Endocrinology, Vol 134, 1960-1963, Copyright © 1994 by Endocrine Society

ARTICLES

Direct detection of insulin-like growth factor II (IGF-II) by chemiluminescence without interference by IGF binding proteins

DD De Leon, C Terry and SP Nissley
National Institutes of Health, National Cancer Institute, Metabolism Branch, Bethesda, MD 20892.

A dot blot method for the detection of picogram quantities of human and rat insulin-like growth factor II (IGF-II) in serum-free conditioned media is described. The crossreactivity of human recombinant IGF-I in the assay was < 10%. None of the IGF binding proteins (IGFBP 1-6) diminished the IGF-II signal. In contrast, significant interference by the IGFBPs was observed when the same concentrations of IGFBPs and 125I- IGF-II were used in a radioimmunoassay which utilized the same antibody. Why IGF-II is detected in the dot blot assay without IGFBP interference is not understood. We speculate that the conformation of the IGF-II/binding protein complex may be altered by binding to the nitrocellulose, exposing the IGF-II epitope that is recognized by the antibody. IGF-II was detected in 1 microliter of serum-free conditioned media from BRL 3A cells (which secrete IGF-II) while no signal was generated by 50 microliters of BRL 3A2 conditioned media (which do not secrete IGF-II). In summary, this method is ideal for screening cells in serum free-culture for production of IGF-II without the need for separation of IGF-II from cell derived IGFBPs.