Studies On The Preparation And Application Of All-solid-State Potentiometric Sensor For The Rapid Detection Of Bacteria

All-solid-state potentiometric biosensing technology has developed rapidly due to its passive,non-invasive,miniaturization and integration into paper-based and flexible materials easily.Recently,due to its portability,potentiometric sensor has attracted widespread interest in environmental monitoring of heavy metal ions,food testing for the related health indicators,clinical biodiagnosis and other aspects.Pathogenic are the important target objects for detection and identification in the fields of medicine,food safety,public health and safety.Bacteria,especially the intestinal bacteria,playing an important role in maintaining our health and is strongly associated with various diseases such as obesity,diabetes,inflammatory bowel disease and allergies.There are about 1,000 species of gut bacteria in a person’s gut,and gut microbes are not only linked to our general health,but also to autoimmune diseases and various intestinal diseases.At present,the common detection technology for bacterial are bacterial cell culture techniques,Raman(SERS),fluorescence,colorimetric and the electrical method.However,the current sensing technology requires a lot of time,causing unnecessary delays,or needs professional personnel operations,large and expensive instrument,besides,the detection precision is easily affected by the sample status which needs to be processed in some complex ways,these factors prevent them be prepared into portable device for field testing.Therefore,it is of great significance for developing a simple,rapid,easily miniaturized and highly sensitive method to applied in water resources monitoring,which is expected to promote the rapid detection of clinical samples.In this paper,we designed two different kinds of all-solid-state potentiometric biosensors based on aptamer for the detection of intestinal bacteria---Salmonella typhimurium and Staphylococcus aureus.The elution method of step-by-step is adopted at the first time to reduce the blank error in the detection which could improve the accuracy.The detection technology based on nucleic acid aptamer was used to amplify the detection response of bacteria and applied in actual samples.Specific research contents are as follows:Chapter One:Introduction the development status of bacterial detectionThis chapter gives a brief overview of foodborne pathogens firstly.Secondly,the development status of bacterial detection is introduced,which is summarized from two categories:the optical method and the electrochemical method.And then,the principle of potentiometric sensing is summarized from three aspects:the all-solid-state potentiometric sensing technology based on polymer membrane response,new molecular recognition mechanisms and the portable all-solid-state potentiometric sensing;Finally,the significance and the main content of this paper are summarized.Chapter Two:All-solid-state potentiometric sensor based on aptamer for detection of Salmonella typhimuriumAn all-solid-state potentiometric aptamer sensor based on disposable screen-printed carbon electrode(SPCE)was designed for the detection of Salmonella typhimurium in water sample.In this paper,the designed sensor was prepared,which was prepared by electrochemical reduction of GO on a disposable SPCE to enhance the electrical conductivity of the electrode,and the effective surface area of the electrode was increased due to the formation of dendritic gold molecules on the electrode.The aptamer was attached to the electrode surface via Au-S,the combination of the aptamer with the target makes the aptamer form a rigid three-dimensional structure,which changes the charge density of the electrode surface and the potential.The detection limit of the sensor was 79 CFU/m L and the detection range was 10~2～10~7 CFU/m L.Besides,the sensor has a better repeatability and stability.This sensing strategy is simple and rapid,which provides a better method for sensitive detection of Salmonella typhi in the real samples.Chapter Three:All-solid-state potentiometric sensor based on double-stranded aptamer for the detection of Staphylococcus aureusIn this paper,a competitive all-solid-state potentiometric sensor based on double-stranded aptamers were proposed for the detection of Staphylococcus aureus.The graphene oxide(GO)and dendritic gold(Den Au)were modified screen-print electrode through self-assembly.Staphylococcus aureus aptamer complementary strand connection on the electrode,the complementary strand connects with staphylococcus aureus aptamer through base complementation,which made complementary strand away from the surface of electrode,the binding of the aptamer to the targrt causes the aptamer leaving the electrode surface,then the complementary strand returns to the electrode surface.The potential change of shedding is greater than that of conformational change,thereby,the sensitivity was amplied.Under the optimal conditions,in the range of 4×10~2～4×10~6 CFU/m L,the sensor shows a linear relationship,and the detection limit is 59 CFU/m L.Besides,the sensor has good reproducibility,selectivity and stability and has been applied to the detection of water samples,so,it provides a simple and accurate scheme for the detection of bacteria.