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Characterization of a monoclonal antibody ITI‐Pl 1 directed against human platelet membrane glycoprotein IIb using extracts of whole platelets and platelet surface and intracellular membranes
Author(s) -
Thorsen L. I.,
Gaudernack G.,
Hack N.,
Wilkinson J. M.,
Brosstad F.,
Solum N. O.,
Crawford N.
Publication year - 1988
Publication title -
british journal of haematology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.907
H-Index - 186
eISSN - 1365-2141
pISSN - 0007-1048
DOI - 10.1111/j.1365-2141.1988.tb04181.x
Subject(s) - platelet , thrombasthenia , monoclonal antibody , chemistry , microbiology and biotechnology , platelet membrane glycoprotein , neuraminidase , glycoprotein , epitope , membrane glycoproteins , biochemistry , antibody , biology , immunology , enzyme , platelet aggregation
Summary. A monoclonal antibody (mAb) termed ITI‐Pl 1 has been prepared by the hybridoma procedure. Using immuno‐absorption and crossed immunoelectrophoresis of Triton X‐100 extracts of untreated and EDTA‐treated human platelets it was shown to be directed against the surface membrane glycoprotein IIb (GP IIb). This mAb binds to whole platelets independently of ADP‐stimulation and the presence of Ca 2+ ‐ions. It saturates at around 870 ng/10 8 cells corresponding to approximately 35 800 molecules/platelet. ITI‐Pl 1 did not significantly inhibit GP IIb‐IIIa dependent functions such as platelet aggregation or fibrinogen binding. Immunofluorescence could be demonstrated using ITI‐Pl 1 and intact normal platelets, but not with platelets from a Glanzmann's thrombasthenia patient. Crossed immuno‐electrophoresis with platelet extracts from four different thrombasthenic patients gave a line precipitate in the intermediate gel with 125 I‐labelled ITI‐Pl 1 and autoradiography indicating trace amounts of free GP IIb or the GP IIb‐IIIa complex. The epitope on GP IIb detected by ITI‐Pl 1 is not destroyed by neuraminidase treatment. Thus the mAb also interacts with neuraminidase‐treated GP IIb‐IIIa complex in highly purified platelet surface membrane fractions as well as with GP IIb‐IIIa from untreated internal membranes isolated by continuous flow electrophoresis.