Enzyme-linked immunosorbent assay for pharmacological studies targeting hypoxia-inducible factor 1 alpha

Abstract

Hypoxia-inducible factor 1 (HIF-1) activates the transcription of a wide range of genes related to oxygen delivery and metabolic adaptation under hypoxic (low-oxygen) conditions. HIF-1 is, in fact, a heterodimer of two subunits, HIF-1 alpha and HIF-1 beta. The only analytical methods available for measuring HIF-1 alpha levels in tumors are immunohistochemistry and Western blotting. Immunohistochemistry has the advantage of allowing the identification and direct examination of HIF-1 alpha-expressing cells, but has the intrinsic limitation, as for Western blotting, of being nonquantitative. We developed and validated an enzyme-linked immunosorbent assay (ELISA) approach to measure HIF-1 alpha levels in cultured tumor cell lines in vitro. HIF-1 alpha was expressed in thirteen tumor cell lines grown under hypoxic conditions; however, the levels differed strongly between cell lines. These data point to intrinsic differences between cell lines for the induction of HIF-1 alpha under hypoxic conditions. The ELISA developed in the present study is thus an interesting alternative to other analytical methods used to measure HIF-1 alpha protein levels and should be useful in preclinical pharmacological studies targeting HIF-1 alpha.