The use of PhastSystem crossed immunoelectrophoresis with immunoblotting to demonstrate a complex between glycoprotein Ib and the actin-binding protein (ABP) of human platelets

Electrophoresis. 1989 Nov;10(11):758-61. doi: 10.1002/elps.1150101105.

Abstract

The study shows how a technique described in an accompanying paper can be applied to solve a biological problem. The technique makes use of the observation that a monoclonal antibody that has been coprecipitated with its antigen during crossed immunoelectrophoresis can be transferred to a nitrocellulose membrane and visualized. Previous studies using crossed immunoelectrophoresis of Triton X-100 extracts of platelets have indicated that a particular immunoprecipitate (peak III) of the membrane receptor glycoprotein Ib (GP Ib) might contain a complex between the receptor and the actin-binding protein (filamin). When a monoclonal antibody (PM6/317) directed towards the actin-binding protein was added to a platelet extract prior to immunoelectrophoresis and blotting, this was visualized on the blot as a replica of the peak III immunoprecipitate. This demonstrates a colocalization of GP Ib and the actin-binding protein in the precipitate, and thus the existence of a complex between the membrane receptor and the cytoskeletal protein.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antibodies, Monoclonal
  • Blood Platelets / analysis*
  • Humans
  • Immunoblotting / methods*
  • Immunoelectrophoresis / methods*
  • Immunoelectrophoresis, Two-Dimensional / methods*
  • Microfilament Proteins / metabolism*
  • Platelet Membrane Glycoproteins / metabolism*
  • Precipitin Tests

Substances

  • Antibodies, Monoclonal
  • Microfilament Proteins
  • Platelet Membrane Glycoproteins