Lab Report - Protein Separation

Biomarker discovery in human breast cancer cells Lab based Practical Molecular Pharmacology BHS006-6 Breast cancer is the most common cancer type among women and the ability to identify potential biomarkers or hallmarks of early disease is critical for early detection and treatment. The aim of this lab based practical session is to utilise an established protein separation technique (SDS-PAGE) to identify possible biomarkers, which are present in 100pM Estradiol, treated breast cancer cell homogenates versus control cells. Students will run protein samples on SDS-PAGE gel as per protocol below to demonstrate protein separation and identify differing bands, between control and treated cells, which may be biomarkers of disease. Equipment & Materials: Running buffer [0.025M Tris/0.192M glycine/0.1% (w/v) SDS, pH8.3] Pre-stained molecular markers (PageRuler) Coomassie Blue Stain Solution Human breast cancer cell homogenates: a) control cells; b) cells treated with 100pM 17β-estradiol Human recombinant histone proteins: H2B; H3.3 [1μg (1μl) added to 20μl sample buffer] Experiment Procedures: Preparation of Sodium Dodecyl Sulphate-Polyacrylamide (SDS-PAGE) gel will be undertaken by laboratory staff prior to the practical. The protocol for the preparation of the gel can be found at the end of this document.