Quantification by affinity perfusion chromatography of phosphorylated BRCAl and BRCA2 proteins from tumor cells after lycopene treatment - Archive ouverte HAL Accéder directement au contenu
Article Dans Une Revue Journal of Chromatography B - Analytical Technologies in the Biomedical and Life Sciences Année : 2005

Quantification by affinity perfusion chromatography of phosphorylated BRCAl and BRCA2 proteins from tumor cells after lycopene treatment

Résumé

A new procedure for the quantification of phosphorylated BRCA1 (P-BRCA1) and BRCA2 (P-BRCA2) proteins in breast cell lines after different treatments was carried out. Cells were cultivated with [35S]-methionine and extracts subjected to three perfusion chromatographies. First heparin affinity chromatography purified cellular DNA-binding proteins. Subsequent specific immunoprecipitation of BRCA1 and BRCA2 proteins was performed with antibodies raised against BRCA1 or BRCA2. The immune complexes were isolated by protein A affinity chromatography. Phosphorylated BRCA1 or BRCA2 proteins were then purified with a Poros 20 AL column where anti-phosphothreonine and anti-phosphoserine antibodies were previously bound. The percentage of phosphorylated BRCA1 or BRCA2 proteins was calculated as follows: 100 × dpm of P-BRCA1 or P-BRCA2 eluted from the POROS® 20AL column/total dpm eluted from POROS® 20AL column. Treatment with 10 μM lycopene increased P-BRCA1 and P-BRCA2 in the breast tumor cell line MCF7 but not in MDA-MB-231 or MCF-10a, breast tumor or fibrocystic cell lines, respectively.

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Cancer

Dates et versions

hal-02071222 , version 1 (18-03-2019)

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Citer

Nassera Chalabi, Jean-Claude Maurizis, Ludovic Le Corre, Laetitia Delort, Yves-Jean Bignon, et al.. Quantification by affinity perfusion chromatography of phosphorylated BRCAl and BRCA2 proteins from tumor cells after lycopene treatment. Journal of Chromatography B - Analytical Technologies in the Biomedical and Life Sciences, 2005, 821 (2), pp.188-193. ⟨10.1016/j.jchromb.2005.05.006⟩. ⟨hal-02071222⟩

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