Study On High-sensitive Immunochromatography Detection Of Escherichia Coli O157:H7

Foodborne illness caused by pathogens are one of the greatest threats to global health problems.Escherichia coli（E.coli）O157:H7 is the most common colorectiperiosis associated with foodborne disease outbreaks,causing extensive attention from researchers.Therefore,it is necessary to conduct real-time and continuous monitoring of pathogen level to obtain valuable information,so as to assist health institutions in guiding the prevention and control of pathogen related epidemic.Up to now,most pathogenic bacteria are detected through traditional techniques,which rely on professionals and are limited by processing time and point-of-care testing availability.Immunochromatographic test strips（ITS）method based on nanotechnology has attracted much attention because of its unique optical properties,it opens up a new way for early and rapid detection of foodborne pathogens.Escherichia coli（E.coli）O157:H7 is the most common colorectiperiosis associated with foodborne disease outbreaks,causing extensive attention from researchers.In this study,two sensitive and efficient new label-free immunochromatographic rapid detection methods were established to monitor E.coli O157:H7.The main research results are as follows:（1）CuS-based ITS and its application to label-free detection of E.coli O157:H7 in food.In this study,we found a kind of molecular recognition chromogenic agent that could directly adsorb with pathogens,which could be used to construct a new sandwich structure of“nanoprobe-pathogen-antibody”.This structure not only did not require paired antibodies,but also did not require a probe labeling process.Therefore,a novel ITS for E.coli O157:H7 detection based on dual recognition factors（nanomaterials and antibody）was developed.Due to the simple synthesis steps of CuS nanoclusters（NCLs）and the good electrostatic adsorption ability between CuS NCLs and E.coli O157:H7 with negative charge on the surface,we only needed to incubate CuS NCLs with pathogenic bacteria at room temperature for a period of time to obtain new probes.After the optimization of CuS NCLs concentration,immunoreaction time and others in the detection process,the specific antibody was used as the recognition factor on the T-line to realize the sensitive detection of E.coli O157:H7.The sensitivity of naked eye detection was 10⁴CFU/m L,there was no crossing reactions with other pathogenic bacteria and showed good specificity.Additionally,E.coli O157:H7 was successfully detected in beef,chicken,milk and honey samples.The v LODs of CuS-ITS were 10⁵,10⁴,10⁴and 10⁵CFU/m L,and the recovery rate was between80%and 120%,which showed a good practical application value.Therefore,the developed CuS NCLs-based novel label-free sandwich ITS provided a simple,reliable,fast and sensitive E.coli O157:H7 detection method.（2）Fe₃O₄@CuS-based ITS and its application to label-free and colorimetric-photothermal dual-readout detection of E.coli O157:H7 in food.In order to improve the detection sensitivity and electrostatic adsorption capacity between nanomaterials and pathogenic bacteria,synthesized Fe₃O₄@CuS nanostructures（NSs）by a simple hydrothermal-chemical deposition（HD-CD）strategy in an aqueous solution procedure.It not only enhanced the strong capture ability to bacteria,but also had good photothermal effects.Under the radiation of 808 nm infrared light,the temperature of the nanomaterial raised,and the temperature difference was recorded with an infrared thermal imager,then a label-free and dual-readout ITS was developed.The concentration,volume of Fe₃O₄@CuS NSs,incubation time and immunoreaction time were optimized.Under the optimal conditions,the high sensitivity colorimetric and photothermal test of E.coli O157:H7 had a wide range（10²-10⁸CFU/m L）and high specificity.And its detection limit,as low as 10³and 10²CFU/m L,could be monitored in colorimetric and photothermal modes.In addition,the detection limits of E.coli O157:H7 in beef,chicken,milk and honey were 10³,10²,10³and 10²CFU/m L by this proposed platform with a recovery of 83%-120%.Through this technique,the reliability of ITS in the detection of pathogens in complex food samples was confirmed.