Optical Thromboelastography to evaluate whole blood coagulation

Measurement of blood viscoelasticity during clotting provides a direct metric of haemostatic conditions. Therefore, technologies that quantify blood viscoelasticity at the point-of-care are invaluable for diagnosing coagulopathies. We present a new approach, Optical Thromboelastography (OTEG) that measures the viscoelastic properties of coagulating blood by evaluating temporal laser speckle fluctuations, reflected from a few blood drops. During coagulation, platelet-fibrin clot formation restricts the mean square displacements (MSD) of scatterers and decelerates speckle fluctuations. Cross-correlation analysis of speckle frames provides the speckle intensity temporal autocorrelation, g2(t), from which MSD is deduced and the viscoelastic modulus of blood is estimated. Our results demonstrate a close correspondence between blood viscoelasticity evaluated by OTEG and mechanical rheometry. Spatio-temporal speckle analyses yield 2-dimensional maps of clot viscoelasticity, enabling the identification of micro-clot formation at distinct rates in normal and coagulopathic specimens. These findings confirm the unique capability of OTEG for the rapid evaluation of patients’ coagulation status and highlight the potential for point-of-care use. Spatial maps of blood viscoelasticity index, G, during clotting obtained from a normal patient (top row) and a hypo-coagulable patient with low levels of clotting factors (bottom row) at 0, 1, 14, and 30 minutes after kaolin activation. Micro-clots of significant G values appear at early times (~1 min) and continue to progress to form a large blood clot over 30 min in the normal patient. In contrast, in the hypo-coagulable sample, micro-clots of comparable G are only visible at 14 min and the extent and overall clot strength is considerably reduced compared to the normal patient even at 30 min. Scale bars are 100 μm long. These results demonstrate the high sensitivity and spatial resolution of OTEG for detecting incipient micro-clots during very early stages of clot formation in patients. Keywords: Blood, coagulation, laser speckle, optical thromboelastography, rheology, thromboelastography, viscoelasticity 1. Introduction Abnormalities in the blood coagulation process, termed coagulopathies, can result from several conditions following acute trauma, surgery, or illness and are often the underlying cause of life-threatening bleeding or thromboembolic conditions. For instance, deficient coagulation or ‘hypo-coagulable’ states can result in dangerous blood loss and organ failure [1]. In other cases, increased clotting tendency or ‘hyper-coagulable’ states can cause potentially fatal arterial or venous thromboemboli resulting in deep vein thrombosis, pulmonary embolism, myocardial infarction or stroke [2, 3]. Monitoring blood coagulation status rapidly at the bedside therefore is of crucial importance for preventing severe bleeding or detecting and treating thrombotic conditions. Unfortunately, due to their long reporting times (1–5 hours), conventional coagulation tests (CCT’s) have limited clinical utility particularly in the context of dynamically changing coagulation conditions in severely injured, surgical, or critically ill patients [4]. This dire situation calls for the development of new technologies for detecting blood coagulation abnormalities at the patient’s bedside to facilitate the early identification and management of coagulation impairments. A direct indicator of blood coagulation status can be obtained by measuring the temporal evolution of whole blood viscoelasticity, closely associated with the time course of several biochemical processes of the coagulation cascade [5]. For instance, rheology-based coagulation technologies, such as Thromboelastography (TEG®) and Rotational Thromboelastometry (ROTEM®), report on blood coagulation status by quantifying the viscoelasticity of clotting blood in real-time by mechanically stirring a whole blood sample with a rod in a cup [4, 5]. While favourable over CCT’s in terms of reporting time and real-time operation, both TEG® and ROTEM® are contact-based methods that are likely to strain the blood sample beyond the linear viscoelastic regime, delay clot formation, and modify fibrin network structure, thereby providing unreliable results [5]. Moreover, these instruments have bulky moving parts, are complex to operate, and require routine calibration, which hinders their widespread clinical acceptance for point-of-care use at the patients’ bedside [6]. In this paper, we introduce Optical Thromboelastography (OTEG), a novel optical approach that will likely overcome the limitations of TEG® and ROTEM® to evaluate blood viscoelasticity in a rapid, non-contact manner using just a few drops of whole blood. In OTEG, the specimen consisting of a few drops of blood is illuminated by a laser source, and temporally fluctuating speckle patterns are captured by a high-speed CMOS sensor. We have previously demonstrated that temporal intensity fluctuations of laser speckle patterns provide information on the viscoelastic properties of tissues and test phantoms [7–12]. In un-clotted blood, the continuous re-arrangements of intrinsic light scattering particles undergoing Brownian motion induce rapid temporal speckle intensity fluctuations. During coagulation, blood viscoelasticity is progressively modified in concert with key aspects of haemostatic clot formation, including platelet aggregation, fibrin polymerization, and fibrin cross-linking [13]. The gradual emergence of a fibrin-platelet clot with increased stiffness restricts the Brownian motion of scattering particles, subsequently culminating in reduced optical phase shifts and slower speckle fluctuations. In this paper, we demonstrate that OTEG measurements leverage from the susceptibility of laser speckle intensities to optical phases shifts caused by sub-wavelength scale particle displacements that in turn closely follow the alterations in the viscoelastic properties of coagulating blood. Thus, the viscoelastic properties of blood during coagulation can be measured accurately using OTEG without requiring any sample manipulation unlike TEG® or ROTEM®. Here, we elaborate on OTEG methods to measure blood viscoelasticity and discuss results obtained using whole blood samples from animals and patients with normal and defective coagulation states.