Construct An Electrochemical Cell Sensor Based On RBL Mast Cells To Detect Allergen Proteins In Food

Food allergy is a disease with frequent,a wide range and a serious reactioa Allergic reactions include skin,respiratory system,nerve eenter system,gastrointestinal system,etc.Severe allergic reactions can even be life-threatening.At present most of the allergens are indirectly evaluated by the detections of proteins and nucleic acids,and the detection results are highly susceptible to the external environment.Conpared to other conventional detection methods,cell sensing technology is more sensitive and specific.In this paper,three kinds of cell sensors with high specificity,high sensitivity,fast detection speed,accurate and reliable results were eonstructed to detect allergenic proteins in food.The results of this study are as follows:(1)In order to detect the peanut allergen protein Ara h2,the working electrode(magnetic glassy carbon electrode)was modified by using 0.40 mg/mL nano gold and 1.00 mg/mL nano magnetic beads to construct a three-eleclrode RBL mast cell electrochemical sensor.The current signal of the sensor had a good correlation with the cell concentration,and 1 ×107 cells/mL was selected as the optimal cell concentration The surface resistance value(Ret)of the magnetic glassy carbon electrode was proportional to the concentration of the peanut allergy protein Ara h2.The linear range of detection was 0.02 to 0.10 ng/mL,the correlation coefficient was 0.996.and the detection limit was 8.00 pg/mL.Combined with scanning electron microscopy,The results of transmisson electron microscopy and detection of Ca2+levels in RBL mast cells confirmed the reliability of the detection results of mast cell electrochemical sensors.(2)To detect casein in milk,and to improve the portability and mobility of the testing equipment and sinplify the post-processing operation of the testing equipment,a paper chip based on cellulose paper was designed and prepared as a testing platform.The working electrode of paper chip was modified by using 7.00 mg/mL carbon nano fiber and 3.00 mg/mL of graphene nanosheet was used to construct an electrochemical paper chro sensor based on RBL mast cell.The paper chip sensor had a good correlation with cell concentration,and 1×107 cells/mL was selected as the optimal cell concentratioa When casein was detected,the peak current value(Ip value)of the paper chip was inversely contrasted to the casein concentration,The linear range of detection was 0.10 to 1.00?g/mL,the correlation coefficient was 0.996,and the detection limit was 0.03 ?/mL.The detection results of electrochemical sensors and ELISA results were mutually validated.The results of scanning electron microscopy,transmission electron microscopy and detection of Ca2+levels in last cells were consistent.The feasibility of using mast cell-based electrochemical paper chip sensor to evaluate casein in milk was verified.(3)In order to better simulate the allergic environment in the body and reduce the cost of the experiment,compound 48/80(C48/80)was used as the detection target.The conceltration of 6.00 mg/mL molybdenum disulfide and 2 mmol/mL chloroauric acid paper were used to modified the working electrode of the chip.Co-culture of RBL cells with B cells constructed an electrochemical paper chip sensor based on cell co-culture system.There was a good correlation betWeen the paper chip sensor and the cell concentration of1×107 cells/mL was selected as the optimal cell concentration.Compound 48/80(C48/80)was a polymer produced by the condensation of N-methyl-p-methoxyphenethylamine with formaldehyde,which had the effect of promoting degranulation of mast cells.The results of electrochemical experiments showed that the current value of the paper chip was inversely proportional to the concentration of C48/80,the linear range was 0.02 to 0.10 ng/mL,the correlation coefficient was 0.991,and the detection limit was 0.21 ng/mL.The detection results of paper chip sensors were consistent with the results of scanning electron microscopy,transmission electron microscopy and detection of Ca2+levels in mast cells,indicating that the constructed paper chip sensor can be used for quantitative detection of C48/80.In summary,in this sstudy,combining electrochemical technology and microfluidic paper chips constructed three novel,sensitive and highly specific sensors based on mast cells,using mast cells as sensing elements.It was also used in the detection of sanples of various food allergens,providing new technical supports for the development of food safety testing technology.