Analysis of mitogenic activity of proteins after separation by gel electrophoresis

• Published in: Cytotechnology (Dordrecht) Vol. 30; no. 1-3; pp. 235 - 240
• Main Authors: HOHENWARTER, O, MARZBAN, G, JISA, E, VORAUER-UHL, K, KATINGER, H
• Format: Conference Proceeding Journal Article
• Language: English
• Published: Dordrecht Springer 01.07.1999; Springer Nature B.V; Kluwer Academic Publishers
• Subjects: Biological and medical sciences; Biotechnology; Cell culture; Colorimetry; Endothelial cells; Fundamental and applied biological sciences. Psychology; Gel electrophoresis; Interleukin 2; Isoelectric focusing; Methods. Procedures. Technologies; Others; Pituitary; Proteins; Tissue culture; Various methods and equipments; Human; Vertebrata; Endothelial cell; Mammalia; Bioassay; Bovine; Gel electrophoresis; Pituitary gland; Mitogen; Artiodactyla; Ungulata; Growth factor
• ISSN: 0920-9069; 1573-0778
• DOI: 10.1023/a:1008063327828

We have used a combination of gel electrophoresis and a cell culture assay in microplates to analyse mitogenic activity in tissue extracts. The procedure is a modification of the method described by Kuo et al. The proteins were separated by native gel electrophoresis or isoelectric focusing. The gel was sliced and defined pieces were transferred into tissue culture inserts fitting in 96 well microplates, which contained the test cells. The proteins diffused from the gel slices directly into the culture supernatant and the mitogenic effects were evaluated by a colorimetric assay (MTT or phosphatase activity). Human interleukin 2 was used to demonstrate the feasibility of the method by evaluating the mitogenic effect on the cell line CTLL-2. Extracts of bovine pituitary glands were separated by native gel electrophoresis and isoelectric focusing and several protein bands could be identified which showed a distinct mitogenic effect on human endothelial cells. The method is very sensitive and allows rapid screening of protein mixtures for bioactive fractions.