Development of a heat-mediated protein blotting method

O'Sullivan, Jack; McMahon, Hilary E.M.

doi:10.1016/j.ab.2016.01.026

Cited by 1 publication

(1 citation statement)

References 21 publications

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“…Next, cells were lysed using RIPA and proteins were collected after centrifuging at 13,000 r/min, 4 ℃ for 20 min. The concentration of protein was determined using Bradford Protein kit [56]. Protein samples were mixed with 1× reducing electrophoresis loading buffer and boiled for 5 min [57].…”

Section: Western Blot Analysismentioning

confidence: 99%

Seleno-short-chain chitosan induces apoptosis in human breast cancer cells through mitochondrial apoptosis pathway in vitro

Zhao

et al. 2018

Cell Cycle

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Seleno-short-chain chitosan (SSCC) was a synthesized chitosan derivative with the molecular weight of 4826.986 Da. The study is aimed to investigate cytotoxicity of SSCC on human breast cancer MCF-7 and BT-20 cells and explore apoptosis-related mechanism in vitro. The MTT (3- [4,5-Dimethylthiazol-2-yl]-2, 5-diphenylterazolium bromide) assay showed that SSCC exhibited significantly cytotoxic effects on MCF-7 and BT-20 cells in a dose- and time-dependent manner, and the effective inhibitory concentration was 100 μg/ml and 200 μg/ml, respectively. Apoptosis assay of these two kinds of cells was determined by Hoechst 33,342/PI and Annexin V-FITC/PI double staining. The cell cycle assay showed that SSCC triggered S and G2/M phase cell cycle arrest in MCF-7 cells and S phase cell cycle arrest in BT-20 cells in a time-dependent manner. Further studies demonstrated that SSCC led to the generation of reactive oxygen species (ROS) and the disruption of mitochondrial membrane potential (MMP) in these two kinds of cells. N- acetyl-L cysteine (NAC), as a radical scavenger, significantly inhibited the generation of ROS and decreased the apoptosis of MCF-7 and BT-20 cells. Moreover, the expression of mitochondrial apoptosis-related proteins was detected by western blot assay. SSCC up-regulated the expression of Bax, down-regulated the expression of Bcl-2, subsequently increased the release of cytochrome c from mitochondria to cytoplasm, and activated the cleavage of caspase-9 and -3, which finally induced apoptosis in MCF-7 and BT-20 cells in vitro. Consequently, these data indicated that SSCC could induce apoptosis of MCF-7and BT-20 cells in vitro by mitochondrial pathway.

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Section: Western Blot Analysismentioning

confidence: 99%

Seleno-short-chain chitosan induces apoptosis in human breast cancer cells through mitochondrial apoptosis pathway in vitro

Zhao

et al. 2018

Cell Cycle

View full text Add to dashboard Cite

show abstract

Development of a heat-mediated protein blotting method

Cited by 1 publication

References 21 publications

Seleno-short-chain chitosan induces apoptosis in human breast cancer cells through mitochondrial apoptosis pathway in vitro

Seleno-short-chain chitosan induces apoptosis in human breast cancer cells through mitochondrial apoptosis pathway in vitro

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