Co-immunoprecipitation was performed according to a previous report.16 Briefly, proteins were extracted with cell lysis buffer (KGP701~KGP701–100, KeyGEN BioTECH, China) with 1% PMSF (KGP610, KeyGEN BioTECH, China). A total of 500 μg of protein extract was then incubated with 2 μg of appropriate immunoprecipitation (IP) antibodies, i.e., rabbit anti-Thy-1 (ab225, Abcam, US) or mouse anti- integrin β3 (sc-46655, Santa Cruz, USA) at 4 °C overnight with shaking. Then, 20 μl of protein A/G plus agarose (Thermo) was added, followed by incubation at 4 °C for 3 h with shaking. The pellets were washed five times with cell lysis buffer for 2 min and resuspended in 40 μl of 2 × electrophoresis loading buffer. After boiling for 10 min, 20 μl samples were used for SDS-PAGE and assessed by Western blot as described above.
Kgp701 100
Manufactured by Keygen Biotech
Sourced in China
The KGP701–100 is a high-precision laboratory centrifuge capable of processing up to 100 samples simultaneously. It features a compact design and advanced temperature control system to ensure consistent and reliable results.
Automatically generated - may contain errors
Lab products found in correlation
Ab225, by Abcam (1 mentions)
Protein a g plus agarose, by Thermo Fisher Scientific (1 mentions)
Sc 46655, by Santa Cruz Biotechnology (1 mentions)
Kgp701, by Keygen Biotech (1 mentions)
Kgp610, by Keygen Biotech (1 mentions)
Nb600 1335, by Novus Biologicals (1 mentions)
A0208, by Beyotime (1 mentions)
2 protocols using kgp701 100
1
Co-Immunoprecipitation Protocol for Protein Interactions
2
Western Blot Analysis of ER Stress Proteins
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Cells were washed once with PBS and lysed by cell lysis buffer (KeyGEN, KGP701‐100) containing 20 mM Tris (pH 7.5), 150 mM NaCl, 1% Triton X‐100 and protease inhibitors for 30 min ice, and then ultrasonicated. The samples were boiled to 100℃ for 10–15 min in loading buffer (EpiZyme, LT101S) with 2% β‐mercaptoethanol (Amersham, CT). Anti α‐TUBULIN (1:10000, Proteintech, 66031‐1‐Ig) was used as endogenous control and anti‐SURF4 (1:1000, Bioswamp, PAB44330), anti‐XBP1 (1:1000, ABclonal, A1731) and anti‐DDIT3 (1:1000, Novus, NB600‐1335) was used. ECL peroxidase‐labelled sheep anti‐mouse antibody (GE Healthcare, NA931V) or HRP‐labelled goat anti‐rabbit antibody (Beyotime, A0208) were used as secondary antibodies.
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