Purification and Characterization of Transforming growth factor - beta1 from Human Platelets.
- Author:
Eun Jo KO
;
Jong Won LEE
;
Sang Uk NHAM
;
Eui Yul CHOI
;
Gie Taek CHUN
;
Se Won YIE
;
Pyeung Hyeun KIM
- Publication Type:In Vitro ; Original Article
- Keywords:
Human platelet;
TGF-p1;
ELISA;
Bioassay;
Cell cycle;
igA
- MeSH:
B-Lymphocytes;
Biological Assay;
Blood Platelets;
Cell Cycle;
Cell Differentiation;
Chromatography;
Chromatography, Gel;
Chromatography, High Pressure Liquid;
Chromatography, Liquid;
Electrophoresis;
Enzyme-Linked Immunosorbent Assay;
Humans*;
Immunoglobulin A;
Lung;
Mink;
Molecular Weight;
Silver Staining;
Spleen;
Transforming Growth Factors*
- From:Korean Journal of Immunology
1998;20(1):1-8
- CountryRepublic of Korea
- Language:Korean
-
Abstract:
Transforming growth factor-j31 (TGF-p1) has potential for therapeutic use in common clinical conditions for which there are no adequate pharmacological agents. However, in vivo studies using TGF-p1 were hindered by high price of this cytokine. As a first step towards large scale purification of TGF-p1, it was purified in a small scale (10 unit platelets) from human platelets by four purification steps: platelet extraction, gel filtration, cation exchange chromatography, and reversed phase high performance liquid chromatography (HPLC). A single protein band with a molecular weight of 25 Kd corresponding to purchased TGF-p1 (R8D Systems) was confirmed by silver staining after SDS-polyacrylamide gel electrophoresis (SDS-PAGE) of eluant from reversed phase HPLC. Recovery (%) of each step was about 50-60%, resulting in the final recovery of 20% based on the detection by a sandwich ELISA. Approximately, 3.7 p,g of purified TGF-p1 was obtained from 18 pg of platelet extracts. This result was confirmed by receptor (TGF-j31 type II) ELISA and bioassay using a mink lung epithelial'cell line (MV1LU). Further, in vitro characterization study showed that purified TGF-p1 inhibits G1/S transition of LPS-activated murine spleen B cells and increases surface IgA expression by the same cell population, which are typical activities of TGF-p1 in B cell differentiation. Taken together, the results from the present study reveals that purified TGF-p1 is fully biologically active and our purification methodology could be usbful to obtain a large scale of recombinant TGF-p1 in the future.
