The use of PhastSystem crossed immunoelectrophoresis with immunoblotting to demonstrate a complex between glycoprotein Ib and the actin-binding protein (ABP) of human platelets.
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.
Aakhus
A M
AM
Research Institute for Internal Medicine, University of Oslo, Rikshospitalet, Norway.
Wilkinson
M
M
Pedersen
T M
TM
Solum
N O
NO
D000911:Antibodies, Monoclonal; D001792:Blood Platelets / Q000032:analysis*; D006801:Humans; D015151:Immunoblotting / Q000379:methods*; D007122:Immunoelectrophoresis / Q000379:methods*; D007123:Immunoelectrophoresis, Two-Dimensional / Q000379:methods*; D008840:Microfilament Proteins / Q000378:metabolism*; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism*; D011233:Precipitin Tests
D000911:Antibodies, Monoclonal; D008840:Microfilament Proteins; D010980:Platelet Membrane Glycoproteins