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Off-axis digital holographic microscopy (DHM) provides both amplitude and phase images, and so it may be used for label-free 3D tracking of micro- and nano-sized particles of different compositions, including biological cells, strongly absorbing particles, and strongly scattering particles. Contrast is provided by differences in either the real or imaginary parts of the refractive index (phase contrast and absorption) and/or by scattering. While numerous studies have focused on phase contrast and improving resolution in DHM, particularly axial resolution, absent have been studies quantifying the limits of detection for unresolved particles. This limit has important implications for microbial detection, including in life-detection missions for space flight. Here we examine the limits of detection of nanosized particles as a function of particle optical properties, microscope optics (including camera well depth and substrate), and data processing techniques and find that DHM provides contrast in both amplitude and phase for unresolved spheres, in rough agreement with Mie theory scattering cross-sections. Amplitude reconstructions are more useful than phase for low-index spheres and should not be neglected in DHM analysis.

 

Vortex fiber nulling (VFN) is a technique for detecting and characterizing faint companions at small separations from their host star. A near-infrared (∼2.3μm) VFN demonstrator mode was deployed on the Keck Planet Imager and Characterizer (KPIC) instrument at the Keck Observatory and presented earlier. In this Letter, we present the first VFN companion detections. Three targets, HIP 21543 Ab, HIP 94666 Ab, and HIP 50319 B, were detected with host–companion flux ratios between 70 and 430 at and within one diffraction beamwidth (λ/D). We complement the spectra from KPIC VFN with flux ratio and position measurements from the CHARA Array to validate the VFN results and provide a more complete characterization of the targets. This Letter reports the first direct detection of these three M dwarf companions, yielding their first spectra and flux ratios. Our observations provide measurements of bulk properties such as effective temperatures, radial velocities, and$v\sin i$, and verify the accuracy of the published orbits. These detections corroborate earlier predictions of the KPIC VFN performance, demonstrating that the instrument mode is ready for science observations.

 

We observed HD 19467 B with JWST’s NIRCam in six filters spanning 2.5–4.6μm with the long-wavelength bar coronagraph. The brown dwarf HD 19467 B was initially identified through a long-period trend in the radial velocity of the G3V star HD 19467. HD 19467 B was subsequently detected via coronagraphic imaging and spectroscopy, and characterized as a late-T type brown dwarf with an approximate temperature ∼1000 K. We observed HD 19467 B as a part of the NIRCam GTO science program, demonstrating the first use of the NIRCam Long Wavelength Bar coronagraphic mask. The object was detected in all six filters (contrast levels of 2 × 10⁻⁴to 2 × 10⁻⁵) at a separation of 1.″6 using angular differential imaging and synthetic reference differential imaging. Due to a guide star failure during the acquisition of a preselected reference star, no reference star data were available for post-processing. However, reference differential imaging was successfully applied using synthetic point-spread functions developed from contemporaneous maps of the telescope’s optical configuration. Additional radial velocity data (from Keck/HIRES) are used to constrain the orbit of HD 19467 B. Photometric data from TESS are used to constrain the properties of the host star, particularly its age. NIRCam photometry, spectra, and photometry from the literature, and improved stellar parameters are used in conjunction with recent spectral and evolutionary substellar models to derive the physical properties of HD 19467 B. Using an age of 9.4 ± 0.9 Gyr inferred from spectroscopy, Gaia astrometry, and TESS asteroseismology, we obtain a model-derived mass of 62 ± 1M_J, which is consistent within 2σwith the dynamically derived mass of${81}_{-12}^{+14}$M_J.

 

Described over 100 years ago, the Gouy phase anomaly refers to the additional$\mathrm{\pi <\#comment/>}$phase shift that is accumulated as a wave passes through focus. It is potentially useful in analyzing any type of phase-sensitive imaging; in light microscopy, digital holographic microscopy (DHM) provides phase information in the encoded hologram. One limitation of DHM is the weak contrast generated by many biological cells, especially unpigmented bacteria. We demonstrate here that the Gouy phase anomaly may be detected directly in the phase image using the z-derivative of the phase, allowing for precise localization of unlabeled, micrometer-sized bacteria. The use of dyes that increase phase contrast does not improve detectability. This approach is less computationally intensive than other procedures such as deconvolution and is relatively insensitive to reconstruction parameters. The software is implemented in an open-source FIJI plug-in.

 

Following the previous article, here we describe the first field demonstration of the ELVIS system, performed at Newport Beach, CA. We examined ocean water to detect microorganisms using the combined holographic and light-field fluorescence microscope and successfully detected both eukaryotes and prokaryotes. The shared field of view provided simultaneous bright-field (amplitude), phase, and fluorescence information from both chlorophyll autofluorescence and acridine orange staining. The entire process was performed in a nearly autonomous manner using a specifically designed sample processing unit (SPU) and custom acquisition software. We also discuss improvements to the system made after the field test that will make it more broadly useful to other types of fluorophores and samples. 

This is the first of two articles on the Extant Life Volumetric Imaging System (ELVIS) describing a combined digital holographic microscope (DHM) and a fluorescence light-field microscope (FLFM). The instrument is modular and robust enough for field use. Each mode uses its own illumination source and camera, but both microscopes share a common objective lens and sample viewing chamber. This allows correlative volumetric imaging in amplitude, quantitative phase, and fluorescence modes. A detailed schematic and parts list is presented, as well as links to open-source software packages for data acquisition and analysis that permits interested researchers to duplicate the design. Instrument performance is quantified using test targets and beads. In the second article on ELVIS, to be published in the next issue of Microscopy Today , analysis of data from field tests and images of microorganisms will be presented.