Study On The Construction And Calibration Of Optical Tweezers Based Force Spectroscopy

Optical tweezers based single-molecule force spectroscopy has opened up new avenue for biological research. Using the radiation pressure produced by a tightly focused continuous wave laser beam, an optical trap is formed stably. The nanometer and pico Newton ranges of distance and force capability to optical tweezers make them particular useful for trapping and manipulating micron-sized or nano-sized particles. So that scientists are gaining essential new, more sophisticated insights into the mechanical properties of biological macromolecules, the dynamics and mechanisms of molecular motors. In our system, we construct a dual-trap optical tweezers from two orthogonally polarized beams generated by a single laser with differential detection. Many optical components and much of the optical path traveled are common to both trapping beams, thus minimizing differential movements of one optical trap relative to the other and highly improving the system detection accuracy and anti-disturbance capacity.The main works in this paper are listed below:1. The single-molecule manipulation techniques, including atomic force microscopy, optical tweezers and magnetic tweezers are reviewed, then with an overview highlighting the respective areas of investigation by the three most common force spectroscopy techniques. According to the details concerning strengths, limitations and practical considerations of them, the construction of an dual-trap optical tweezers with differential detection is selected to apply pico Newton-level forces to micron-sized particles while simultaneously measuring displacement with nanometer-level precision. Then based on the mechanical principles of optical tweezers, the two physical diagrams of radiation pressure are discussed. 2. Design the optical system of dual-trap optical tweezers, and divide the optical path into several parts. More discussions have been given to the system requirements for the selection of the key technical parameters of optical components. Design related structures and mechanical clamping devices and arrange the positions of the elements on the optical platform. 3. The mathematical analysis of the conjugate structure is done, taking one part from piezo-driven mirror to the back focal plane of objective to study the effect of the off-center error on the conjugate performance. An approach to realize high quality optical conjugate in optical tweezers is proposed. Finally, study the critical problems appeared in the process of building the light path and provide some effective guidance. 4. The system is calibrated, including the optical trap displacement and the trap stiffness. With high precision piezoelectric displacement units installed as a reference sample stage and the CCD, the correspondence between the piezo-driven mirror and the trap displacement can be quantified. Then, several calibration methods of trap stiffness are discussed, such as power spectrum analysis, equipartition of motion and drag force method. 5. The tests of the mechanical characteristics of the samples are done, composing 1?m and 3?m silicon particles without being coted and 1?m particles adhered by ?-DNA strand.