Elsevier

Cytokine

Volume 7, Issue 5, July 1995, Pages 389-397
Cytokine

Regular Article
Characterization of recombinant soluble human transforming growth factor-β receptor Type II (rhTGF-βsRII)

https://doi.org/10.1006/cyto.1995.0054Get rights and content

Abstract

Recombinant human transforming growth factor soluble receptor Type II (rhTGF-βsRII) corresponding to the 159 amino acid extracellular domain of hTGF-βRII has been expressed in insect cells using the baculovirus expression system or in a mouse myeloma cell line. N-terminal sequence analysis of the purified protein revealed the removal of the 23 amino acid signal peptide. In SDS-PAGE, the rhTGF-βsRII resolves into multiple bands due to N-linked glycosylation. Recombinant hTGF-βsRII is a TGF-β antagonist and will inhibit the biological activities of TGF-β1, TGF-β3, and TGF-β5 on TGF-β-responsive cell lines, such as murine HT-2 or human TF-1 with an ED50of approximately 0.3 μg/mL. However, hTGF-βsRII does not inhibit TGF-β2 bioactivities in these cell lines, suggesting that hTGF-βRII has low affinity for TGF-β2. Polyclonal antibodies to hTGF-βsRII have been produced in goats and purified on Protein-G affinity columns. This antibody can inhibit TGF-β1,2,3,5-dependent bioactivities on human cell lines such as TF-1. Additionally, this antibody has species cross-reactivity and will also inhibit TGF-β-dependent bioactivities on murine cells. These results, taken together, suggest that hTGF-βRII is required for signal-transduction of all TGF-β isoforms, including TGF-β2. rhTGF-βsRII has been used as a capture reagent in conjunction with a TGF-β1-specific polyclonal antibody as the detection antibody in a highly sensitive enzyme-linked immunosorbent assay (ELISA) for the specific measurement of TGF-β1 in complex biological fluids with a minimum detectable dose of approximately 10 pg/mL. Latent TGF-β1 and other TGF-β isoforms do not cross-react in this assay. rhTGF-βsRII can also sandwich effectively with anti-TGF-β3 and TGF-β5 antibodies to detect TGF-β3 and TGF-β5, respectively. Consistent with the bioassay results, rhTGF-βsRII is not suitable for use either as a capture or as a detection reagent in TGF-β2-specific ELlSAs because the sensitivity is much lower than for the other isoforms of TGF-β.

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