Chemiluminescent enhanced CD47 detection on Western blotting.
CD47 is a leucocyte antigen of wide tissue distribution including platelets and erythrocytes. It has been found that anti-CD47 antibodies may cause platelet activation. Sensitive techniques are therefore needed for biochemical investigation of platelet glycoproteins including CD47. Conventional Western blotting techniques using horseradish peroxidase-conjugated antibody and 3,3'-diaminobenzidine (DAB) substrate resulted in unsatisfactory results with CD47 due to low sensitivity. When chemiluminescent detection with horseradish peroxidase-conjugated antibody was used in conjunction with Western blotting, detection of the CD47 molecule was greatly enhanced. A protein of 47--55 kDa was immunoblotted from platelets by anti-CD47 while the mAb immunoblotted a broader band (45--60 kDa) from erythrocyte membranes. Thus, there may be different forms of CD47 present on platelets and erythrocytes. The results indicate that Western blotting with the chemiluminescent detection technique is a useful and sensitive alternative for the study of human platelet antigens.
NSW Red Cross Blood Transfusion Service, Sydney, Australia.
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Yuan
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Mihrshahi
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Fletcher
D015703:Antigens, CD / Q000097:blood*; D001792:Blood Platelets / Q000276:immunology*; D015153:Blotting, Western; D051928:CD47 Antigen; D002352:Carrier Proteins / Q000097:blood*; D004910:Erythrocyte Membrane / Q000276:immunology*; D006801:Humans; D008163:Luminescent Measurements; D008970:Molecular Weight; D012680:Sensitivity and Specificity
D015703:Antigens, CD; D051928:CD47 Antigen; C497894:CD47 protein, human; D002352:Carrier Proteins