Helmut Lill

Beta-adrenergic receptor mediated release of tissue plasminogen activator in anaesthetized dogs

Drugs like epinephrine or isoproterenol can enhance fibrinolytic activity of blood. It is still a matter of debate whether this effect can be totally or only partially blocked by beta-receptor blockers. We have developed a dog model for ex vivo measurement of the fibrinolytic activity using a spectrophotometric assay employing human plasminogen, chromogenic plasmin substrate and human fibrinogen-BrCN digests for the stimulation of t-PA. After i.v. administration of isoproterenol the fibrinolytic activity increased, but this could be seen only in the presence of fibrinogen-BrCN digest in the test system. This suggests that isoproterenol caused higher levels of dog t-PA. Pretreatment with the beta-blocker propranolol completely blocked the increase in t-PA.

Development of a photometric assay for activated partial thromboplastin time and its application to the cobas(R) biocentrifugal analyzer

We describe a two-step procedure for APTT that can be performed on photometric devices. It includes preincubation of diluted plasma with ellagic acid and phospholipids and a starting reagent that contains calcium and a chromogenic peptide substrate for thrombin, Tos-Gly-Pro-Arg-pNA. Reaction time is recorded from addition of the starting reagent until thrombin formation occurs, and a prefixed amount of substrate is cleaved. The pattern of sensitivity to clotting factors and heparin was similar to clotting assays and the substrate used did not interfere with the activity of factor Xa. An application of the method was made for the Cobas(R) Bio centrifugal analyzer. Absorbance readings were sent to an external computer and were transformed into reaction times by a computer program. Although the results are independent on fibrinogen concentrations, from kinetic data of the reaction curve fibrinogen concentrations can be estimated. Correlation studies showed good correspondence to clotting methods (r = 0.92, n = 53) as well as an excellent precision (CV 3% for inter-assays, n = 15) and high throughput of samples (greater than 100/h) in the automated assay.