Identification of an insulin-like growth factor-binding protein in human cerebrospinal fluid with a selective affinity for IGF-II.

Human cerebrospinal fluid (CSF) has been found to contain several different molecular forms of IGF-specific binding proteins (BPs). Qualitatively, they are similar to those present in serum, although their relative proportions are very different, as well as to those present in the culture media of brain tissue from which these BPs presumably arise. One particular form of BP is predominant in CSF. It has an Mr of 34,000, as estimated by SDS-polyacrylamide gel electrophoresis followed by transfer onto nitrocellulose, and an isoelectric point around 5.0 based on chromatofocusing. It has a selective affinity for IGF-II (approximately 4 X 10(10) M-1) as shown by competitive binding experiments in which biosynthetic IGF-I was about 40-times less potent than native IGF-II in displacing 125I-labelled IGF-II. These findings are in agreement with the preponderance of IGF-II in nervous tissue and in CSF and suggest that this BP plays an important role in the interaction of IGF-II with its target cells.

Analysis of serum insulin-like growth factor binding proteins using western blotting: use of the method for titration of the binding proteins and competitive binding studies.

A nitrocellulose gel transfer technique has been adapted to study the insulin-like growth factor (IGF) binding proteins of human serum. Normal and hypopituitary sera were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, followed by electroblotting to nitrocellulose or nylon membrane. Nonidet-P40 (3%) and Tween 20 (0.1%) were required for quenching and to allow detection of the IGF binding proteins by autoradiography after overlay with either 125I-labeled IGF I or IGF II. Several forms of IGF binding protein have been identified with molecular weights of 41,500, 38,500, 34,000, 30,000, and 24,000. Titration and competitive binding studies with IGF were performed on the transferred IGF binding proteins, indicating that binding proteins isolated by this technique can be characterized.