Dynamic Digital Holographic Microscopy: A Noninvasive Contrast Imaging Technique Allowing Quantitative Visualization

To a great degree, the breakthrough in the field of the research of the life sciences dependon varies novel and powerful analysis measurement instruments. The microscopyapparatus is the most familiar employed. However, the conventional microscopyapparatus can not meet the development trend of the scientific research. Therefore, forthe active demand of the research of the life sciences, it is necessary to grope for a novelmicroscopical imaging technique that can achieve real-time, non-interference andquantitative analysis. This immediately concerns some important scientific problems,such as the exploration of mechanism of disease, the evaluation of the medical results,the drug screening and so on.With the rapid development of the computer and the charge coupled device (CCD)camera sensor technology, great progress had been made in technology of digital imageprocessing. At the same time, in combination with the holography and the optical fringeanalysis method, the digital holographic microscopy, a noninvasive contrast imagingtechnique allowing quantitative visualization of living cells, is developed. A CCD camerais employed to record a hologram onto a computer and numerical methods aresubsequently applied to reconstruct the holographic image to enable direct access to bothphase and amplitude information of the object wave in digital holography. It offers asignificant advantage for dynamic real-time analysis.The most popular numerical reconstruction methods for digital holography includethe well-known Fresnel diffraction integral method, the angular spectrum method and theconvolution-based method. In order to filter out the zero-order term, the twin image termand the parasitic interferences, the process of the spatial filtering must be carried out inthese methods. When some noises and parasitic interferences are introduced into thehologram, the spectrum of the virtual image would be disturbed by some other spectrum.It brings difficulties to define the spatial filter because of the blurry boundary andnon-regular distribution of the spectrum. Therefore, the quality of the reconstructedimage is mainly limited by the process of the spatial filtering. For the analysis ofdifferent object waves with respective spectrum, manual spatial filters with different pass-band are proposed. However, it shows that different results would be obtainedaccording to the spatial filters at different standard in the experiments, and it still bringsdifficulties to define a proper spatial filter at the uniform standard. On the other hand, forthe dynamic analysis, lots of holograms are recorded. Defining different manual spatialfilters would consume plenty of time for dynamic analysis. Thereby, the process of thespatial filtering limits the application of the digital holography for the dynamic andautomatic analysis. Therefore, it is necessary to grope for a numerical reconstructiontechnique that can be performed automatically at the uniform standard.In order to overcome the defect of the most popular numerical reconstructionmethods for the dynamic analysis, a numerical reconstruction technique of digitalholography by means of wavelet transform is described. Appling the wavelet transformto digital holography, the object wave can be reconstructed by calculating the waveletcoefficients of the hologram at the ridge or the peak of the wavelet coefficientsautomatically. At the same time the effect of the zero-order diffraction image and thetwin-image are eliminated without the spatial filtering. The theory, the results of thesimulations and experiments are demonstrated in detail in this paper.The theory mainly includes the following aspects:(1) Based on the processing ofthe spatial filtering, an automatic spatial filtering to obtain the virtual image term indigital holographic microscopy is presented.(2) A numerical reconstruction technique fordigital holography by means of the one-dimensional and two-dimensional Gabor wavelettransform is presented to achieve the dynamic and automatic analysis. The researchincludes the choice of the optimum mother wavelet, the decision of the values of thescale parameter and the accurate determination of the ridge of the Gabor wavelettransform.(3) A numerical reconstruction technique employing the de-multiplexing bymeans of the Gabor wavelet transform in digital holography is described.The simulations and experiments researches are mainly include the followingaspects: first, the numerical simulations are performed to demonstrate the validity of theaforementioned methods, and develop the programs and the application system byMatlab. Second, employ the apparatus analogous to a Mach-Zehnder interferometer forexperiments. Employ a CCD camera to recorder the holograms of the onion specimen and gastric cancer cells and perform the numerical reconstruction method. Finally, asequence of holograms of a grub is recorded for the dynamic and automatic analysis.The creative idea of this paper is that the Gabor wavelet transform, a tool excellingin multiresolution and localization in the time-or space-frequency domains, is applied tothe digital holography. The object wave can be reconstructed by calculating the waveletcoefficients of the hologram at the ridge or the peak of the wavelet coefficientsautomatically without the processing of the spatial filtering. This technique provides anovel method for the analysis of the biological samples, special for the living cells. It isof great significance in the development of the research of the life sciences and thedynamic analysis.