GPIIIa-related PLA1 antigens with different molecular weights: studies in platelets, endothelial cells, and megakaryocytes.
The mol wt of the glycoprotein(s) carrying the PLA1 antigen was examined on platelets, megakaryocytes and endothelial cells by immunoblotting with a human polyclonal anti-PLA1 antibody (BE), as well as on four different monoclonal antibodies (MoAbs; DEK-1, DEK-2C, DEK-10, and DEK-16) raised against GPIIIa, the 100,000-mol wt platelet glycoprotein known to carry the PLA1 antigen. BE reacted with PLA1 positive but not with PLA1 negative platelets. DEK-1 reacted strongly with PLA1 positive platelets but weakly with PLA1 negative platelets. The remaining three MoAbs reacted equally with PLA1 positive as well as negative platelets. BE, DEK-1, DEK-10, and DEK-16 reacted with a 120,000- as well as 100,000-mol wt band on immunoblot of PLA1 positive platelets. The 120,000-mol wt band copurified with affinity purified 100,000-mol wt GPIIIa. Megakaryocytes had a prominent 120,000- as well as 105,000-mol wt band that reacted with BE on immunoblot (the 100,000-mol wt band was not detectable). Umbilical cord endothelial cells from presumed PLA-positive infants had a prominent 100,000-mol wt band that reacted with BE, DEK-16, and DEK-1 (the 120,000-mol wt band was not visualized). The 120,000- and 100,000-mol wt PLA1-positive bands could be digested with proteolytic enzymes to 55,000- to 65,000-mol wt-resistant fragments that retain PLA1 epitopes. Further digestion with endoglycosidase-H lowered the apparent mol wt by approximately 2,000 to 6,000 daltons without affecting PLA1 reactivity. We conclude that the PLA1 antigen is present on a 120,000- as well as 100,000-mol wt glycoprotein of platelets and megakaryocytes, a 105,000-mol wt band of megakaryocytes, and a 100,000-mol wt glycoprotein of endothelial cells. We postulate that the 120,000-mol wt glycoprotein, which shares three or more epitopes with the 100,000-mol wt GPIIIa, may be a post-translational precursor of this species.
Dancis
A
A
Department of Medicine, New York University Medical School 10016.
Ehmann
C
C
Ferziger
R
R
Grima
K
K
Karpatkin
S
S
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000937:Antigen-Antibody Reactions; D016824:Antigens, Human Platelet; D001792:Blood Platelets / Q000276:immunology*; D002918:Chymotrypsin; D004591:Electrophoresis, Polyacrylamide Gel; D004730:Endothelium, Vascular / Q000276:immunology*; D006801:Humans; D007118:Immunoassay; D039661:Integrin beta3; D007519:Isoantigens / Q000276:immunology / Q000302:isolation & purification*; D008533:Megakaryocytes / Q000276:immunology*; D051379:Mice; D008970:Molecular Weight; D010980:Platelet Membrane Glycoproteins / Q000276:immunology / Q000302:isolation & purification*
D000911:Antibodies, Monoclonal; D016824:Antigens, Human Platelet; C061248:ITGB3 protein, human; D039661:Integrin beta3; D007519:Isoantigens; D010980:Platelet Membrane Glycoproteins; D002918:Chymotrypsin