Distance-Based Heparin Sensing using Optodes embedded in Agarose Gel
Heparin is a polyanionic blood anticoagulant used in several surgical procedures. Its effects are reversed with protamine, an arginine-rich peptide. Heparin concentration changes with time in the body. Thus, it is crucial to monitor it quickly to keep it constant during the procedure. Too low of a heparin concentration increases the clotting risk, whereas too high can induce uncontrolled bleeding . Knowing precisely the heparin concentration is also required to determine the appropriate amount of protamine to neutralize the anticoagulant effect at the end of surgery.
Currently, the anti-Xa assay is used to measure heparin . It binds to antithrombin, which inhibits factor Xa. A reporter is then added to quantify the unbound fraction of factor Xa optically. This method requires specialised equipment, trained staff, significant amount of time and sample treatment, complicating quantification during surgical procedures.
Within our group, we have developed emulsion-based microparticles sensitive towards protamine based on previous research . They contain a cationic solvatochromic dye (X4), a hydrophobic anionic exchanger (NaTFPB) and a polar anionic pseudo-ionophore towards protamine (DNNS). When the latter binds to protamine, polarity of the dye environment decreases resulting in an absorption shift, which can be related to protamine concentration and later to heparin level. Patient plasma have been successfully measured using this sensor. However, our current approach still requires a calibration curve and a large sample volume. It is also unsuitable for whole blood measurements.
Moving to a distance-based approach with agarose gel has the potential to remove the need of calibration for each measurement, reduce the required sample volume and make the sensor suitable for whole blood analysis. Agarose gels are known to be able to filter red blood cells , which are the main interfering species when analysing blood samples by colorimetry. Our device is currently composed of a polystyrene semi-micro cuvette filled with an agarose gel containing protamine sensitive particles. Dropping the sample on top allows the quantification of protamine concentration using image analysis to determine the distance in which a color change is induced. In the talk, we will the present the most recent advances in the development of our distance-based device and data analysis procedure.
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