Samples containing 30 µg protein were diluted with 4x LDS sample buffer containing DTT (Invitrogen), boiled for 5 minutes and separated by gel electrophoresis at 100 volts. Gels were transferred to nitrocellulose membranes with iBlot2 transfer system (P3 for 7 minutes) and visualized with Odyssey Sa imager at 800 nm. Membranes were then blocked with Odyssey blocking buffer for 2 hours at rt or overnight at 4 °C. The blots were incubated with desired antibodies (anti-HDAC1 (Abcam ab7028), anti-HDAC2 (Abcam, ab12169), anti-HDAC3 (Abcam, ab7030), anti-HDAC8 (Abcam, ab187139) and anti-HDAC4,5,9 (Abcam, ab131524)) at the recommended dilutions for each in Odyssey blocking buffer (LI-COR) overnight at 4 °C. The blots were then washed 3 × 5 minutes with PBST, incubated with relevant species of IRDye® 680RD secondary antibody (LI-COR) for 1 hour and visualized with Odyssey Sa imager at both 700 nm and 800 nm. If additional antibody probing was necessary, membranes were stripped with 0.2 N NaOH for 10 minutes and re-blockedwith Odyssey blocking buffer.
Ab131524
Manufactured by Abcam
Ab131524 is a lab equipment product manufactured by Abcam. It serves a core function, but a detailed description while maintaining an unbiased and factual approach is not available.
Automatically generated - may contain errors
Lab products found in correlation
Odyssey blocking buffer, by LI COR (1 mentions)
Irdye 680rd secondary antibody, by LI COR (1 mentions)
Dithiothreitol (dtt), by Thermo Fisher Scientific (1 mentions)
Ab7028, by Abcam (1 mentions)
Ab7030, by Abcam (1 mentions)
Ab187139, by Abcam (1 mentions)
Ab12169, by Abcam (1 mentions)
Flag antibody, by Merck Group (1 mentions)
Bz x810 all in one fluorescence microscope, by Keyence (1 mentions)
H3k27ac, by Abcam (1 mentions)
2 protocols using ab131524
1
Western Blot Analysis of HDACs
2
Immunofluorescence Assay for Cardiac Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Anti-Hdac4 + 5 + 9 antibody (Abcam, ab131524) is the antibody used to detect the endogenous condensate formation. This antibody directed target the 14-3-3 bind motif of Hdac4 around phosphorylated site S246. Flag antibody (Sigma F1804), H3K27ac (Abcam ab245911), V5 (Abcam ab9116), Mef2c (CST 5030), HA (ab9110), cTnT (Thermo Fisher Scientific), and GFP (Thermo Fisher Scientific) Antibodies were used. Cells were first fixed with 4% formaldehyde for 15 min. Then they were permeabilized with 0.1% Triton X-100 in PBS for another 15 min at room temperature. Cells were blocked with 4% horse serum in PBS for 1 h and then incubated with primary antibodies overnight at 4°C followed by incubation with appropriate fluorogenic secondary antibodies (Thermo Fisher Scientific) at room temperature for 1 h. The images were captured using the Keyence All-in-One Fluorescence Microscope BZ-X810. cTnT and α-actinin positive cells were manually quantified by single-blind method from randomly HPFs within each well.64 (link)
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