A strategic approach for the visualisation of the low abundance protein fatty acid binding protein 7

Dean, Sarah JR; Perks, Claire CM; Rhodes, Anthony; Holly, Jeff MP; Jarrett, Caroline

Abstract

Fatty acid binding protein 7 (FABP7) is a 15kDa protein that plays a role in fatty acid transport, solubilisation and metabolism. It has been found to be overexpressed in triple negative/basal like breast cancer patients. The role of FABP7 in these breast cancers is not fully understood; in order to better understand how it may be involved with breast cancer prognosis cell line models are needed for mechanistic studies. However using a standard western blotting protocol FABP7 was not detectable in a selection of breast cancer cell lines and mRNA was in low abundance. This paper outlines the approach of modifying a western blot protocol and presents an optimised western blot protocol for the detection of FABP7 in breast cancer cell lines. The main areas considered during optimisation were, titration of primary and secondary antibodies, choice of protein lysis buffer, lysate preparation and a comparison between in-house 12% polyacrylamide gels and commercially available gradient polyacrylamide gels. This strategy could be used for other low abundant and small proteins.

• Journal Article Type: Article
• Publication Date: Jan 1, 2015
• Deposit Date: Mar 10, 2015
• Publicly Available Date: Apr 17, 2016
• Journal: American Journal of Biomedical Science
• Print ISSN: 1937-9080
• Peer Reviewed: Peer Reviewed
• Volume: 7
• Issue: 1
• Pages: 15-25
• DOI: https://doi.org/10.5099/aj150100015
• Keywords: FABP7, western-blotting, breast cancer, protein, lysis
• Public URL: https://uwe-repository.worktribe.com/output/841610
• Publisher URL: http://dx.doi.org/10.5099/aj150100015
• Contract Date: Apr 17, 2016