Research On Refractive Index Decoupling And Optical Path Integration In Multi Wavelength Dual Channel Phase Imaging

Cells are the most basic unit of life,and the quality of cells is related to human survival,health,immunity and heredity,and its quality is closely related to the physical characteristics of cells.Blood cells are cells that exist in the blood and can flow along with the blood throughout the body.Blood cells account for about 45 percent of the blood volume.Many blood cells have substructure morphology.Therefore,the research on the detection technology of substructure morphology of cells and their organelles is very important.The morphological and structural characteristics of blood cells reflect the detail structure of the interior of the cell and characteristics change,and the characteristics of its substructure morphological surface formation are mainly closely related to the cell refractive index.Therefore,it is of great significance to obtain the refractive index distribution of cells from the imaging information of cell morphological structure.Due to the light transparency of cells,traditional imaging methods cannot image the substructure morphology of cells.The recent development of phase microscopy imaging has been boon to imaging the morphology of cellular substructures,but in optical phase imaging the phase information contains both the physical thickness and the refractive index parameter of the cell,therefore,to determine the refractive index in this context requires decoupling the refractive index and thickness parameters of the cell’s phase information.In recent years,the development of phase micro-imaging technology brings good news to the morphological imaging of cell substructure.However,in optical phase imaging,phase information contains both the physical thickness and the refractive index parameter of the cell.Therefore,to determine the refractive index in this case requires decoupling the refractive index and thickness parameters of the cell’s phase information.It follows that the study of the decoupling method of cell refractive index and thickness is a very important research work in the field of cell quality detection technology.Current methods for refractive index decoupling of cells are mainly based on optical imaging or based on physicochemical principles,however,these methods have their own different disadvantages,such as: the structure of experimental equipment is complex,the calculation process is time-cons-uming,many hypothetical conditions and restrictive conditions,damage cell activity,etc.In this paper,we develop a new method to decouple cell refractive index and thickness based on phase diagrams using advanced optical phase micro-imaging techniques,in order to achieve a fast,non-destructive,and simple decoupling of cell refractive index and thickness.The main innovative research content of this paper is as follows:(1)For the complex morphological structure of heteromorphic nuclear cells,the nuclear refractive index is difficult to determine,using the three-dimensional phase images under dualwavelength phase micro-imaging,based on the dual-wavelength phase shift mathematical expression,according to the characteristic analysis of dispersion and phase gradient,the decoupling method of the cell refractive index and thickness is proposed.This method mainly applies the phase shift and dispersion theory,uses the phase distribution and phase gradient distribution characteristics and their relationship with the cell structure morphology,and obtains the mathematical expression results of the refractive index and thickness distribution under the representation of the dual-wavelength phase distribution function,thereby using stable phase imaging technology and relatively simple mathematical calculation to achieve the decoupling of the refractive index and thickness of single medium and double medium outer cell spheroid blood cells.This method can effectively determine the refractive index of heteromorphic nucleus of cells,meanwhile simulation experiment verification and result analysis were performed.(2)In view of the difficulties associated with refractive index decoupling for cells of arbitrary morphology,based on the optical phase micro-imaging method,the effect of threewavelength dispersion on phase imaging and its relationship with the characteristics of phase distribution is studied,and a three-wavelength refractive index decoupling method is proposed.This method utilizes the Cauchy’s law of dispersion after ignoring the high-power square,solved by mathematical solution,and finally the cytoplasmic and nuclear refractive indices can be obtained,and the corresponding thickness distribution can be further achieved.Simulation experiments and analysis of the results are performed,which show that the method in this paper achieves refractive index decoupling for either sphere-like or non-spherical cells and is applicable to any dispersive medium.From these,the thickness distribution of cytoplasm and nucleus could be reconstructed,which provides an important basis for analyzing the morphological characteristics of cell substructure.(3)Based on the technical basis of multi-channel phase micro-imaging in this paper,aiming at the shortcomings of complex structure and high operation requirements in the existing dualchannel phase imaging technology,a dual-channel orthogonal phase micro-imaging pentaprism sampling system is proposed.The sampling system utilizes the refraction and reflection characteristics of pentaprism and wavelength division multiplexing,which can solve the three shortcomings of the traditional dual-optical path orthogonal phase microsco-imaging optical path.One is that due to the imbalance of the devices,the two light sources will produce different systematic errors in the two orthogonal channels,which will affect the accuracy of morphological reconstruction.Second,in the traditional imaging,the sample is often carried on a static slide,here,the method of flow channel is adopted,and the normal incidence of both beams to the sampling channel is ensured,which can effectively improve the imaging sampling speed.Third,the traditional orthogonal dual-channel phase micro-imaging optical path has complex structure,many devices and high maintenance cost.The optical path structure with pentaprism as optical path modulation realizes efficient integration,and effectively solves the above problems.In addition,this paper also reports the research work on the improvement of photoelectric coupler devices during the internship at the graduate workstation(Jiangsu Allray optoelectronics Co.,Ltd.).Through the study and scientific research in the master’s study stage,the expected goal has been achieved.