 or 800 nm infrared dye-conjugated antibodies (Nos. 926-68020 and 926-32211; LI-COR Biosciences). The membrane was imaged by scanning at 800 and 700 nm with an Odyssey Infrared Imaging System (LI-COR Biosciences). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as an internal control (ab8245; Abcam) and the control group was set as reference.
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