Western blot and ip lysis buffer

Manufactured by Beyotime

Sourced in China

Western blot and IP lysis buffer is a solution used for the extraction and solubilization of proteins from cells or tissues. It is designed to preserve the native structure and activity of the proteins during the lysis process, making it suitable for downstream applications such as Western blotting and immunoprecipitation.

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Lab products found in correlation

Protein a g magnetic beads, by Selleck Chemicals (1 mentions) Sds loading buffer, by Beyotime (1 mentions) Anti flag m2 affinity gel, by Merck Group (1 mentions)

Spelling variants of this product

Lysis buffer (984 citations) IP lysis buffer (228 citations) IP buffer (46 citations) Western/IP lysis buffer (38 citations) Western and IP lysis buffer (33 citations) Western and IP buffer (10 citations) Cell lysis buffer for Western blot and IP (8 citations) Western blot lysis buffer (5 citations)

2 protocols using western blot and ip lysis buffer

Co-Immunoprecipitation and Western Blot Analysis

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For co-IP, total proteins were extracted with western blot and IP lysis buffer (Beyotime Biotech) from SSCs and C18-4, 293T or NIH3T3 cells. After preclearing with 30 μL protein A/G magnetic beads (Selleck) at 4°C, the proteins were incubated with 40 μL magnetic beads and primary antibodies or control IgG at 4°C overnight. Then, the beads were washed with PBS three times for 10 min each. Finally, the beads were resuspended in 50 μL of western blot and IP lysis buffer with SDS loading buffer (Beyotime) and boiled at 100°C for 5 min. Mass spectrometry analysis was conducted by OE Biotech Co. Ltd.
Total proteins of cultured cells were extracted with western blot and IP lysis buffer (Beyotime) and boiled with SDS loading buffer at 100°C for 5 min. Then, western blotting was performed with standard procedures. For testicular protein extraction, fresh testis removed from the albuginea was sheared into pieces in PBS and washed with PBS 3 times. Then, total protein was extracted from the pieces with western blotting and IP lysis buffer. Finally, the total proteins were used for western blotting. The antibodies used in the co-IP and western blots are listed in key resources table.

Min Z., Xin H., Liu X., Wan J., Fan Z., Rao X., Fan J., Yang L, & Li D. (2022). Chromodomain helicase DNA-binding domain 2 maintains spermatogonial self-renewal by promoting chromatin accessibility and mRNA stability. iScience, 25(12), 105552.

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Endogenous and Exogenous Co-Immunoprecipitation Assay

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For co-immunoprecipitation (co-IP) assays, endogenous verification was performed in conjunction with exogenous expression. For exogenous verification, PAMs (10⁵ cells) were cotransfected with 2 μg of NS3-EGFP and 2 μg of CMV-TRAF6 plasmids. After 36 h, the cells were washed with PBS and were harvested with Western blot and IP lysis buffer (Beyotime, China) containing PMSF. After centrifugation for 30 min at 4 °C, a quarter of the supernatant was subjected to input assays. The rest were used for co-IP experiments with ANTI-FLAG M2 Affinity Gel (Sigma, USA) according to the manufacturer’s instructions. In brief, 50 μL of resin stored in 50% glycerol was centrifuged for 30 s at 9000 × g and rinsed twice with 1 mL of TBS. The cell lysate was added to the equilibrated resin and rocked gently overnight at 4 °C. The resin was washed three times with 1 mL of TBS and resuspended with 2 × SDS sample buffer for Western blot with anti-GFP and anti-Flag antibodies. For further endogenous verification, PAMs (10⁵ cells) were transfected with 4 μg of CMV-NS3 plasmid, and the samples were detected with rabbit anti-TRAF6 and anti-Flag pAb. The other steps were similar to those of exogenous verification.

Lv H., Dong W., Cao Z., Li X., Wang J., Qian G., Lv Q., Wang C., Guo K, & Zhang Y. (2017). TRAF6 is a novel NS3-interacting protein that inhibits classical swine fever virus replication. Scientific Reports, 7, 6737.

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