Development of impedimetric immunosensors for biomarkers of cardiac pathology

Immunosensors rely ·on antibody-antigen binding with a range of possible detection methodologies. In this study, electrochemical impedance spectroscopy (EIS) was used to monitor the sensor surface assembly and recognition of three different analytes of cardiac pathology, myoglobin, cardiac troponin I (cTnl) and soluble lectin-like oxidized LDL receptor-1 (sLOX-1). Myoglobin is rapidly released into the circulatory system after an acute myocardial infarction (AMI) and rapidly rising levels make it the first biochemical marker of myocardial damag~, whereas, cTnl is more persistence and specific making a diagnosis more reliable. Circulating levels of sLOX-1 serve as a marker for plaque instability or rupture before an AMI becomes apparent and facilitates early diagnosis of acute coronary syndrome (ACS). The immunosensor fabrication steps comprised of (a) formation of mixed self-assembled monolayers (mSAM) on gold electrodes using a mixture of biotinyl-phospholipid and m.ercaptohexadecanoic acid; (b) Neutravidin functionalisation and (c) attachment of biotinylated --~_._----.. _-- -----._------_.. --- ..- antibodies. The immunosensors were highly spe-afi~--sjnce··a-clinicaIlY--'----' relevant range of analyte concentrations was successfully detected in phosphate buffered saline and in serum concentration ranging from 10% . (v/v) to 100% (v/v) serum without alteration to the slope of the calibration curve. Detection limits for myoglobin and cTnl immunosensor were 10-12 and 10-13 M respectively while sLOX-1 immunosensor achieved 10-13 M. In addition, myoglobin immunosensors were constructed using an arylthiol ester SAM with sulfo-SMCC mediated covalent immobilisation of halfantibody (FAb) fragments, yiel~ing somewhat more sensitive myoglobin detection (10-13 M). Quartz crystal microbalance and atomic force microscopy studies were carried out to study .each step of fabrication to elucidate binding characteristics and surface nanotopography of mSAM based immunosensors. Additionally, radio-labelling studies of myoglobin, sLOX-1 and cTnl were carried out to determine the apparent binding affinity (kd) of the immunosensors.EThOS - Electronic Theses Online ServiceGBUnited Kingdo