Blood coagulation analysis using ElastoSens™ Bio

This is a short report of a study performed by Shiva Naseri, Newsha Koushki, Ehsan Rezabeigi, Allen Ehrlicher and Showan N. Nazhat at McGill University (Montreal, Canada) which title is A nondestructive contactless technique to assess the viscoelasticity of blood clots in real-time (published in 2020 in the Journal of the Mechanical Behavior of Biomedical Materials, 110, 103921).

SUMMARY

The measurement of the absolute blood viscoelastic properties during coagulation is not possible with most conventional techniques.
ElastoSens™ Bio has shown to provide repeatable and sensitive measurements to test blood coagulation through the viscoelastic properties of the forming clot.
The concentration of CaCl2 in blood affects the initialization of coagulation (showing a non-linear behavior). The stiffness of the developed clot is proportional to the concentration of CaCl2.
ElastoSens™ Bio’s measurements were similar to those obtained with a rotational rheometer. The ElastoSens™ Bio’s data showed a better correlation to CaCl2 concentration compared to TEG.

INTRODUCTION

The viscoelastic properties of coagulating blood can be correlated with several diseases and genetic conditions that affect the natural blood coagulation process including bleeding disorders, hemophilia, rare factor deficiencies, von Willebrand disease and platelet function disorders. Therefore, the evaluation of blood clot properties can be valuable for the study, diagnosis and eventually treatment of these diseases. Conventional and more recent techniques used to assess blood coagulation such as thromboelastography (TEG), rotational thromboelastometry (ROTEM) and magnetomotive optical coherence elastography do not directly measure the absolute viscoelasticity of the clot. Furthermore, they normally offer a destructive testing which prevents complementary characterization of the clots and/or the monitoring of the dynamic changes during blood coagulation. Other issues related to these techniques also include low sensitivity and repeatability. In this short application note, ElastoSens™ Bio was used for monitoring the shear storage modulus (G’) of whole blood during coagulation induced by different concentrations of CaCl₂ [1]. In addition, similar tests were performed using TEG and rheometer for comparison.

MATERIALS AND METHODS

Citrated bovine blood (Cedarlane, ON, Canada) was pre-warmed at 37 °C and recalcified with 0.5 mL of CaCl₂ (Sigma-Aldrich) at different concentrations (15, 25, 35, and 45 mM). After 10 gentle vial inversions, the mixture (4 mL of citrated blood + 0.5 mL of CaCl₂) was immediately loaded into the ElastoSens™ Bio sample holders which were previously heated and placed in the chamber of the instrument. The test was performed at 37 °C for 120 minutes. In the case of the rheometer (MCR 302 Anton Paar, Austria), 0.65 mL of recalcified blood was loaded between its parallel plates. Measurements were performed under a constant oscillation frequency of 1.5 Hz and an amplitude of 0.1 % at 37 °C for 120 minutes. For the TEG (Haemoscope Corporation, USA), measurements were conducted according to the manufacturer’s specifications in native mode at 37 °C for 40 min.

RESULTS AND DISCUSSION

Fig. 1 shows the shear storage modulus (G’) evolution of blood samples as a function of time after the addition of CaCl₂ at different concentrations. The increase of the CaCl₂ concentration from 15 mM to 25 mM led to a shorter reaction time (the time at which G’ begins increasing). However, a further increase from 35 mM to 45 mM resulted in longer reaction time. The final G’ values increased with increasing calcium concentration (Fig. 2) showing the formation of stiffer clots. This trend was consistent with the results obtained with the Rheometer and TEG. The relation between the CaCl₂ concentration and blood coagulation obtained with the ElastoSens™ Bio and the rheometer were closer when compared with those from the TEG. This is explained by the fact that the two first techniques directly measure the elastic modulus while TEG measures an indirect parameter (typically rotational displacement) that correlates with viscoelasticity [2,3].

Fig. 1: Shear storage modulus G’ time evolution during coagulation of bovine whole blood after recalcification with 15, 25, 35 and 45 mM CaCl₂ (Error bars: Standard deviation: SD, n = 3).

Fig. 2: The correlation between the shear storage modulus (Pa) and CaCl₂ concentration (mM) measured by ElastoSens™ Bio and Rheometer (left y-axis). TEG results show the correlation of the maximum amplitude with CaCl₂ concentration (right y-axis).

CONCLUSION

The increasing concentration of CaCl₂ from 15 mM to 35 mM reduced the clotting time while a further increase (to 45 mM) led to a longer reaction time. The higher concentration of CaCl₂ resulted in higher clot stiffness. The relation between CaCl2 concentration and the clot final storage moduli measured by the ElastoSens™ Bio was close to those measured by the rheometer. In turn, this information was correlated to the results obtained by TEG (the conventional instrument used in clinics).

PERSPECTIVES

ElastoSens™ Bio can directly measure the evolution of viscoelasticity during clot formation. It provides robust measurements and an easy-to-use platform. The instrument can be used for:

R&D: providing superior quantitative data to better investigate blood coagulation.
Preclinical studies: to study the effect of medication on blood coagulation.

REFERENCES

[1] Naseri, S., Koushki, N., Rezabeigi, E., Ehrlicher, A., & Nazhat, S. N. (2020). A nondestructive contactless technique to assess the viscoelasticity of blood clots in real-time. Journal of the Mechanical Behavior of Biomedical Materials, 110, 103921

[2] Evans, P. A., Hawkins, K., Lawrence, M., Williams, R. L., Barrow, M. S., Thirumalai, N., & Williams, P. R. (2008). Rheometry and associated techniques for blood coagulation studies. Medical engineering & physics, 30(6), 671-679.

[3] Whiting, D., & DiNardo, J. A. (2014). TEG and ROTEM: technology and clinical applications. American journal of hematology, 89(2), 228-232.

How to measure the coagulation kinetics of blood plasma using ElastoSens™ Bio?

The human body has a natural mechanism to stop bleeding after an injury. Platelets migrate to the site of injury and start to form a soft blood clot. This activates other clotting factors in the bloodstream triggering a chain reaction to form a harder blood clot that will stay firmly in place.

Effect of hemostatic agents on the coagulation of blood : in vitro quantitative characterization using ElastoSens™ Bio

The ElastoSens™ Bio was used to analyze the effects of hemostatic agents (HAs) on blood coagulation. The technology identified HAs that can alter clotting independently of the body's natural processes. These findings can optimize current HAs and aid in developing new ones.

How do hemostatic agents work ?

Hemostatic agents (HAs) can be absorbable, biological, or synthetic. Absorbable HAs, like gelatin or oxidized cellulose, speed up clotting and are naturally absorbed by the body. Biological HAs include thrombin, fibrinogen, and platelets which are key to blood clotting. Synthetic HAs, such as polyethylene glycol, form strong sealant matrices. The choice of HA depends on the type of bleeding, tissue interaction, and patient's coagulation profile. Instruments like the ElastoSens™ Bio provide valuable data on HA efficacy by measuring blood absorption and coagulation kinetics.

An inorganic biomaterial with great hemostatic potential

Scientists from Dalhousie University, led by Dr. Mark Joseph Filiaggi, investigated the sodium polyphosphate (NaPP) polymer as a potential hemostatic agent. They tested six formulations of the biomaterial, with varying degrees of polymerization and types of divalent cations. The hemostatic potential of these formulations was evaluated using various blood clotting assays. The biomaterial was mixed with coagulation reagents and recalcified blood or plasma in a tube, which was then shaken to visually assess blood or plasma flow. The clotting time was noted as the time required to achieve no flow. Surgifoam®, a commercial hemostatic agent, was used as a control.

In addition to vaccines, the viscoelasticity of coagulating blood can help optimizing outcomes in COVID-19 patients

In a recent study published in the Critical Care Medicine Journal, researchers from Western Michigan University, the University of Texas, and healthcare-related offices across the United States investigated the relationship between COVID-19 and blood coagulation disorders. They found that analyzing the viscoelastic changes in coagulating blood can provide personalized information on a patient's coagulation state, offering valuable insights for treatment optimization. The study highlights the importance of a personalized patient-oriented approach due to the diverse clinical profiles observed in COVID-19 cases.

Measuring the efficacy of hemostatic agents using ElastoSens™ Bio

Hemostatic agents (HA, e.g. powders, gauzes, adhesives and sealants) have been used for decades to control bleeding. The demand for these agents is growing due to two major trends in surgical practice: the expansion of minimally invasive surgery and complex reconstructive procedures that are more limited in their capacity to obtain hemostasis.