Double Stranded DNA Fluorescence Probes Are Used For Fluorescence Imaging And Quantitative Determination Of Cell Surface Molecules

In biological experiments,it is often necessary to perform staining imaging on the surface or internal molecules of the cells and to determine the content of the molecules.Normally,the two experiments require two sets of systems to be separated.A double-stranded DNA-based fluorescent probe has been successfully constructed in our laboratory for fluorescent imaging of cell surface molecules.respectively biotin-DNA/SYBR GREEN and biotin-DNA-ROX.There are two major defects:(1)The fluorescence of the biotin-DNA-ROXprobe was poor when the cell imaging was performed because only coupling one ROX;(2)Both types of probes can only be specifically recognized and bound by biotin(biotin)and Streptavidin.If a certain protein on the cell surface is to be stained,multi-level coupling is needed(for example,the middle one needs to Protein antibodies),multi-level coupling not only increase the number of staining steps,but also allow not simultaneous imaging of two molecules.The purpose of this topic is twofold:(1)To improve the double-stranded DNA fluorescent probes we have constructed and make it more perfect in the fluorescence imaging of cell surface molecules;(2)To construct a novel method for in situ quantification of cell surface molecules so that our double-stranded DNA fluorescent probes can be used not only for imaging of cell surface molecules but also for in situ molecular quantification of cell surface molecules.This topic consists of three parts:(1)preparation and identification DNA(preparation of DNA,then identification at the molecular and cellular levels);(2)Solving two problems in our existing double-stranded DNA fluorescent probes,and verify that the double-stranded DNA fluorescent probe can be used for fluorescence imaging of molecules on the cell surface;(3)Verifying that the double-stranded DNA fluorescent probe can be used for in situ quantitative quantification of molecules on the cell surface.The experimental results prove that,our double-stranded DNA fluorescent probes can be used not only for the imaging of cell surface molecules,but also for quantifying the number of molecules on the surface of cells in situ.Moreover,the method is simple,rapid,effective,and inexpensive.