The Co-IP experiments were performed to determine the interaction between RCN2 and CALR using a Co-IP assay kit (#26,146; Thermo Fisher Scientific). CNE2 and 5-8 F cells were lysed in 500 µL solubilization Co-IP buffer. We use 10–50 µg of protein for positive control. Then, 30 µL of RCN2 (10193-2-AP; Proteintech, Illinois, USA) or 26 µL of CALR (PA3-900; Invitrogen) were added to the rest of the supernatant for batch binding. After the beads were collected and washed 5 times, we eluted the beads with 200 µL of spent regenerant.
Co ip assay kit
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Co-IP assay kit is a laboratory tool designed to facilitate the identification and study of protein-protein interactions. It provides the necessary reagents and protocols to perform co-immunoprecipitation experiments, which are a widely used technique in molecular biology and biochemistry research.
Automatically generated - may contain errors
Lab products found in correlation
Enhanced chemiluminescence ecl kit, by Santa Cruz Biotechnology (2 mentions)
Cell cycle detection kit, by Keygen Biotech (2 mentions)
Pa3 900, by Thermo Fisher Scientific (1 mentions)
Primescript 2 1st strand cdna synthesis kit, by Takara Bio (1 mentions)
Trizol reagent, by Thermo Fisher Scientific (1 mentions)
Ros assay kit, by Beyotime (1 mentions)
Tunel assay kit, by Roche (1 mentions)
Anti keap1, by Proteintech (1 mentions)
P gsk3β ser9, by Cell Signaling Technology (1 mentions)
Ml364, by MedChemExpress (1 mentions)
Anti collagen 3 antibody, by Abcam (1 mentions)
Ang 2, by Merck Group (1 mentions)
Gsk3β, by Cell Signaling Technology (1 mentions)
Mg132, by Enzo Life Sciences (1 mentions)
4 protocols using co ip assay kit
1
Co-IP Assay for RCN2-CALR Interaction
2
Coimmunoprecipitation of PGC-1α and ERRα
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Coimmunoprecipitation was conducted using a co-IP assay kit (Thermo Fisher Scientific, USA), and the total protein concentrations of the cell lysates were measured by BCA assay. An anti-PGC-1α antibody (see Table S1 ) was used to pull down PGC-1α, and an anti-IgG antibody was used as a negative control. The protein collected was analysed by Western blot analysis using an anti-ERRα antibody (see Table S1 ) to detect the co-combination between PGC-1α and ERRα.
3
Investigating USP7-mediated Nrf2 Regulation
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P5091 was produced by Selleck (USA). Cell signaling technologies provided anti-USP7 and anti-bax antibodies (USA). Proteintech (China) produced anti-Keap1. Bioworld (Bioworld Technology, USA) provided anti-Nrf2 and anti-GAPDH antibodies. Antibodies produced against ubiquitin were bought from Life Technologies, as was the Co-IP assay kit (USA). Invitrogen produced the TRIzol reagent (USA). Takara Biomedical Technology (Beijing) Co., Ltd. provided the PrimeScript II 1st Strand cDNA Synthesis Kit. Keygen created the Cell Cycle Detection Kit (China). Santa Cruz Biotechnology provided the enhanced chemiluminescence (ECL) kit (USA). ROS Assay Kit was bought from Beyotime Biotechnology (China). Cusabio Technology (China) provided ELISA Kit. TUNEL Assay Kit was acquired from Roche (China). For immunoblot, immunofluorescence, and Co-IP assays, antibodies were diluted at 1:1000, 1:2500, 1:500, and 1:50, respectively.
4
Signaling Pathways in Fibroblast Regulation
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ML364 was manufactured by MedChemExpress (USA). Ang II was provided by Sigma‐Aldrich (USA). MG132 was a product of Enzo (USA). Anti‐USP2 and anti‐periostin antibodies were purchased from Proteintech (China). Anti‐collagen‐III antibody was manufactured by Abcam (United Kingdom). Anti‐CTGF and anti‐GAPDH antibodies were obtained from Boster (China) and Bioworld (Bioworld Technology, USA), respectively. Antibodies raised against ubiquitin, cyclin D1, P27, β‐catenin, GSK‐3β and P‐ GSK‐3β(Ser9) were provided by Cell Signaling Technology (USA). MTS assay reagents were manufactured by Promega (USA). The Co‐IP assay kit was purchased from Life Technologies (USA). Cell Cycle Detection Kit was manufactured by Keygen (China). The enhanced chemiluminescence (ECL) kit was provided by Santa Cruz Biotechnology (USA). Antibodies were diluted at 1:1000, 1:500 and 1:50 for immunoblot, immunofluorescence and Co‐IP assay, respectively.
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