Research Of Electrochemical Immunosensor For The Detection Of Tumor Biomarker

Cancer is still the main cause of the human death. Although great advances have been made in the treatment of the malignant tumor so far, much importance is attached to its early diagnosis and treatment, which is still considered as the most effective means. In screening the tumor, to detect the tumor-specific marker is the most effective way of making a diagnosis. The advances in the biological technology have made it possible to find many early markers of the tumor, which are characterized by a low concentration in serum. The developed detections such as ELISA, radiation immune, fluorescence immunoassay, etc., have shortcomings of time consumption, radioactive danger, light pollution, and low sensitivity. Recently, chemiluminescence detection, electrochemiluminescence test and protein microarray have further promoted the development of the technology of the clinical testing, but there still exist certain disadvantages to be further overcome. In the current study, a novel sensitive immune analysis was developed for the detection of HCC markers, And the preliminary evaluation of clinical results was given as follows:1. An electrochemical sensor array was constructed based on monoclonal/polyclone antibody sandwich, and applied to the quantitative detection of AFP. Through physical adsorption, one type of and two types of antibodies on the two capture probes were fixed on one chip of the printing electrode, the target protein bonded with the signal probe and with polyclonal antibody, respectively, and the secondary antibody with enzyme markers linked to the complexes. With the portable multi-channel electrochemical device, the electric current was detected for test during the development of the substrate catalyzed via horseradish peroxidase.2. Poly-HRP amplification technology applied to the current signal to detect alpha-fetoprotein (AFP). The polylysine-coated HRP enzyme complexes, labeled an antibody of sheep anti-rabbit, supramolecular IgG, were used as the signal carrier to enhance the signal, thus increasing the sensitivity of the measurement sensor.The second amplification was involved with a double monoclonal antibody sandwich to detect AFP, the target protein and biotinylated monoclonal antibodies bonded with the signal probe, respectively, on the chip of the printing electrode. Streptavidin-HRP was bonded with biotin onto the surface of the printing electrode, and the electric current was further amplified during the development of a new substrate catalyzed via horseradish peroxidase.The results of the first construction showed that AFP detection limit reached56pg/ml, with two linear ranges, a low concentration interval at64p-1000p and a high concentration of interval at1n-250n, as compared with that by the routinely used ELISA kit at ng/ml level, and by the electrochemical luminescence at605pg/ml level.The results of the second construction indicated that AFP detection limit arrived at34pg/ml, producing two linear ranges, one low concentration interval at64p-1000p, and a high concentration of interval atl n-500n. In addition, the immune sensor was proved to be capable of detecting simultaneously16serum samples.The literature has indicated that the early diagnosis and treatment of HCC is still the most effective means, and the advances in biological technology has made it possible to detect the tumor markers early, which are characterized by extremely low serum concentration. Therefore, the testing means of such markers are required to more accurate and sensitive in detecting their serum trace amounts. However, the routinely employed approaches such as ELISA, radiation immune, fluorescence immunoassay and others have been found to be insufficient sensitivity, radioactive and lighting pollution and high time-consumption. Therefore, to develop a higher sensitive immune analysis for the detection of HCC markers is of great significance.In the current study, a novel technology of electrochemical immune sensing was devolved based on the specific union between antigen and antibody by combining a particular immune analysis and an electrochemical sensor. It was found that the AFP detection limit was significantly reduced via two amplifications, thereby expanding the scope of testing, i.e., the AFP detection limit reaching34pg/ml with two linear ranges, a low concentration interval at64p-1000p and a high concentration of interval at1n-500n.A conclusion can be drawn from the results is that the sensor possesses such advantages as high sensitivity, fast speed of immune analysis, simple operation, low consumption of energy and low cost, suggesting the technology can be better applied to the direct serum detection for AFP, as a good prospect in detecting early HCC markers.