Monitoring modulators of platelet aggregation in a microtiter plate assay.
Platelets play a central role in maintaining biological hemostasis. Inappropriate platelet activation is responsible for thrombotic diseases such as myocardial infarction and stroke. Therefore, novel agents that can inhibit platelet activation are necessary. However, assays that monitor platelet aggregation are generally time-consuming and require high volumes of blood and specialized equipment. Therefore, a medium- to high-throughput assay that can monitor platelet aggregation would be considered useful. Such an assay should be sensitive, comparable to the "gold standard" assay of platelet aggregometry, and able to monitor multiple samples simultaneously but with low assay volumes. We have developed such a microtiter assay. It can assay an average of 60 independent treatments per 60 ml blood donation and demonstrates greater sensitivity than the current gold standard assay, namely platelet aggregation in stirring conditions in a platelet aggregometer. The microtiter plate (MTP) assay can detect known inhibitors of platelet function such as indomethacin, aspirin, and ReoPro. It is highly reproducible when using standard doses of agonists such as thrombin receptor-activating peptide (20 microM) and collagen (0.19 mg/ml). Finally, the MTP assay is rapid and sensitive and can detect unknown platelet-modulating agents from a library of compounds.
Moran
N
N
Department of Clinical Pharmacology, Royal College of Surgeons in Ireland, Dublin 2, Ireland. nmoran@rcsi.ie
Kiernan
A
A
Dunne
E
E
Edwards
R J
RJ
Shields
D C
DC
Kenny
D
D
D000595:Amino Acid Sequence; D001681:Biological Assay; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism; D003094:Collagen / Q000494:pharmacology; D004353:Drug Evaluation, Preclinical / Q000295:instrumentation*; D006801:Humans; D008969:Molecular Sequence Data; D010974:Platelet Aggregation / Q000502:physiology*; D010975:Platelet Aggregation Inhibitors / Q000032:analysis*; D018179:Receptors, Thrombin / Q000737:chemistry; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013917:Thrombin / Q000494:pharmacology
D010975:Platelet Aggregation Inhibitors; D018179:Receptors, Thrombin; D003094:Collagen; D013917:Thrombin