Quantification of transforming growth factor-beta in biological material using cells transfected with a plasminogen activator inhibitor-1 promoter-luciferase construct

Anal Biochem. 1997 Apr 5;247(1):45-51. doi: 10.1006/abio.1997.2026.


Transforming growth factor-beta (TGF-beta), a multifunctional cytokine, can be quantified by a variety of bioassays or immunoassays. One of the disadvantages of these techniques is that they require sample purification to remove components that interfere with the TGF-beta signal. In the current study the feasibility of quantifying TGF-beta in complex biological fluids directly with a recently developed bioassay was examined. This assay is based on the ability of TGF-beta to induce plasminogen activator inhibitor-1 (PAI-1) expression. Mature TGF-beta binds to the receptors of mink lung epithelial cells transfected with a plasminogen activator inhibitor-1 promoter-luciferase construct (PAI/L), resulting in a dose-dependent increase of luciferase activity. Specificity for TGF-beta was proven by treatment of the samples with neutralizing antibodies. The sensitivity and the intraassay precision are comparable to the ELISA. It is demonstrated, however, that, unlike the ELISA, a purification step by, e.g., acid-ethanol extraction prior to the PAI/L assay, is not required. This not only simplifies the assay but also reduces the minimal sample volume and allows to discriminate between latent and mature TGF-beta. The present study furthermore provides insight in the critical steps for accurate TGF-beta determination, which include careful blood collection and sample handling (storage and preparation). With this protocol TGF-beta has been quantified in human plasma, rat plasma, rat saliva, tissue extracts from rat lung, and in culture medium of TGF-beta-producing cells.

Publication types

  • Comparative Study

MeSH terms

  • Animals
  • Biological Assay / methods*
  • Biological Assay / standards
  • Biological Assay / statistics & numerical data
  • Blood Chemical Analysis / methods
  • Blood Chemical Analysis / standards
  • Blood Chemical Analysis / statistics & numerical data
  • Cells, Cultured
  • Enzyme-Linked Immunosorbent Assay / statistics & numerical data
  • Evaluation Studies as Topic
  • Female
  • Gene Expression
  • Humans
  • Luciferases / biosynthesis
  • Luciferases / genetics
  • Lung / chemistry
  • Male
  • Mink
  • Plasminogen Activator Inhibitor 1 / genetics*
  • Promoter Regions, Genetic
  • Rats
  • Reference Standards
  • Reproducibility of Results
  • Saliva / chemistry
  • Sensitivity and Specificity
  • Transfection*
  • Transforming Growth Factor beta / analysis*
  • Transforming Growth Factor beta / isolation & purification
  • Transforming Growth Factor beta / metabolism


  • Plasminogen Activator Inhibitor 1
  • Transforming Growth Factor beta
  • Luciferases