The Prepration And Detection Of The Proteins In Cells Of The Semiconductor Quantum Dots Monoclonal Antibody Fluorescent Probes

Objective : To prepare Semiconductor 605nm Quantum Dots (605QDs)-Smad4,525nm Quantum Dots (525QDs)-Smad2 monoclonal antibody Fluorescent probes;to detect the optical qualities of 605QDs,525QDs , 605QDs-Smad4,525QDs-Smad2 monoclonal antibody Fluorescent probes ; To study the biocompatibility of semiconductor quantum dot to the rat dental papillae cells (RDPCs);to detect the ability to Specific Recognizing Smad4,2 in Rat dental papillae cells of 605QDs-Smad4,525QDs-Smad2 monoclonal antibody Fluorescent probes.Methods:①RDPCs isolated and cultured with enzyme digestion. Wave form fibroin and cytokeratin were used to identify the cells resource and the growth rules of cells were observed.②To study the effect of QDs to the RDPCs, The RDPCs were cultured with the 605QDs ,525QDs and those mixture in different density.③With the MTT, to study the cytotoxicity of QDs to the RDPCs, The RDPCs were cultured with the 605QDs ,525QDs and those mixture in different density.④605QDs ,525QDs were chemically modified with Smad4 and Smad2 proteins to prepare water soluble 605QDs-Smad4,525QDs-Smad2 monoclonal antibody Fluorescent probes which were purified after the preparation.⑤The properties of 605QDs ,525QDs,605QDs-Smad4,525QDs-Smad2 monoclonal antibody Fluorescent probes were studied through absorption band analyses, fluorescence spectra,fluorescence spectrum analyses and confocal laser scanning fluorescence microscopy.⑥Before the location of Smad4 and Smad2 proteins in RDPCs was studied with anti-Smad4,Smad2 immunocytochemical method and direct immunofluorescence imaging, RDPCs were incubated with TGF-β1. ; and the properties of 605QDs-Smad4,525QDs-Smad2 monoclonal antibody Fluorescent probes in RDPCs were detectedResults:①RDPCs cultured in vitro were well growing in DMEM/F12. Wave form fibroin staining was positive and cytokeratin staining was negative in RDPCs.②The 605QDs, 525QDs and those mixture in different density show no negative effect and cytotoxicity on the growth of the RDPCs.③QDs and monoclonal antibody covalently bond to form the Fluorescent probes which could specifically and effectively recognize Smad4,2 proteins in RDPCs. These Fluorescent probes still had good properties,including broad excite spectra,narrow emission spectra ,high fluorescence intensity and photostability.Conclusion:①The different QDs in a range of density have well biocompatibility to the live cells.②QDs and monoclonal antibody could covalently bond to form the Fluorescent probes with distinct optics character and photostability , which provides the scientific evidence that QDs trace the molecular movement in living cells in long-term, in situ and in real time .