Pororus Layer Open Tubular Capillary Based Immunosensor For Senitive Detection Of Biomarkers

Conventional enzyme-linked immunosorbent assay(ELISA)for biomarker detection in clinical diagnosis faces some challenges because of their relatively low sensitivity,time-consuming process,and multiplex detection incapability.In recent years,capillary-based immunosensors that utilize capillary tubes as supports for bio-reagent immobilization offer several advantages,including fast and sensitive assay,convenient operation,and low reagent and sample consumption,which being used in the filed of fermentation industry,biological medicine and biochemical research in recent years,and how to further improve the detection sensitivity of capillary immune sensor has been a hot research topic in this field.The Porous layer open tubular(PLOT)has a three-dimensional uniform Porous structure,and its large specific surface area can increase the load of the target molecule,while reducing the mass transfer distance and shortening the analysis time.The present paper mainly focused on the research of surface modification-mediated porous layer open tubular(PLOT)capillary immunosensor.The contents were listed below:1.We developed a sensitive,rapid,and multiplex PLOT capillary-mediated fluorescence immunosensor for the simultaneous identification of alpha fetoprotein(AFP)and carcinoembryonic antigen(CEA)in clinical serum samples.Via an in situ biphasic reaction,we successfully prepared an innovative surface modification-mediated porous layer open tubular(PLOT)capillary,which has three-dimensional uniform porous structure.We immobilized two capture antibodies on the inner surface of PLOT capillary via glutaraldehyde chemical covalent bonding,and then connected two separate PLOT capillaries by utilizing a suitable sleeve tube,and perform multiplex targets simultaneously detection in the same sample by combining double antibody sandwich ELISA and laser induced fluorescence(LIF)detection.This capillary comprised three-dimensional homogeneous and porous structures,which could increase the surface-area-to-volume ratio for antibody immobilization.Thus,this capillary was shown as an ideal immunoreaction base to enhance the sensitivity of immunoassays and shorten analysis time.The detection sensitivity of this immunosensor improved by 50-70 times compared with that of conventional ELISA,and the whole analysis could be completed within 1 h.Furthermore,we investigated the intra and inter-assay coefficients of variation of the immunosensoe were less than 6.0%,which results proved the acceptable reproducibility of the proposed immunoassay.The recovery of AFP and CEA were 84.6%-113.4% and 85.7%-110.8% in the spiked sample,respectively.The developed immunosensor were used to detected the human serum sample,which results werecompared with the clinic test results.Good correlation was found between the two methods(R2=0.9).This work suggest that suitable surface modification strategy is an effective tool to improve the analytical performance of conventional immunoassay and our study provided a feasible,sensitive,and multi-target assay for the detection of cancer biomarkers,which would be of valuable application in clinical diagnosis.2.We developed a novel,sensitive porous layer open tubular(PLOT)capillary-based chemiluminescence immunosensor for the detection of the procalcitonin.In this work,we immobilized the caputure antibody,target analyte,HRP enzyme-labeled detection antibody on the PLOT capillary inner surface via chemical covalent bonding by sandwich ELISA,and immediately measured the chemiluminescence intensity for the quantitative determined of analytes through the HRP catalyze the substrate luminol and hydrogen peroxide,which was successfully applied to the determination of procalcitonin in the actual serum samples.It is easy to operate,real-time online detection with strong specificity and high sensitivity.Under the optimized experimental conditions,due to the porous layer structure modification,the PLOT capillary immunosensor enhanced 6.7 times improvement in the analytical sensitivity.The PLOT capillary based immunosensor was expected to be further applied to other tumor markets in clinic,and has a good application prospect for disease diagonsis.