Flow cytometric analysis of changes in cytoskeletal proteins during platelet destruction and activation using a monoclonal antibody against platelet myosin.
We developed a new monoclonal antibody directed against platelet myosin (NNKY6-19). Using this antibody, we analyzed platelet cytoskeletal changes related to stimulation with thrombin and to long-term storage. Immunoelectron microscopy showed increased binding of NNKY6-19 to pseudopods and the open canalicular system during treatment with thrombin (0.1 U/ml) and during storage for 7 days. Flow cytometry also showed increased binding to platelets by NNKY6-19 and an antiactin monoclonal antibody during storage. The binding of NNKY6-19 showed an increase greater than that with the antiactin antibody after storage of platelets for 7 days and after thrombin treatment. These findings indicated that the increased binding of NNKY6-19 had some relationship to changes in intracellular myosin and platelet morphology. Thus use of NNKY6-19 allowed analysis of subtle changes related to platelet activation, which differed from those detected by antibodies against platelet glycoproteins or by the antiactin antibody. This antibody appears to provide a simple method for studying changes in platelet cytoskeletal and surface proteins.
Yamaguchi
K
K
First Department of Internal Medicine, Kansai Medical University, Osaka, Japan.
Nomura
S
S
Kido
H
H
Kawakatsu
T
T
Fukuroi
T
T
Suzuki
M
M
Hamamoto
K
K
Yanabu
M
M
Kokawa
T
T
Yasunaga
K
K
D000199:Actins / Q000032:analysis / Q000276:immunology; D000906:Antibodies / Q000276:immunology*; D001792:Blood Platelets / Q000737:chemistry* / Q000166:cytology / Q000502:physiology; D003598:Cytoskeletal Proteins / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel; D005434:Flow Cytometry; D006801:Humans; D015151:Immunoblotting; D016253:Microscopy, Immunoelectron; D009218:Myosins / Q000032:analysis* / Q000276:immunology*; D015539:Platelet Activation / Q000502:physiology*
D000199:Actins; D000906:Antibodies; D003598:Cytoskeletal Proteins; D009218:Myosins