New-Generation Coagulation Laboratory – A Glimpse
In the hundred years since the first clotting time tests were proposed, the development of coagulation assay technology has progressed tremendously. The range and the quality of the tests available have also increased significantly, allowing various measures of clotting time, platelet function, factor activity, and concentration. These tests have progressed alongside an increased understanding of the processes involved in hemostasis and the development of anticoagulant drug therapies.
Automated technologies have revolutionized the monitoring of coagulation disorders in the central hospital laboratory setting, allowing for high-throughput testing, improved accuracy, and precision, accompanied by a marked reduction in human error. Failure pattern sensors, which detect insufficient sample levels and clots and cap piercing, which enhances safety as well as sample integrity. have been introduced. However, they still require trained operators and sample transportation. Owing to the relatively long turnaround time of conventional laboratory tests, and with the aim of reducing inappropriate transfusion, there is increasing interest in point-of-care coagulation testing (POCCT), which can provide results within minutes.
Several studies have highlighted the need for POCCT in clinical settings including liver transplantation, cardiothoracic surgery, and trauma because of the high risk of hemorrhage and the resulting requirements for blood-component support. Others have identified that POCCT in the form of platelet count, PT, aPTT, and fibrinogen concentration has resulted in reduced blood loss, shorter surgical times, and reduced transfusion requirements. While the initial cost of POCCT may be greater, the wider patient, economic, and societal benefits offered look promising. In a recent review the effects of POCCT on the rate of perioperative transfusion of allogenic blood products, the frequency of hemostatic treatment, and the clinical outcome were assessed. The wider usage of POCCT devices may potentially reduce rates of transfusion and lower costs associated with hemotherapy.
Technological advances
The central event in coagulation pathways is the production of thrombin, which acts upon fibrinogen to produce fibrin and thus the fibrin clot. For several decades, coagulation analyzers have detected clot formation in one of two ways – optical or mechanical. Most automated analyzers currently used in hemostasis laboratories are capable of performing other types of testing as well, such as chromogenic, latex agglutination, and even enzymatic immunoassays.
Several of the novel devices offer possible alternative means to study platelet aggregation – a laboratory test procedure that is currently complex and time-consuming to perform. The drive for POC has also driven technological solutions, which can operate at the microscale, can be run at lower costs, and use less blood volume. Some of the technological advances are summarized below.
Thromboelastography (TEG) and Thromboelastometry (ROTEM) provide global information on the dynamics of clot development, stabilization and dissolution that reflect in vivo hemostasis. Although TE has not been subjected to the same evaluation processes as conventional hemostatic tests, its use as a POCCT monitor in complex major surgery has been shown to significantly reduce the use of blood-component therapy and overall blood loss.
T2 magnetic resonance (T2MR) is one new and emerging technology that is able to detect clot formation, as used in a novel device. T2MR is able to detect clot formation based on partitioning of red blood cells and proteins, which occurs during fibrin formation and platelet-mediated clot contraction. This device can be used to measure clotting times, individual coagulation factors, and platelet function, and has also revealed a novel hypercoagulable signature that needs further study to determine if it can be used to predict patients at higher risk of thrombosis.
Acoustic waves are alternatively used in another novel, miniaturized point-of-care device capable of using only a small amount of citrated whole blood, measuring the time required for fluorescent microspheres to cease motion due to clot formation. Overall, 1 mL of whole blood is initially needed to be collected, but only <10 µL is loaded onto the device. The result provided is a clotting time in seconds. This system may be useful for assessing anticoagulant effects, and the device has been studied in patients receiving different types of anticoagulants such as heparin, argatroban, rivaroxaban, dabigatran, and warfarin.
Infrared spectroscopy is instead used to detect clot formation in the Perosphere Technologies’ hand-held point-of-care coagulometers. This device uses fresh or citrated whole blood (<10 µL) with clotting activation initiated by glass contact.
The turnaround time is fast, providing a clotting time within 3 to 10 minutes. Preliminary data showed that this device may be useful in assessing coagulation response to any of the direct oral anticoagulants (DOACs) as well as the antithrombin-dependent activated factor X (FXa), inhibiting anticoagulants such as heparin, low molecular weight heparin, and fondaparinux.
Laser speckle rheology (LSR) is employed to detect clot formation using another novel optical hand-held point-of-care device that performs several coagulation tests including prothrombin time, activated clotting time, clot polymerization rate (α-angle), clot stiffness, fibrinolysis, and platelet function. LSR quantifies tissue viscoelasticity from light scattering patterns called laser speckle, using small amounts (40 µL) of whole blood and providing test results within 10 minutes.
Another instrument uses automated digital microfluidics in coagulation testing, enabling the performance of up to 12 different tests at once on <50 µL whole blood, using small electric fields to manipulate droplets of fluid on a printed circuit board. This system has been used to perform genetic testing for FV Leiden and prothrombin G20210A, functional assays for antithrombin, protein C, and FVIII, as well as antigen assays for antithrombin, protein C, protein S, anticardiolipin antibody (immunoglobulinG [IgG], IgM), FVIII,VWF, and homocysteine, as well as for anticoagulant detection by anti-FXa.
Novel analyzers have also been developed, which more specifically assess the role of platelets in human pathologies, including bleeding and thrombotic disorders, cancer, sickle-cell disease, stroke, ischemic heart disease, and others. In development stages, the platelet contraction cytometer is a small device that can assess platelet contractile forces, and preliminary studies suggest that it may identify some patients with a bleeding tendency that cannot otherwise be identified by existing laboratory tests. Microfluidic flow devices are potentially useful in diagnosing primary hemostasis disorders. These devices are also helpful in assessing clinical bleeding risk in the setting of trauma and surgery, and antiplatelet therapy in the setting of platelet thrombus prevention in acquired and inherited hypercoagulable states.
The technologies inherent in these devices offer a combination of physiology accuracy and small blood volume requirements in the evaluation of mild VWD and platelet function in flowing whole blood, with the potential to individualize therapeutic options for, and to achieve greater diagnostic accuracy in platelet disorders and VWD.
Way forward
The coagulation and hemostasis testing has come a long way from PT, APTT to a whole gamut of complex tests, which are of paramount importance in management of coagulopathies (bleeders) and thrombosis. Besides an accurate diagnosis, timely diagnosis is of equal importance since many of these can be rapidly fatal. Advances in conventional instruments with their ever-increasing menu of specialized tests make complex tests like factor assays, platelet function tests, vWD, and the like, available to a far greater proportion of clinicians and patients. At the same time, improving technologies in point-of-care testing devices enables patients to do home monitoring and anesthesiologists and surgeons to manage intraoperative complications better.
POCCT results may not necessarily mirror those values from laboratory-based testing. The reliability of these tests depends on the experience of the operator and appropriate calibration. A thorough familiarity of the devices’ functioning, methodology, and strengths and weakness is imperative. POC INR testing is widely used in outpatient labs and anticoagulation clinics, although clinical lab INR testing remains the reference standard. Integration of the clinical laboratories and POCCT testing so that the two complement each other and clinicians and patients get the best of both worlds is of paramount importance.
