Cells were homogenized with ice-cold RIPA buffer for 30 min to provide whole cell lysates. Concentrations of total proteins were determined using enhanced BCA protein assay kit (TransGen Biotech, Beijing, China). Lysates were separated by 8%–12% SDS-PAGE and subsequently transferred to polyvinylidene fluoride (PVDF) membranes. Membranes were blocked in 5% non-fat milk, probed with primary antibody (1:1000 diluted) for 2 h, and further incubated with secondary antibody (1:1000 diluted) for 1 h. Then, membranes were washed and developed with enhanced chemiluminescence (ECL) substrate. Images were captured using Tanon 5200 Imaging Analysis System (Tanon, Shanghai, China). Relative protein expressions were shown by densitometry analysis using Image J software.
Enhanced bca protein assay kit
Manufactured by Transgene
Sourced in China
The Enhanced BCA protein assay kit is a colorimetric assay used to determine the concentration of protein in a sample. It is based on the reduction of copper ions by proteins in an alkaline medium, which results in the formation of a purple-colored complex that can be measured using a spectrophotometer.
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Lab products found in correlation
Antibodies against cleaved caspase 3, by Cell Signaling Technology (1 mentions)
Dulbecco s modified eagle s medium dmem, by PAN Biotech (1 mentions)
3 4 5 dimethylthiazol 2 yl 2 5 diphenyltetrazolium bromide mtt, by Merck Group (1 mentions)
α tubulin, by Cell Signaling Technology (1 mentions)
Histone h3, by Cell Signaling Technology (1 mentions)
Rabbit igg, by Cell Signaling Technology (1 mentions)
Foxo3a, by Cell Signaling Technology (1 mentions)
Foxo1, by Cell Signaling Technology (1 mentions)
Fetal bovine serum (fbs), by PAN Biotech (1 mentions)
Hoechst 33258, by Solarbio (1 mentions)
Lactate dehydrogenase ldh assay kit, by Nanjing Jiancheng (1 mentions)
2 protocols using enhanced bca protein assay kit
1
Western Blot Analysis of Protein Expression
2
BSHX Granules and AGEs Effects
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BSHX granules were purchased from Beijing Kangrentang Pharmaceutical Co. Ltd. (Beijing, China). The medicinal components of BSHX are listed in Table 1 . AGEs were purchased from Beijing Biosynthesis Biotechnology Co. Ltd. (Beijing, China) with a purity greater than 95% by high-performance liquid chromatography (HPLC). Antibodies against cleaved caspase-3, Bim, Histone H3, FoxO1, FoxO3a, α-Tubulin, and rabbit IgG were obtained from Cell Signaling Technology (Boston, MA, USA). 3-(4,5-Dimethylthiazol-2-yl)2,5-diphenyltetrazolium bromide (MTT) was bought from Sigma-Aldrich Chemical Co. (Saint Louis, MO, USA). The Dulbecco's modified eagle's medium (DMEM) and fetal bovine serum (FBS) were purchased from PAN-Biotech GmbH (Aidenbach, Germany). The endothelial cell medium (ECM) was obtained from Beijing M&C Gene Technology Ltd. (Beijing, China). The lactate dehydrogenase (LDH) assay kit was purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing, China). Hoechst 33258 was obtained from Beijing Solarbio Science & Technology Co. Ltd. (Beijing, China). The enhanced BCA protein assay kit was obtained from Beijing TransGen Biotech Co. Ltd. (Beijing, China).
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