Hemothorax is a condition where blood and pleural fluid become trapped between the lung(s) and chest wall [1-3], typically from blunt or penetrating trauma . Intrapleural fibrinolytic therapy (IPFT) is a less invasive treatment option that is used to treat retained hemothorax, allowing it to be broken down and drained through tube thoracostomy. IPFT has been successfully modeled in a rabbit system, thus allowing the examination of various treatment options pre-clinically. To help support the use of this animal model, blood halo assay (BHA) was developed to assess the efficacy of various plasminogen activators for use with IPFT. The BHA was optimized in a 96-well format using rabbit as well as human blood with both tPA or uPA and it was found that a blood volume of 10 µL/well was optimal for reproducible halo clot formation. In addition, comparison of halo clots formed with Tissue factor (Innovin) versus thrombin/Ca2+ indicated the latter as providing the most reproducible results. Efficacy of human tPA versus uPA (5 nM) in lysis of human and rabbit halo clots (with or without exogenous human PLG 0-29 nM) was examined by monitoring the increase in the optical density within the well at 510 nm (OD510) over time. Rates of clot lysis induced by tPA were found to be higher than those obtained with uPA for both human and rabbit clots and were independent of the presence of exogenously added PLG. Prepared clot halos in BHA plates were found to be stable at 4ºC from 24 hrs to 6 days as determined by the similar rates of clot lysis obtained. The rate of clot lysis was found to be reduced on day 7, and blood halos were found to be unstable on day 8, possibly due to changes in the fibrin clot structure. The use of an I.DOT system to prepare BHA plates was found to increase productivity, stability, and reproducibility. Overall, the BHA allows rapid in vivo/ex vivo testing of different types of IPFT treatments and shows potential for facilitating personalized treatment of retained hemothorax in a clinical setting.
Date of publication
Pierre Neuenschwander, Galina Florova, Ali Azghani, Alan Cook
Masters of Biotechnology
Huckestein, Charlotte S., "Development of a Companion Assay to Assist in the Study of Fibrinolytic Therapy for Retained Hemothorax" (2022). Biotechnology Theses. Paper 5.