| High-throughput single-molecule force spectroscopy(H-SMFS)is one of the most important tools for abundant biodynamic such as single-molecule dynamics.The experimental efficiency and reliability are improved via measuring tens even hundreds of single-molecules simultaneously.However,there is no matching high-precision and real-time algorithm for its mass data.Digital holographic microscopy(DHM),as one of the hotpots in the field of optical micro-nanometer detection in the past 30 years,has the characteristics of contactless,high accuracy and real-time.In this paper,DHM and H-SMFS are combined to develop a digital holographic centrifugal force microscopy(DH-CFM)system,which realizes high-throughput and high-precision detection of single molecules in real time directly.The main research contains,1.Analyzing the method of tracing the beads in three-dimensions by DHM systematically,including the optical path,principles,and different implementation methods.And it is verified that the feasibility of applying it to H-SMFS systems.2.Based on the DHM and CFM,DH-CFM system is proposed.The contents include system principles,optical path’s building,measurement theories and methods.The errors caused by mechanical deformation,sample cell deformation and particle Brownian motion are analyzed and simulated.3.The specimen preparing method for SMFS systems is described.Analyzing the problems arising from the current SMFS specimen and the relevant researches such as viscous damping and specific linkage.Then,we demonstrate a new method of specimen preparation.Experimental results show that the proposed method improves the efficiency of H-SMFS more than 15.2 times.4.The step response and DNA stretching procession have detected following the proposed method by DH-CFM.The results manifest the resolution is within 5nm and DNA extension curves accord with Worm-like-chain(WLC)which was accepted worldly.It verifies the validity both the DH-CFM system and the specimen preparing method. |
PDF Full Text Request