Detection Of Cardiac Disease Markers Based On Soybean Peroxidase Immunosensing

Cardiovascular disease is the“killer”threat to human health,and the acute myocardial infarction?AMI?is a major cardiovascular disease.The onset of AMI is Sudden,and the acute mortality rate of about30%.In the common diagnostic methods of AMI,the detection of its biochemical markers is extremely important.Cardiac troponin I?cTnI?has high myocardial specificity and is therefore considered the"gold standard"for the diagnosis of AMI.To achieve sensitive and accurate detection of cTnI for the early diagnosis and treatment of AMI disease has great practical significance.At the same time,creatine kinase?CK-MB?is an enzyme with a higher myocardial specificity in myocardial enzyme spectrum.Myoglobin?Myo?has a small molecular weight,and its concentration increases significantly in the early stage of AMI,therefore,it has a higher sensitivity for the diagnosis of AMI.The detection of cTnI,CK-MB and Myo in serum is integrated into the same chip,and the simultaneous detection of three biomarkers of myocardial diseases can provide the basis and guarantee for the rapid,sensitive and accurate diagnosis and treatment of AMI.Based on this,this paper constructed immunosensors based on soybean peroxidase?SBP?catalyzed luminol-hydrogen peroxide?H₂O₂?system for the detection of biochemical markers of AMI in serum,including the following two parts:?1?Polyethylenimine-functionalized graphene?PEI-RGO?was prepared based on the good electrical conductivity,biofilm formation and reducibility of polyethyleneimine?PEI?,and the capture antibody?Ab₁?was immobilized onto its surface by amide reaction.Afterwards,the cTnI antigen?Ag?and the soybean peroxidase-labeled detection antibody?SBP-Ab₂?were incubated continuously to form a sandwich immune complex by relying on the specific recognition between antigen and antibody.In the presence of hydrogen peroxide?H₂O₂?,the sensitive detection of cTnI was achieved using the electrochemical luminescent signal generated by SBP catalyzed oxidation of luminol.The linear range for cTnI detection was 5-30000 pg/mL,and the detection limit was 3.3 pg/mL.The proposed immunosensor had good selectivity and reproducibility for the detection of cTnI.At the same time,it was used for the detection of real plasma samples and showed acceptable accuracy,which provided a choice for the detection of cTnI.?2?A biological immune chip was constructed to achieve the simultaneous detection of three biochemical markers of cTnI,CK-MB and Myo.The three corresponding capture antibodies were immobilized onto the surface of the nitrocellulose membrane,followed by immobilization of the antigens and the corresponding detection antibodies Ab₂-SiO₂-SBP to form a double antibody sandwich immunological structure.In the presence of the enhancer,SBP catalyzed luminol-H₂O₂ to generate a strong chemiluminescent signal,and the luminescent signal value at the corresponding position was acquired to perform simultaneous detection of the three biomarkers.The results showed that the linear detection range for cTnI was 0.02-80 ng/mL?detection limit of 0.006 ng/mL?,the linear detection range for CK-MB was0.2-120 ng/mL?detection limit of 0.067 ng/mL?,and the linear detection range for Myo was 0.3-480 ng/mL?detection limit of 0.1 ng/mL?.