Nanomagnetic Immunochromatographic Technology And Its Application In Pathogenic Microorganism Detection

Magnetic immunochromatography is based on immunochromatographictechnology which combines immunochromatographic technology withmagnetic nanoparticle. It would promote the development of the technologyto be more quantitative, high sensitivity, multielement and comprehensive.Nanobeads are superparamagnetic nanoparticles containing iron compounds,which are coated with polyethylene polymer material and cross-linked withcarboxyl or amino groups conjugated with functional group of proteinmolecules. Because of spherical and large surface area, the nanobeads couldcombine to pathogenic antigens in the three-dimensional space. Theproperties of magnetic and magnetic enrichment can quantitatively detecttrace amounts (pg level) of biomarkers and could be developed into usefulin-vitro diagnostic reagents. Furthermore, Magnetic signal can be interpretedby magnetic reading instrument into quantitative results. The advantages of magnetic immunochromatography make it become one of the important partof POCT (point of care testing) technology in the future.In this research，superparamagnetic beads were coupled with differentantibodies separately in standard EDC/NHS method according to thereference. We constructed the single-test and muti-testimmunochromatographic platform by optimizing chromatographic conditionsand materials. On the base of the platform, we designed theimmunochromatographic strips which could be applied on quantitativedetection of2009H1N1influenza virus and blood transfusion relatedpathogenic microorganisms (such as type B hepatitis, type C hepatitis,syphilis and HIV) in single or multiple test. During the experiment, weevaluated the sensitivity，specificity，stability and other associated charactersof the immunochromatographic strip. At the same time, we also identified theoptimizing reagents and materials in this system (such as the size of magneticbeads, sample volume, concentration and quantity of antibody) and analyzedthe influence from surfactant, macromolecule, stabilizing agent and salt in thechromatographic system.Our results showed that the constructed immunochromatographicplatforms were feasible in the detection of pathogenic microorganisms. TheHCG test strip and the HBsAg test strip have a high sensitivity andspecificity，which could meet the standards of commercial colloidal goldproducts. Sometime our immunochromatographic platforms even exceededthe results of ELISA and colloidal gold products. Especially, in the sensitivitydetection of HCG, our system could trace much lower concentration of HCG on1miu/ml in5min (concordance rate is99%). In the detection of HBsAg,the strip also exhibited a high sensitivity (0.1ng/ml) and reached to thestandard of similar products in ELISA analysis within5min. The results ofquantitative detection of2009/A/H1N1antigens showed that the visualdetection limitation of the magnetic immunochromatographic test strip was10.0ng/ml and the limitation of magnetic signal was100pg/ml, which is muchsensitive than those of ELISA and colloidal gold products. According to theresults of80clinical samples, it indicated that the sensitivity of our magneticstrip was85%. In the detection of influenza H3N2samples， thecross-reactivity was90%and no cross-reactivity with influenza B virussamples. In this study, we also found that the blood transfusionmulti-immunochromatographic strip system got a poor result compared to thesimple-immunochromatographic strip system in sensitivity and reaction time.It needs further work on optimizing the materials and reagents to constructmulti-immunochromatographic strip system for the detection of associatedpathogenic microorganisms in one system simultaneously.In this study, we explored a new approach towards the development ofbiomarker labeled superparamagnetic beads, which owns much advantage,such as more convenient and quick, higher sensitivity and stability, than thetraditional colloidal gold particles in lateral flow based onimmunochromatography. The detection results are objective and facilitatereal-time record-keeping at any time which is very important in the inspectionof the new biomarker during normal clinical application or battlefront.Furthermore, our immunochromatography system provides a solution to set more associated detections in one test to perform multiple pathogenicorganisms testing at the same time for conquering the problem of poorconditions and development in the inspection and prevention of sudden publichealth incidents and major infectious diseases in primary hygiene hospital...