The treated GBC cell lines were harvested and lysed in RIPA buffer (Cell Signaling, Danvers, USA). The protein concentration determined by the bicinchoninic acid (BCA) protein reagent (Beyotime) Equal amounts of protein samples were run on the 10% SDS-PAGE and transferred onto polyvinylidene difluoride (PVDF) membranes. The membranes were probed with primary antibodies against human Src, p-Src(Y416), T-Akt, p-Akt(Ser473), Bcl-2, Bax, or GAPDH (Cell Signaling Technology, USA). After incubated with the secondary antibodies (HRP-conjugated goat anti-rabbit or goat anti-mouse IgG), the signal bands were visualized by enhanced chemiluminescence (ECL) western blotting detection reagent. GAPDH was served as the loading control.
Bca protein reagent
Manufactured by Beyotime
Sourced in China
The BCA protein reagent is a colorimetric detection and quantification method used to measure the total protein concentration in a sample. It is based on the reduction of copper ions (Cu2+) to cuprous ions (Cu+) by proteins in an alkaline environment, followed by the chelation of the cuprous ions with bicinchoninic acid (BCA) to produce a purple-colored complex that can be measured spectrophotometrically.
Automatically generated - may contain errors
Lab products found in correlation
Ripa buffer, by Cell Signaling Technology (2 mentions)
Bcl2 associated x (bax), by Cell Signaling Technology (1 mentions)
Gapdh, by Cell Signaling Technology (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
3 4 5 dimethylthiazol 2 yl 2 5 diphenyltetrazolium bromide mtt, by Thermo Fisher Scientific (1 mentions)
Trizol reagent, by Thermo Fisher Scientific (1 mentions)
Phenylmethanesulfonyl fluoride pmsf, by Thermo Fisher Scientific (1 mentions)
Sybr green real time pcr master mix, by Takara Bio (1 mentions)
Pierce ecl western blotting substrate, by Thermo Fisher Scientific (1 mentions)
Nuclear and cytoplasmic protein extraction kit, by Beyotime (1 mentions)
Radioimmunoprecipitation assay (ripa), by Applygen (1 mentions)
4 protocols using bca protein reagent
1
Western Blot Analysis of GBC Cells
2
Gallic Acid Effects on LDL Metabolism
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Gallic acid (GA) (HPLC ≥ 98%) from Shanghai yuanye Bio-Technology Co., Ltd., Shanghai, China; Trypsin–EDTA (0.25%), penicillin–streptomycin, phenylmethanesulfonyl fluoride (PMSF), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), and Trizol reagent from Thermo Fisher Scientific, Shanghai, China; Dulbecco’s modified Eagle’s medium (DMEM) and Fetal Bovine Serum (FBS) from MeilunBio; 4′,6-diamidino-2-phenylindole (DAPI) and Human 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanide perchlorate–low-density lipoprotein (Human Dil-LDL) from Yeasen Biotechnology (Shanghai) Co., Ltd., Shanghai, China; SYBR Green real-time PCR Master Mix from Takara Biomedical Technology (Beijing) Co., Ltd., Beijing, China; Kit for the nuclear extraction of the cell from Beijing Solarbio Science & Technology Co., Ltd., Beijing, China; BCA Protein Reagent from Shanghai Beyotime Biotechnology Co., Ltd., Shanghai, China; anti-LDLR, ERK1/2, phospho-ERK1/2, EGFR, phospho-EGFR, and β-tubulin from HuaBio, Hangzhou, China; and Anti-PCSK9, HNF1α, FOXO3, and SREBP-2 from Wuhan ABclonal Biotechnology Co., Ltd., Wuhan, China.
3
Bcl-2 Protein Detection by Western Blot
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Western blotting assay was performed as described previously [13] . In brief, the cells were harvested and lysed in RIPA buffer (Cell Signaling, Danvers, USA). The protein concentration determined by the bicinchoninic acid (BCA) protein reagent (Beyotime) Equal amounts of protein samples were run on the 10% SDS-PAGE and transferred onto polyvinylidene difluoride (PVDF) membranes. The membranes were probed with primary antibodies against human Bcl-2 and GAPDH (Santa Cruz, USA). After incubated with the secondary antibodies (HRP-conjugated goat anti-rabbit or goat anti-mouse IgG) and the signal bands were visualized by chemiluminescence using Pierce ECL Western Blotting Substrate (Thermo Fisher Scientific, USA).
4
Protein Extraction and Quantification Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total protein was extracted from L02 cells and liver tissue samples by radioimmunoprecipitation assay lysis buffer (RIPA; Applygen, China; cat: C1053). The nuclear and cytoplasmic protein extraction kit (Beyotime, China; catalog: P0028) was used to separate cytoplasmic and nuclear proteins. Protein concentration was determined with bicinchoninic acid (BCA) protein reagent (Beyotime, China; catalog: P0012). The protein lysate was electrophoresed, transferred to the membrane, and blocked with nonfat milk powder. The primary antibody was incubated overnight at 4°C, and after washing with TBST, the secondary antibody was incubated for 1 hours. Finally, a chemiluminescence reagent (NCM Biotech, China; cat: P10100) was used to detect the reaction. An ImageQuantLAS4000 (GE Co., USA) chemiluminescence imaging system was used to acquire visual images of target proteins. Quantify protein expression using Image J software version 1.80, and expressed as a percentage of the control.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!