DarkFocus: numerical autofocusing in digital in-line holographic microscopy using variance of computational dark-field gradient
Abstract We report on a novel computational technique for automatic numerical refocusing in digital in-line holographic microscopy. It is based on the adaptive filtering of the recorded on-axis hologram to eliminate its background term and extract interference intensity-component connected with light scattered on the sample (interference fringes). Numerical propagation of such filtered hologram yields the computationally generated dark-field imaging coming from the amplitude part of the complex field. We propose a simple measure in the form of the variance of the dark-field gradient, which attains its maximum value in the focal planes for all types of objects (phase, amplitude and mixed pha…
Quantitative phase imaging by single-shot Hilbert-Huang phase microscopy.
We propose a novel single-shot Hilbert-Huang transform-based algorithm applied to digital holographic microscopy (DHM) for robust, fast, and accurate single-shot quantitative phase imaging in on-axis and off-axis configurations. Fringe pattern with possible defects and closed fringes are adaptively filtered and accurately phase demodulated using local fringe direction estimation. Experimental validation of the proposed techniques is presented as the DHM study of microbeads and red blood cells phase samples. Obtained results compare very favorably with the Fourier approach (off-axis) and temporal phase shifting (on-axis).
Hilbert-Huang single-shot spatially multiplexed interferometric microscopy.
Hilbert-Huang single-shot spatially multiplexed interferometric microscopy (H2S2MIM) is presented as the implementation of a robust, fast, and accurate single-shot phase estimation algorithm with an extremely simple, low-cost, and highly stable way to convert a bright field microscope into a holographic one using partially coherent illumination. Altogether, H2S2MIM adds high-speed (video frame rate) quantitative phase imaging capability to a commercially available nonholographic microscope with improved phase reconstruction (coherence noise reduction). The technique has been validated using a 20×/0.46 NA objective in a regular Olympus BX-60 upright microscope for static, as well as dynamic…
Off-axis digital holographic multiplexing for rapid wavefront acquisition and processing
Off-axis holographic multiplexing involves capturing several complex wavefronts, each encoded into off-axis holograms with different interference fringe orientations, simultaneously, with a single camera acquisition. Thus, the multiplexed off-axis hologram can capture several wavefronts at once, where each one encodes different information from the sample, using the same number of pixels typically required for acquiring a single conventional off-axis hologram encoding only one sample wavefront. This gives rise to many possible applications, with focus on acquisition of dynamic samples, with hundreds of scientific papers already published in the last decade. These include field-of-view multi…
Automatic fringe pattern enhancement using truly adaptive period-guided bidimensional empirical mode decomposition.
Fringe patterns encode the information about the result of a measurement performed via widely used optical full-field testing methods, e.g., interferometry, digital holographic microscopy, moiré techniques, structured illumination etc. Affected by the optical setup, changing environment and the sample itself fringe patterns are often corrupted with substantial noise, strong and uneven background illumination and exhibit low contrast. Fringe pattern enhancement, i.e., noise minimization and background term removal, at the pre-processing stage prior to the phase map calculation (for the measurement result decoding) is therefore essential to minimize the jeopardizing effect the mentioned error…
Versatile optimization-based speed-up method for autofocusing in digital holographic microscopy
We propose a speed-up method for the in-focus plane detection in digital holographic microscopy that can be applied to a broad class of autofocusing algorithms that involve repetitive propagation of an object wave to various axial locations to decide the in-focus position. The classical autofocusing algorithms apply a uniform search strategy, i.e., they probe multiple, uniformly distributed axial locations, which leads to heavy computational overhead. Our method substantially reduces the computational load, without sacrificing the accuracy, by skillfully selecting the next location to investigate, which results in a decreased total number of probed propagation distances. This is achieved by…
Single-shot two-frame π-shifted spatially multiplexed interference phase microscopy
Single-shot, two-frame, π-shifted spatially multiplexed interference microscopy (π-SMIM) is presented as an improvement to previous SMIM implementations, introducing a versatile, robust, fast, and accurate method for cumbersome, noisy, and low-contrast phase object analysis. The proposed π-SMIM equips a commercially available nonholographic microscope with a high-speed (video frame rate) enhanced quantitative phase imaging (QPI) capability by properly placing a beam-splitter in the microscope embodiment to simultaneously (in a single shot) record two holograms mutually phase shifted by π radians at the expense of reducing the field of view. Upon subsequent subtractive superimposition of hol…
Variable zoom digital in-line holographic microscopy
Abstract We report on a novel layout providing variable zoom in digital in-line holographic microscopy (VZ-DIHM). The implementation is in virtue of an electrically tunable lens (ETL) which enables to slightly shift the illumination source axial position without mechanical movement of any system component. Magnifications ranging from ~15X to ~35X are easily achievable using the same layout and resulting in a substantial variation of the total field of view (FOV). The performance of the proposed setup is, first, validated using a resolution test target where the main parameters are analyzed (theoretically and experimentally) and, second, corroborated analyzing biological sample (prostate can…
Upgrading a brightfield optical microscope into a robust numerically advanced interference-based phase imager
The approach to convert a brightfield microscope into an interference-based versatile quantitative phase imaging unit is presented. It employs partially coherent illumination and diffraction grating. Enhanced interferogram bio-phase retrieval is performed by two-shot numerically-robust Hilbert-Huang method.
Single-shot slightly off-axis digital holographic microscopy with add-on module based on beamsplitter cube
Slightly off-axis digital holographic microscopy (SO-DHM) has recently emerged as a novel experimental arrangement for quantitative phase imaging (QPI). It offers improved capabilities in conventional on-axis and off-axis interferometric configurations. In this contribution, we report on a single-shot SO-DHM approach based on an add-on module adapted to the exit port of a regular microscope. The module employs a beamsplitter (BS) cube interferometer and includes, in addition, a Stokes lens (SL) for astigmatism compensation. Each recorded frame contains two fields of view (FOVs) of the sample, where each FOV is a hologram which is phase shifted by π rads with respect to the other. These two …