Research On Label-free Quantitative Microscopic Imaging Based On Digital Holography

Imaging of biological cells and tissues is important of biological research and medical diagnosis,and microscopy has became one of the most commonly tools in medicine and biology.However,the traditional microscopic technology can only obtain the two-dimensional imaging of the sample,but can't obtain its three-dimensional information.We can not only measure the volume of the sample,but also gain the physical characteristics of the cell,such as dry weight and the refractive index,and we can realize high-precision observation and diagnostic research by using these information.However,existing tomographic technologies are often restricted by such factors as low contrast,slow imaging speed and the fluorescence-label.Therefore,the study of new quantitative microscopic tomography imaging technology has guiding significance for the development of multi disciplines,and can meet the needs of the biological and medical communities for accurate observation of tiny objects.Free-label quantitative microscopic tomography based on digital holography is a popular technology in recent years.This technology is based on holography and combined with the relationship of phase and the refractive index and united the theory of fourier diffraction to restore the refractive index of transparent sample.This tomographic technology does not need fluorescent labeling of cells or tissues,so it's not affected by photo-bleaching,and this technology has a quickly imaging speed.At the same time,it has a accurate result of reconstruction and can retrieve the three-dimensional information of the sample.As a non-contact and fast 3D quantitative microscopic tomography,its development will have an important effect on biomedical specific diagnosis,special plant seed structure determination and other fields.This thesis mainly etomography imaging technology based on two kinds of interference techniques,and provesxpounds the theory and experiment of label-free quantitative microscopic the feasibility of the theory through simulation and experiment.The article mainly contains the following work:(1)Theoretical and experimental research on label-free quantitative microscopic tomography based on dual-wavelength digital holography.Firstly,after a brief explanation of the basic theory,the information recovery process is introduced and the recovery process is numerically simulated using Matlab.Secondly,we introduces the advantages of LED as a lighting source,and built an experimental device based on the Linnik interference device.The phase distribution information of each layer was obtained by processing holograms of different layers of the sample.Using the relationship between the reproduction phase and the refractive index,the refractive index distribution of the transparent sample is reconstructed.(2)The theoretical and experimental methods for quantitative microscopic measurement of free-label samples based on quadriwave lateral shearing interferometry were reviewed.The observation of the internal refractive index of the sample was realized by multi-angle illumination recording.After introducing the quadriwave lateral shearing theory and the Fourier diffraction theory,we use the Mtalab to simulate the quadriwave lateral shearing interferometry to recover the amplitude and phase information of the wavefront,at the same time,we used LED as the illumination source and designed a specific lighting hole,and this hole can change the different positions by moving the two-dimensional translation stage in the plane of the light source to obtain the sample information under different illumination angles.After doing this,we can use the quadriwave lateral shearing interferometry to recover the complex amplitude information of the sample to be tested under the multi-angle.By using a special algorithm based on Fourier diffiraction theory and an iterative algorithm to reproduce the refractive index distribution of the weakly absorbing sample.At the same time,the internal refractive index distribution of 3t3-Ll preadipocytes was measured to prove the feasibility of the theoretical and experimental methods,and we were evaluated its imaging characteristics.