Labeling of platelet surface glycoproteins with biotin derivatives.
We describe a non-radioactive method for the labeling of platelet surface proteins, consisting of platelet protein biotinylation by means of N-hydroxysuccinimido-biotin (NHS-B) and biotin-hydrazide (H-B); NHS-B labels proteins amino residues while H-B binds to periodate-modified sialoglycoproteins. Washed platelets were biotinylated and protein bands were detected after SDS-electrophoresis and western-blot using avidin-peroxidase and luminol as substrate to enhance the signal which was then detected by X-ray film. Biotin-labeled platelet proteins were also immunoprecipitated with monoclonal antibodies against glycoproteins Ib and the IIb-IIIa complex. The use of periodate induced biotinylation is the method of choice for labeling platelet surface glycoproteins while NHS-B also labels internal proteins. The sensitivity of this new procedure is similar to that obtained with radiolabeling techniques; biotinylation does not interfere with the antigenic properties of Ib and IIb-IIIa glycoproteins.
Fabris
F
F
Istituto di Semeiotica Medica, Padua University, Medical School, Italy.
Cordiano
I
I
Mazzuccato
M
M
Pradella
P
P
Randi
M L
ML
Girolami
A
A
D000911:Antibodies, Monoclonal / Q000276:immunology; D016824:Antigens, Human Platelet / Q000276:immunology; D001710:Biotin / Q000031:analogs & derivatives*; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D010504:Periodic Acid; D010980:Platelet Membrane Glycoproteins / Q000276:immunology; D011233:Precipitin Tests; D013388:Succinimides
D000911:Antibodies, Monoclonal; D016824:Antigens, Human Platelet; D010980:Platelet Membrane Glycoproteins; D013388:Succinimides; C033788:biotinyl N-hydroxysuccinimide ester; D010504:Periodic Acid; C053311:biotin hydrazide; D001710:Biotin; C009288:metaperiodate