Soluble T cell receptors: detection and quantitative assay in fluid phase via ELISA or immuno-PCR

J Immunol Methods. 1995 Oct 26;186(2):181-94. doi: 10.1016/0022-1759(95)00129-x.

Abstract

To establish the concentration range in which soluble murine T cell receptors (sTCR), derived from the Th2 clone D10, exhibited biological activity, and to follow production and purification of D10 sTCR, we devised four quantitative immunoassays: three ELISA systems, and an immuno-PCR assay. The direct ELISA, employed hamster anti-TCR beta monoclonal antibody (H57), which detects all types of alpha beta TCR, regardless of their variable regions, and had a detection limit of about 6 ng/ml sTCR. The indirect sandwich ELISA employed anti-V beta 8 as capture antibody, and had a detection limit of 600 pg/ml. With the direct sandwich ELISA, that also employed anti-V beta 8, TCR concentrations as low as 100 pg/ml could be detected. The ELISA assays were specific for soluble alpha beta TCR, and showed no cross-reactivity when employing two control hamster anti-gamma delta TCR mAbs (GL3 and UC7), or with anti-TCR beta and monoclonal hamster IgG as a control antigen. Further, we demonstrated that in some assays where use of passive binding ELISA plates resulted in a high background, replacement with covalent binding ELISA plates resulted in an acceptable low background value. With the immuno-PCR assay, concentrations of sTCR as little as 0.8 pg/ml could be detected. In summary, the assays described here may prove valuable in investigating the occurrence and amount of sTCR in vitro and in vivo.

Publication types

  • Comparative Study
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Antibodies, Monoclonal / immunology
  • Antibodies, Monoclonal / isolation & purification
  • Base Sequence
  • Chromatography, Affinity
  • Chromatography, High Pressure Liquid
  • Chromatography, Ion Exchange
  • Cricetinae
  • Enzyme-Linked Immunosorbent Assay / instrumentation
  • Enzyme-Linked Immunosorbent Assay / methods*
  • Humans
  • Mice
  • Molecular Sequence Data
  • Osmolar Concentration
  • Polymerase Chain Reaction / methods*
  • Receptors, Antigen, T-Cell, alpha-beta / analysis*
  • Receptors, Antigen, T-Cell, alpha-beta / isolation & purification
  • Receptors, Antigen, T-Cell, gamma-delta / analysis
  • Recombinant Proteins / immunology
  • Sensitivity and Specificity
  • Solubility
  • Th2 Cells / chemistry*

Substances

  • Antibodies, Monoclonal
  • Receptors, Antigen, T-Cell, alpha-beta
  • Receptors, Antigen, T-Cell, gamma-delta
  • Recombinant Proteins