Hongyue Sun

In Situ Underwater Electronic Holographic Camera for Studies of Plankton

In this paper, we describe an underwater electronic holographic camera (eHoloCam) that has been developed for in situ studies of the distribution and dynamics of plankton and other marine organisms and particles. The eHoloCam uses an Nd-YAG pulsed laser to freeze-frame fast moving particles and a complementary metal-oxide-semiconductor (CMOS) sensor for high-resolution image capture. Digital holograms and holographic videos are recorded at rates from 5 to 25 Hz over a period of several hours. Data is stored locally on an embedded computer. The eHoloCam is capable of recording all organisms and particles located in a water volume of 36.8 cm3 in a single hologram frame. The recorded holographic videos may subsequently be reconstructed numerically at a desired image plane. The main optical and mechanical specifications for eHoloCam are also described. To record electronic holographic videos of marine organisms, the eHoloCam was deployed from a towed sampling frame [autorecording instrumented environmental sampler (ARIES)] on the research vessel RV Scotia at speeds up to 4 kn (about 2 mldrs-1) in the North Sea off the Shetland Isles. Various images of marine organisms obtained from this deployment are shown, together with preliminary measurements on the distribution of Calanus copepods.

Underwater digital holography for studies of marine plankton

Conventional and digital holographies are proving to be increasingly important for studies of marine zooplankton and other underwater biological applications. This paper reports on the use of a subsea digital holographic camera (eHoloCam) for the analysis and identification of marine organisms and other subsea particles. Unlike recording on a photographic film, a digital hologram (e-hologram) is recorded on an electronic sensor and reconstructed numerically in a computer by simulating the propagation of the optical field in space. By comparison with other imaging techniques, an e-hologram has several advantages such as three-dimensional spatial reconstruction, non-intrusive and non-destructive interrogation of the recording sampling volume and the ability to record holographic videos. The basis of much work in optics lies in Maxwells electromagnetic theory and holography is no exception: we report here on two of the numerical reconstruction algorithms we have used to reconstruct holograms obtained using eHoloCam and how their starting point lies in Maxwells equations. Derivation of the angular spectrum algorithm for plane waves is provided as an exact method for the in-line numerical reconstruction of digital holograms. The Fresnel numerical reconstruction algorithm is derived from the angular spectrum method. In-line holograms are numerically processed before and after reconstruction to remove periodic noise from captured images and to increase image contrast. The ability of the Fresnel integration reconstruction algorithm to extend the reconstructed volume beyond the recording sensor dimensions is also shown with a 50% extension of the reconstruction area. Finally, we present some images obtained from recent deployments of eHoloCam in the North Sea and Faeroes Channel.

Visualization of fast-moving cells in vivo using digital holographic video microscopy

Digital in-line holography offers some significant advantages over conventional optical holography and microscopy to image biological specimens. By combining holography with digital video microscopy, an in-line holographic video microscope is developed and is capable of recording spatial 3D holographic images of biological specimens, while preserving the time dimension. The system enables high-speed video recording of fast cell movement, such as the rapid movement of blood cells in the blood stream in vivo. This capability is demonstrated with observations of fast 3-D movement of live cells in suspension cultures in response to a gentle shake to the Petri dish. The experimental and numerical procedures are incorporated with a fast reconstruction algorithm for reconstruction of holographic video frames at various planes (z axis) from the hologram and along the time axis. The current system enables both lateral and longitudinal resolutions down to a few micrometers. Postreconstruction processing of background subtraction is utilized to eliminate noise caused by scattered light, thereby enabling visualization of, for example, blood streams of live Xenopos tadpoles. The combination of digital holography and microscopy offers unique advantages for imaging of fast moving cells and other biological particles in three dimensions in vivo with high spatial and temporal resolution.

Observations of coastal sediment erosion using in-line holography

This paper demonstrates how in-line particle field holography may be used to study erosion processes in coastal sediments, in particular to visualize the morphologies of eroded sediment particles and to investigate how organic matter affects the floc size and erosion threshold in incipient sediment erosion. In-line holography produces 3D images and provides higher resolution over larger recording volume compared with other imaging techniques. The study examines the dependence of the observed shearing stress, particle size distributions and morphologies of particles on the duration of desiccation of the sediment bed, and on the concentration of extra-cellular polymeric substance. Both artificial sediment (produced from cleaned sediments by the addition of natural extra-cellular polymeric substance) and intact sediment were used. The required hydrodynamic shear stress to produce erosion on the sediment beds was generated using a cohesive strength meter (CSM). Characterization of the eroded particle properties was carried out using 2D image analysis.

eHoloCam - an electronic holographic camera for subsea analysis

Recently we have developed an underwater holographic camera for the analysis of plankton and other marine organisms. This camera (HoloMar) was unique in that it was able to record simultaneous in-line and off-axis holograms to cover a range of size of marine organisms from a few microns to tens of millimetres and at concentrations from a few particles per cubic centimetre to dense aggregates. However, HoloMar suffered for being physically large and heavy and difficult to deploy. It also was based on the use of photographic emulsions to record the holograms. To overcome some of these difficulties we have started to develop a new holographic camera (eHoloCam) based on digital holography. In electronic or digital holography (eHolography) an electronic hologram is directly recorded onto a CCD or CMOS sensor and then numerically reconstructed by simulation of the optical hologram reconstruction. In this paper, we discuss some of the possible optical layouts and algorithms under consideration. We present some eHolograms produced in the laboratory prototypes of the eHoloCam

Ocean Plankton Imaging Using an Electronic Holographic Camera

In this paper, we describe an electronic holographic camera that has been developed for in situ underwater studies of the distribution and dynamics of plankton and other marine organisms and particles. Holographic data are stored on an embedded computer in the camera for later data extraction and analysis. We describe the main optical and mechanical specifications and outline the design, development and operation of eHoloCam. We summarise the eHoloCam performance in four in situ deployments in North Sea and Faeroe Channels at water depths ranging from about 10 m to 450 m with research vessel Scotia. eHoloCam is capable of capturing opaque and transparent organisms in the size range from about 50 mum up to a 10 millimetres. A clear advantage of eHoloCam over other imaging and counting techniques is the ability to capture high-resolution images without destroying the organism. This feature is extremely valuable in facilitating new in-situ studies of plankton dynamics. The recorded holographic videos are reconstructed numerically using one of our reconstruction algorithms at various planes through the light path. The overall system resolution for the recorded images is 8 mum and 36 mum at a distance of 100 mm and 470 mm, respectively. We show various images of marine organisms recorded on these 4 cruises, and preliminary data on size distributions of Calanus are also presented and discussed.

Optical configurations for oceanographic applications of eHolography

eHolography (electronic or digital holography) has been used for observing and measuring particle size and distribution in biomedicine, sedimentology and oceanology. In these areas, eHolography possesses advantages over other imaging techniques in terms of its capability for recording live organisms (e.g. plankton) or events (e.g. sediment erosion) in a non-disturbed environment. Compared with classical holography, an eHoIogram is recorded digitally on an electronic imaging sensor and reconstructed numerically, to produce a three-dimensional map of the scene. In this paper, we present our progress in developing an electronic Holographic Camera for high-resolution imaging, both subsea and in air (eHoloCam). Many optical configurations have been successfully used in classical (photographic) holography, including a back-illuminating inline beam with and without a separated reference beam, and side/front illuminated object with an off-axis reference beam. For eHolography, it is necessary to reexamine these and other optical geometries with specific reference to the constraints of reduced resolution and recording area of current CCD or CMOS imaging sensors. For example, using a divergent beam in recording back-illuminated in-line eHolograms can give an improvement in image resolution over the more usual collimated beam geometry. Furthermore, when using an off-axis beam in eHolography considerable limitations are encountered in obtaining useable holograms, such as the need to record at small reference beam angles. In this paper, we will discuss the various geometries we have used to overcome these difficulties.

Digital Holography for Imaging Tissue Cells Using Coherent Lights

Visible and near-infrared lasers are examined to see how laser coherence length and wavelength affect the image quality in digital holographic microscopy. With opaque and partially transparent animal tissues, NIR-lasers show advantages over visible laser. Article not available.

Holographic Image Acquisition and Remote Underwater Visualization

Currently there are two approaches to large volume data retrieval for underwater holography: optical methods using high resolution emulsions and digital holography. Underwater digital holography has enabled high resolution real-time data to be obtained with depths-of-field otherwise unobtainable by conventional non-holographic optical techniques. However, conventional emulsion methods provide large volume retrieval at a superior resolution. This paper will discuss a twin camera automated image retrieval system for in-line emulsion holographic reconstruction and future alternative real-time reconstruction methods using digital holography. In both techniques image acquisition and data retrieval from reconstructed holograms presents a number of challenges that hinder the ability to readily extract information. The application of holography to store high resolution three-dimensional data for large volumes is well understood, however here also belies a problem for data retrieval and large information storage, the data extraction is time-consuming and requires manual intervention. The automated holographic reconstruction system has been developed to aid the analysis, databasing and identification of objects captured by in-line holography. Whereas, the digital off-axis system presents an alternative real-time visualisation approach for underwater analysis. Digital off-axis real-time reconstruction will be presented using numerical pre-processing of captured holograms for data extraction and visualization using a spatial light modulator (SLM). This method has the advantage of recording surface information and optimizing the recording quality of holograms in-situ without the necessity of numerical post-processing. Preliminary experimental research has demonstrated optical and numerical reconstructions at interactive frame rates. Observed interlacing artifacts caused by the CCD camera used are presented and discussed. Optical and numerical reconstructions are presented using a spatial light modulator. Fourier methods are presented using a Negative Laplacian finite impulse response filter to pre-process holograms for optimizing the numerical and optical reconstructionn.