A Study On Germ Cell-based Taste Sensor For The Application Of Bitterness Detection

As a defense mechanism of people,the bitter feeling plays an important role in people’s daily life.Bitter evaluation is widely applicated in many fields,such as food,toxicity detection,research of the mask of bitter taste.Due to the big uncertainty of the traditional method-people taste bitter substance has,in recent years,the electronic tongue based on chemical and biological gets widely development.However it is difficult to make one film to detect several substance,and for the biological method,the cell which can detect bitters is difficult to get.Taste cells use G protein-coupled receptor(GPCR)to detect bitter compounds,and the research finds that the mice sperm cells contain slot of bitter taste gene expression,which can be used to detect bitter tastes.When the GPCR is activated by bitter compounds,a series of cascades can lead to changes of the cytoskeletal and the addition of the intracellular calcium concentration.Cell impedance sensor can detect the change of the cytoskeletal resulted by GPCR.And use the fluorescence probe,the intracellular calcium concentration can be detected.Based on the above,this paper proposes two kinds of method to detect bitter compounds concentration.The main work and innovation of this paper are as follows:1.The design and application of electric cell-substrate impedance sensor and its detection system are studied deeply.This paper studies the cell impedance model of electric cell-substrate impedance sensor,the detection principle of the cell state of the impedance sensor,and the application of electric cell-substrate impedance sensor in GPCR detection.Then the design and processing of the cell impedance sensor chip and the size selection of the interdigital electrode are studied.It provides a detection system for subsequent experiments.2.Building a cell impedance sensor system based on sperm cells to detect bitter compounds.In this paper,we construct a cell impedance sensor system based on sperm cells to detect bitter compounds.Using sperm cells as primary sensing element,impedance sensor as transducer element to detect the bitters.With this sensor,we has detected three compounds,the linear detection of concentration range of the phenylthiourea is 10-200μmol/L,the detection limit is 4μmol/L;the linear detection of concentration range of the quinine is 62.5-1000μmol/L,the detection limit is 40μmol/L;the linear detection of concentration range of the phenylthiourea is 50-1000μmol/L,the detection limit is 13.5μmol/L.And this sensor has a good specificity on bitter detection.Compared with the traditional method that used reprogrammed cells to detect bitter substances,the germ cells have advantages of easily acquired,large numbers,and the impedance sensor can monitor the change of the cells in real-time without prejudice.3.Presenting a method based on sperm cells and fluorescence probe to detect bitter compounds.we have proposed a method that using sperm cell as primary sensing element,using Fluo4-AM and flow cytometry to detect the concentration of intracellular calcium ion to detect bitter compounds.With this sensor,we has detected three compounds,the linear detection of concentration range of the phenylthiourea is 25-1000μmol/L,the detection limit is 25.1μmol/L;the linear detection of concentration range of the quinine is 50-2000μmol/L,the detection limit is 5.7μmol/L;the linear detection of concentration range of the phenylthiourea is 50-4000μmol/L,the detection limit is 15.6μmol/L.And this sensor has a good specificity on bitter detection.Compared with the traditional method of calcium fluorescent imaging,this method’s operation is simple,calculation is easy.It will have a great potential for applications in bitter evaluation.