A Study on Integrated Dual-Mode Holographic Tomography and Adaptive Wavefront Correction Technique for Free- Floating Single Live Cell Label-Free Imaging

Abstract

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Three-dimensional (3D) label-free refractive index (RI) imaging and analysis of a free-floating single live cell is a puzzling keyword for the biological research communities. Digital holographic tomography (DHT) is a potential technique to generate 3D RI profile of the biological specimens without labelling. The DHT uses interferometer configuration followed by either by sample rotation method or by beam rotation method. However, there is no such experimental system is developed to perform both full-angle sample rotation method and beam rotation method; this study developed such an integrated dual-mode tomography (IDT) system for the first time. The IDT system is developed by combining holographic optical tweezers (HOT) with digital holographic microscopy (DHM). The HOT system is used to control the free-floating live sample and the DHM records the transmitted wavefronts of the sample simultaneously. In this manner, the developed IDT system is capable of collecting and combing the spatial frequencies of full angle sample rotation with beam rotation method to extend the spatial frequency coverages along lateral and axial directions. Consequently, a novel unidentified flying object (UFO) like shaped experimental transfer function is obtained. To demonstrate the potential capability of the developed IDT method, a free-floating live Candida rugosa were used as a sample and its label-free 3D RI profile is generated at its sub-cellular level. The experimental results revealed that the IDT method can quantitatively enhance the lateral and axial resolutions without using any complicated image processing algorithm One of the major issues in an imaging system is the aberrations induced by the optical components and its alignments, which severely degrades the resolution and the imaging performance of the experimental system. This research study also focused on the development of a novel computer-generated hologram (CGH) based adaptive wavefront compensation technique demonstrated on a compactly developed structured illumination holographic tomography (SI-HT) system. A series of CGHs comprises of phase Fresnel lens and binary blazed grating are designed and displayed on a phase-only spatial light modulator to generate structured light pattern on the sample. The method is validated using Siemens star target and the potential application is verified using live sample candiada rugosa and its sub-cellular level 3D label-free RI profile is generated. The experimental results demonstrated the capability of the method to enhance the resolution in lateral and axial directions.