Development Of Array Sensors Based On Fluorescent Protein And Nanomaterials

Array sensor is a new sensing method inspired by the animal olfactory system.It consists of a series of sensing units and is a method based on analysis of the characteristic fingerprints obtained from the interactive response of the analytes.Because it has many advantages over traditional detection methods,such as fast,simple,and small damage to test samples,this method is widely used in environmental monitoring,safety screening,food inspection and biosensing.various heavy metal ion pollutions endanger human health and deteriorate our living environment,and it is an urgent need for simple,fast,sensitive,and low-cost detection methods in the field.Here,we developed a three-color fluorescent protein array sensor for the differentiation of metal ions.In addition,we constructed a RBC-Ti₃C₂/FPs fluorescence array sensor based on fluorescent protein?FPs?and erythrocyte membrane modified titanium carbide?RBC-Ti₃C₂?for cancer cell differentiation.The research content of this paper is as follows:?1?Cloning and expression of three fluorescent proteins with His-tag at both ends.Two plasmids,pET28a-h-mtagbfp-h and pET28a-h-sfcherry-h,were constructed by molecular cloning,and three fluorescent proteins,H-mTagBFP-H,H-EGFP-H and H-sfCherry-H were expressed and purified?pET28a-h-egfp-h was previously constructed in our lab?.The His-tag at both ends of the proteins can enhance the interaction between metal ions and the proteins,which is then used to design array sensors for the differentiation of metal ions and cancer cells.?2?The three-color fluorescent protein-based array sensor for differentiation of metal ions.Absorption and emission spectra of H-mTagBFP-H,H-EGFP-H,H-sfCherry-H have good overlapping,and fluorescence energy resonance transfer?F RET?can occur when the proteins are close enough.Coordination effects between His-tag at the ends of the proteins and metal ions,and electrostatic interaction between the negatively charged protein and metal ions,close the distance among proteins,and initiate different degrees of FRET.In addition,some metal ions(such as Cu²⁺)interact with proteins and cause electron transfer to quench the fluorescence.Different metal ions exhibit different mechanisms,resulting in different fluorescent signal changes,and linear discrimination analysis?LDA?was used to distinguish various metal ions.Metal ions in actual water samples were successfully measured by the sensor.?3?RBC-Ti₃C₂/FPs fluorescence array sensors based on FPs and RBC-Ti₃C₂ for differentiation of cancer cells.The RBC-Ti₃C₂/FPs fluorescence array sensor was constructed by modifying Ti₃C₂ with RBC and combining with four-color fluorescent protein?H-mTagBFP-H,ScGFP,H-sfCherry-H,IRFP?.Ti₃C₂ is capable of causing different degrees of fluorescence quenching of the four fluorescent proteins.In the presence of the cancer cells,part of the fluorescent proteins bound to the surface of the RBC is competed and achieve a certain degree of fluorescence recovery,due to the binding between the cancer cell membrane and the RBC.Different interaction between different cancer cells and RBC would cause different fluorescence signals.Four cancer cells?MCF-7,A549,HepG2,MDA-MB-231?was distinguished from the normal cell?LO2?,achieved by principal component analysis?PCA?.