Preparation of positive control platelets for detection of canine platelet surface-associated IgG, IgM and complement (C3) by flow cytometry.
An assay for detection of platelet surface-associated (PSA-) IgG, IgM and/or complement (C3) in dogs was modified by preparation of artificial positive control platelets. Flow cytometry of fluorescein isothiocyanate (FITC)-conjugated anti-dog IgG, anti-dog IgM and anti-dog C3 antibodies was used to detect the PSA proteins. IgM single, IgM/C3 double and IgG/IgM/C3 triple positive platelets were prepared. FITC-conjugated anti-IgG antibody bound strongly only to the triple positive platelets. Binding of FITC-conjugated anti-IgM or anti-C3 antibody to the double and triple positive platelets was specifically blocked by preincubation with the respective non-FITC-conjugated same-origin antibodies. These results confirm that FITC-conjugated antibodies specifically detect PSA proteins and that the control platelets prepared in this study are appropriate positive controls for detection of PSA proteins by flow cytometry.
Laboratory of Internal Medicine II, School of Veterinary Medicine, Azabu University, Kanagawa, Japan. tsutiyar@azabu-u.ac.jp
Ryo
R
Tsuchiya
Takahiro
T
Komatsu
Takefumi
T
Ishikawa
Sakurako
S
Neo
Mary F
MF
McConnell
Masaharu
M
Hisasue
Takatsugu
T
Yamada
D000818:Animals; D001792:Blood Platelets / Q000187:drug effects / Q000276:immunology*; D002462:Cell Membrane / Q000276:immunology; D003176:Complement C3 / Q000378:metabolism; D004285:Dogs; D005260:Female; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate; D007074:Immunoglobulin G / Q000097:blood*; D007075:Immunoglobulin M / Q000097:blood*; D008070:Lipopolysaccharides / Q000494:pharmacology; D008297:Male; D010976:Platelet Count / Q000662:veterinary
D003176:Complement C3; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D008070:Lipopolysaccharides; D016650:Fluorescein-5-isothiocyanate