NPC cells were lysed in RIPA lysis buffer (Meilun Biotechnology Co., Ltd., Dalian, China) containing protease inhibitor cocktails (Fudebio, Hangzhou, China). Then, total protein from different samples (30 μg/per lane) was separated by SDS-PAGE and transferred onto 0.22 μM PVDF membranes (Amersham Bioscience, Piscataway, NJ, United States). After that, the membranes were blocked with 5% skimmed milk in TBST for 1 h and incubated with specific primary antibody overnight at 4°C. After being washed with TBST, the membranes were incubated with the secondary antibody for 1 h at room temperature. Next, the protein bands were developed using an enhanced chemiluminescence kit (FD8030, Fudebio, Hangzhou, China). Antibodies were listed: anti-E-cadherin (Abcam, ab76319, 1:2,000), anti-Vimentin (Abcam, ab92547, 1:2,000), anti-PIK3R1 (Abcam, ab191606, 1:1,000), anti-ERBB2 (Abcam, ab237715, 1:1,000), anti-GAPDH (Abcam, ab8245, 1:3,000), goat anti-mouse IgG H&L (HRP) (Abcam, ab205719, 1:5000), and goat anti-rabbit IgG H&L (HRP) (Abcam, ab205718, 1:5,000).
Protease inhibitor cocktails
Manufactured by Fudebio
Sourced in China
Protease inhibitor cocktails are a type of lab equipment used to inhibit the activity of proteases, which are enzymes that break down proteins. These cocktails contain a mixture of chemical compounds that target and inactivate different classes of proteases, helping to preserve the integrity of protein samples during various experimental procedures.
Automatically generated - may contain errors
Lab products found in correlation
Ripa lysis buffer, by Meilun (2 mentions)
Ab76319, by Abcam (1 mentions)
Ab237715, by Abcam (1 mentions)
Ab205719, by Abcam (1 mentions)
Ab205718, by Abcam (1 mentions)
Fd8030, by Fudebio (1 mentions)
Ab191606, by Abcam (1 mentions)
Pvdf membrane, by Cytiva (1 mentions)
Ab92547, by Abcam (1 mentions)
Ab8245, by Abcam (1 mentions)
Anti hk2, by Abcam (1 mentions)
Pvdf membrane, by Abcam (1 mentions)
Anti stat3, by Abcam (1 mentions)
2 protocols using protease inhibitor cocktails
1
Western Blot Analysis of EMT Markers
2
Protein Expression Analysis via Western Blot
Check if the same lab product or an alternative is used in the 5 most similar protocols
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GC cells were lysed for protein lysis on ice using radioimmunoprecipitation assay (RIPA) lysis buffer (Dalian Meilun Biotechnology, China) containing protease inhibitor cocktails (Fudebio, Hangzhou, China). Total protein from each sample was separated by SDS-PAGE gels and transferred onto polyvinylidene fluoride (PVDF, 0.22-μm) membranes (Amersham Bioscience, Piscataway, NJ, USA). Subsequently, the PVDF membranes were blocked with skim milk powder (5%) in Tris-buffered saline with Tween 20 (TBST) at room temperature and incubated with primary antibodies (anti-STAT3, anti-HK2, Abcam, 1:1,000) overnight at 4°C. Then, the membranes were washed with TBST and incubated by horseradish peroxidase (HRP)-conjugated secondary antibody (anti-ACTINB). Quantitative analysis of band intensity was determined using ImageJ software.
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