3D Reconstruction of unstained cells from a single defocused hologram

We investigate the problem of 3D complex field reconstruction corresponding to unstained red blood cells (RBCs) with a single defocused off-axis digital hologram. We employ recently introduced mean gradient descent (MGD) optimization framework, to solve the 3D recovery problem. While investigating volume recovery problem for a continuous phase object like RBC, we came across an interesting feature of the back-propagated field that it does not show clear focusing effect. Therefore the sparsity enforcement within the iterative optimization framework given the single hologram data cannot effectively restrict the true object volume. For phase objects, it is known that the amplitude contrast of the back-propagated object field at the focus plane is minimum and it increases at the defocus planes. We therefore use this information available in the detector field data to device weights as a function of inverse of amplitude contrast. This weight function is employed in the iterative steps of the optimization algorithm to assist the object volume localization. The experimental illustrations of 3D volume reconstruction of the healthy as well as the malaria infected RBCs are presented. The proposed methodology is simple to implement experimentally and provides an approximate tomographic solution which is axially restricted and is consistent with the object field data.

Quantification of Pollen Viability in Lantana camara By Digital Holographic Microscopy

Pollen grains represent the male gametes of seed plants and their viability is critical for efficient sexual reproduction in the plant life cycle. Pollen analysis is used in diverse research thematics to address a range of botanical, ecological and geological questions. More recently it has been recognized that pollen may also be a vector for transgene escape from genetically modified crops, and the importance of pollen viability in invasion biology has also been emphasized. In this work, we analyse and report an efficient visual method for assessing the viability of pollen using digital holographic microscopy (DHM). We test this method on pollen grains of the invasive Lantana camara, a well known plant invader known to most of the tropical world. We image pollen grains and show that the quantitative phase information provided by the DHM technique can be readily related to the chromatin content of the individual cells and thereby to pollen viability. Our results offer a new technique for pollen viability assessment that does not require staining, and can be applied to a number of emerging areas in plant science.

Robust phase retrieval with complexity-guidance for coherent X-ray imaging

Reconstruction of a stable and reliable solution from noisy incomplete Fourier intensity data recorded in a coherent X-ray imaging (CXI) experiment is a challenging problem. The Relaxed Averaged Alternating Reflections (RAAR) algorithm that is concluded with a number of Error Reduction (ER) iterations is a popular choice. The RAAR-ER algorithm is usually employed for several hundreds of times starting with independent random guesses to obtain trial solutions that are then averaged to obtain the phase retrieval transfer function (PRTF). In this paper, we examine the phase retrieval solution obtained using the RAAR-ER methodology from perspective of the complexity parameter that was introduced by us in recent works. We observe that a single run of the RAAR-ER algorithm produces a solution with higher complexity compared to what is expected based on the complexity parameter as manifested by spurious high frequency grainy artifacts in the solution that do not seem to go away completely even after a number of trial solutions are averaged. We then describe a CG-RAAR (Complexity Guided RAAR) phase retrieval method that can effectively address this inconsistency problem and provides artifact-free solutions. The CG-RAAR methodology is first illustrated with simulated unblocked noisy Fourier intensity data and later applied to centrally-blocked noisy cyanobacterium data which is available from the CXIDB database. Our simulation and experimental results using CG-RAAR suggest two important improvements over the popular RAAR-ER algorithm. The CG-RAAR solutions after the averaging procedure is more reliable in the sense that it contains smallest features consistent with the resolution estimated by the PRTF curve. Secondly, since the single run of the CG-RAAR solution does not have grainy artifacts, the number of trial solutions needed for the averaging process is reduced.

Quantitative phase imaging of single particles from a cryoEM micrograph

We show that de-focused single particle images recorded using a cryo-electron microscope (cryoEM) system may be processed like a Fresnel zone in-line hologram to obtain physically meaningful quantitative phase maps associated with individual particles. In particular, a region-of-interest (ROI) of the de-focused image surrounding a particle can be numerically back-propagated, in order to determine accurate de-focus information based on the sparsity-of-gradient merit function. Further with the knowledge of de-focus information, an iterative Fresnel zone phase retrieval algorithm using image sparsity constraints can accurately estimate the quantitative phase information associated with a single particle. The proposed methodology which can correct for both de-focus and spherical aberrations is a deviation from the image processing chain currently used in single particle cryoEM reconstructions. Our illustrations as presented here suggest that the phase retrieval approach applies uniformly to de-focused image data recorded using the traditional CCD detectors as well as the newer direct electron detectors.

Dictionary based Approach to Edge Detection

Edge detection is a very essential part of image processing, as quality and accuracy of detection determines the success of further processing. We have developed a new self learning technique for edge detection using dictionary comprised of eigenfilters constructed using features of the input image. The dictionary based method eliminates the need of pre or post processing of the image and accounts for noise, blurriness, class of image and variation of illumination during the detection process itself. Since, this method depends on the characteristics of the image, the new technique can detect edges more accurately and capture greater detail than existing algorithms such as Sobel, Prewitt Laplacian of Gaussian, Canny method etc which use generic filters and operators. We have demonstrated its application on various classes of images such as text, face, barcodes, traffic and cell images. An application of this technique to cell counting in a microscopic image is also presented.

Sparsity assisted solution to the twin image problem in phase retrieval

The iterative phase retrieval problem for complex-valued objects from Fourier transform magnitude data is known to suffer from the twin image problem. In particular, when the object support is centro-symmetric, the iterative solution often stagnates such that the resultant complex image contains the features of both the desired solution and its inverted and complex-conjugated replica. The conventional approach to address the twin image problem is to modify the object support during initial iterations which can possibly lead to elimination of one of the twin images. However, at present there seems to be no deterministic procedure to make sure that the twin image will always be very weak or absent. In this work we make an important observation that the ideal solution without the twin image is typically more sparse (in some suitable transform domain) as compared to the stagnated solution containing the twin image. We further show that introducing a sparsity enhancing step in the iterative algorithm can address the twin image problem without the need to change the object support throughout the iterative process even when the object support is centro-symmetric. In a simulation study, we use binary and gray-scale pure phase objects and illustrate the effectiveness of the sparsity assisted phase recovery in the context of the twin image problem. The results have important implications for a wide range of topics in Physics where the phase retrieval problem plays a central role.