Biological Cell Imaging Research By Full-angle Digital Holographic Micro-tomography

Digital holographic micro-tomography is a new three-dimensional microscopic imaging technology.It records the digital holograms of samples under different illumination angles through traditional digital holographic microscopes and reproduces the complex amplitude data,and the three-dimensional image of the sample is obtained by tomographic reconstruction algorithm.Digital holographic micro-tomography has the ability to perform non-destructive three-dimensional imaging of small biological samples or special materials without labels.It can quantitatively measure the threedimensional refractive index distribution of the sample to reveal the internal structure of the sample.It has a broad application prospect in biological research,medical diagnosis,material analysis.In digital holographic micro-tomography,there are two methods for obtaining digital holograms of samples at different observation angles.The first is the beam scanning method,which keeps the sample fixed and changes the direction of beam illumination to scan the sample.The other is the sample rotation method,that is,the sample is rotated without changing the direction of the illumination side.The beam scanning method is limited by the numerical aperture of the micro-objective lens at the recording angle of view.Only the observation data of the sample in the range of about 140 degree can be obtained,and the problem of missing spectrum will occur.The sample rotation method allows the sample to be rotated 360 degree to obtain a fullangle digital hologram,thereby improving the accuracy of phase measurement and the uniformity of reconstruction results.However,in the process of obtaining a projection using the sample rotation method,because the position of the sample on the turntable is not easy to be completely centered or the vibration caused by the inertia of the turntable and the bad environment,it will inevitably cause the sample to shift axially and laterally.Because the ideal depth of field is small,the shift error can cause the sample to deviate from the ideal depth of field and reduce the quality of the digital hologram.In this paper,the imaging process is improved based on the sample rotation method in digital holographic microtomography technology.The depth of field tracking combined with digital refocusing is used to correct the axial displacement error,and the cross-correlation algorithm is used to correct the lateral displacement error of the sample.The basic principle of digital holographic microtomography technology was expounded.The basic process of off-axis hologram recording of samples at different observation angles was described.The principle of reproducing complex amplitude distribution of object light field by off-axis hologram was introduced.The reconstruction algorithm based on Fourier center slice theorem and the reconstruction algorithm based on Fourier diffraction projection theorem was expounded.We propose a correction method for sample offset errors and optimized reconstruction methods based on full-angle digital holographic micro-tomography.For the near-circle motion displacement error in the rotation process caused by the sample not being completely centered,the sample stage compensates the displacement of the sample during the rotation process,so that the sample is always maintained within the ideal depth of field.The angular spectroscopy method is used to phase retrieve.The digital refocusing process was performed,that is,the reproduction distance was gradually increased and the focus criterion function was used on the amplitude diagram to find the best reproduction distance to further correct the axial error.Quality-oriented unwrapping algorithm is applied to improve the anti-noise performance.Polynomial fitting is used to remove the background phase gradient to ensure continuous hologram capture to reduce the impact of environmental changes.The effect of correcting the shift error is verified.Experimental results show that depth-of-field tracking combined with digital refocusing technology can effectively reduce the axial offset error during sample rotation,improve the accuracy of phase measurement,and the application of cross-correlation algorithm can effectively correct the lateral error.The three-dimensional refractive index of porcine adipocytes was obtained using the imaging method.We tried the application of digital holographic micro-tomography technology in the pathological diagnosis of cancer cells.We took the mouse breast cancer cells as the research object and measured its three-dimensional refractive index distribution to obtaine its three-dimensional pathological model.Compared with traditional pathological sections,the application potential of this imaging technique in cytological pathological diagnosis is verified.