Isolation of EGF-dependent transforming growth factor (TGFβ-like) activity from culture medium conditioned by fetal rat calvariae

Open Access
Publisher Website
Google Scholar
Abstract
A transforming growth factor of the beta class (TGF-β), defined by its ability to induce normal rat kidney cells (NRK, clone 49F) to form anchorage-independent large colonies in soft agar in the obligate presence of epidermal growth factor, has been prepared from culture medium conditioned by fetal rat calvariae. This activity was purified by acetic acid extraction, gel permeation chromatography, and two reversed-phase HPLC (rpHPLC) steps. Bone culture derived-TGFβ-like activity was soluble in 1.0 M acetic acid, eluted from Sephadex G-75 at relative molecular mass (Mr) 25,000, from μBondapak C18 rpHPLC at 63 ± 5% methanol in 0.1 M acetic acid, and from μBondapak CN rpHPLC at 36 ± 2% n-propanol in 0.1% trifluoroacetic acid. Based on specific activity estimations at each stage of purification, TGFβ-like activity was purified 2500-fold with a 14% recovery, and 1 I of conditioned medium yielded 1–2 μg of factor. Silver-stained polyacrylamide gels of this material after CN μBondapak rpHPLC revealed a predominant band of (Mr) 24,000.
Funding Information
  • NIAMSD (AM21707)

This publication has 22 references indexed in Scilit: