The determination of recombinant human tissue-type plasminogen activator activity by turbidimetry using a microcentrifugal analyzer.
A method is described for determining the activity of recombinant human tissue-type plasminogen activator (rt-PA) by turbidimetry using a microcentrifugal analyzer (MCA). A mixture of thrombin and rt-PA is centrifuged into a mixture of fibrinogen and plasminogen to initiate clot formation and subsequent clot dissolution. The resultant profile of absorbance versus time is analyzed to determine the assay endpoint. Different rt-PA assay concentration ranges were studied in conjunction with profile endpoints for assay optimization. Spiked placebo recovery studies were used to evaluate assay accuracy and precision, which were determined to be 99.5 and 5% (relative standard deviation or RSD), respectively. Assay sensitivity was 0.5 ng/ml. Typical analysis time, including calculations, for a standard curve plus 14 samples was less than 30 min. The application of turbidimetry with the MCA for determining rt-PA activity provides rapid sample analysis and high throughput while maintaining accuracy and precision.
D002498:Centrifugation; D005342:Fibrinolysis; D006801:Humans; D009391:Nephelometry and Turbidimetry; D011994:Recombinant Proteins / Q000032:analysis; D010959:Tissue Plasminogen Activator / Q000032:analysis*
D011994:Recombinant Proteins; D010959:Tissue Plasminogen Activator