Heart diseases, and mainly heart attacks, are the leading cause of death all over the world. The measurement of cardiac marker proteins in blood of patients presented to the emergency room with chest pain is an important parameter for the accurate diagnosis or proper exclusion of acute myocardial infarction (AMI). The aim of this Thesis is to introduce a novel cardiac marker cardiac troponin I (cTnI) and to develop reliable immunoassays for cTnI measurements....[ Read more ]

Heart diseases, and mainly heart attacks, are the leading cause of death all over the world. The measurement of cardiac marker proteins in blood of patients presented to the emergency room with chest pain is an important parameter for the accurate diagnosis or proper exclusion of acute myocardial infarction (AMI). The aim of this Thesis is to introduce a novel cardiac marker cardiac troponin I (cTnI) and to develop reliable immunoassays for cTnI measurements.

cTnI is one of the most sensitive and specific markers for heart infarction available right now. To develop a sensitive and specific immunoassay for cTnI, monoclonal antibodies (MAbs) have been produced by hybridoma technique. At the current stage, four positive clones TA, TB, TI and TG were identified. The MAb production was generally successful but the amounts of MAbs produced are still not sufficient for subsequent purification.

Since purified MAbs are still not available now, data about the binding kinetic and epitope specificity of the MAbs produced cannot be obtained by Surface Plasmon Resonance (SPR) technology. Therefore, SPR was used to confirm the cTnI ELISA sandwich system. The binding between antigens and antibodies to form the sandwich system can be observed in real time which cannot be achieved in the real cTnI ELISA experiments.

As shown by SPR and cTnI ELISA experiments, the cross-reactivities of the cTnI ELISA to the possible interferents skeletal TnI and TnC are very low, which is good for a specific immunoassay. But the reactivity of the cTnI ELISA to complexed cTnI is also very low which makes it unsuitable to measure the total cTnI in real samples. EDTA was added into the ELISA system as a cleaving agent to cleave Tn complexes but an effect was not obvious. Thus, the cTnI ELISA should be modified for practical measurement of the total cTnI in patient samples.

One of the possible ways to modify the cTnI ELISA is the change of materials used which can enhance the performance to recognize complexed cTnI. To use other commercial antibodies is an option. When the MAbs produced by us are well-characterized and ready for use, then we can employ them in the cTnI ELISA and hopefully construct our own practical cTnI ELISA setup and cTnI rapid tests.