Biology subjects

McFadden, C.

Publications and source records attributed to McFadden, C..

6 recordsLinked to original sources

bioRxiv2025

A deconvolution solution to aliasing confusion in apparently undersampled oblique plane microscopy

In oblique plane microscopy, a remote refocussing system is used to create an aberration-free 3D image of the object, from which an inclined plane is sampled using a tilted imaging system. As a result, unlike in conventional microscopes, the point spread function (PSF) and optical transfer function (OTF) are tilted with respect to the optical axis of the imaging system. This suggests that a small sample-scanning step size is required to satisfy the Nyquist-Shannon sampling criterion. Recently, we have shown that a careful choice of apparent undersampling leads to aliased OTF copies that can be separated and stitched together in Fourier space to form a properly sampled OTF. In our demonstrations, this led to acquisition speed gains of between twoand four-fold. Here, we introduce a deconvolution-based method to reconstruct properly sampled volumes from apparently undersampled datasets that outperforms our previous Fourier-stitching approach.

biophysics↗

bioRxiv2024

Increasing the acquisition speed in oblique plane microscopy via Aliasing

Oblique plane microcopy (OPM), a variant of light-sheet fluorescence microscopy (LSFM), enables rapid volumetric imaging without mechanically scanning the sample or an objective. In an OPM, the sample space is mapped to a distortion free image space via remote focusing, and the oblique light-sheet plane is mapped via a tilted tertiary imaging system onto a camera. As a result, the 3D point-spread function and optical transfer function are tilted to the optical axis of the tertiary imaging system. To satisfy Nyquist sampling, small scanning steps are required to encompass the tilted 3D OTF, slowing down acquisition and increasing sample exposure. Here we show that a judicious amount of under-sampling can lead to a form of aliasing in OPM that can be recovered without a loss of spatial resolution or introducing artifacts. The resulting speed gain depends on the optical parameters of the system and can reach 2-4-fold in our demonstrations. We leverage this speed gain for rapid subcellular 3D imaging of mitochondrial dynamics.

biophysics↗

bioRxiv2024

Adaptive Optics in an Oblique Plane Microscope

Adaptive optics (AO) can restore diffraction limited performance when imaging beyond superficial cell layers in vivo and in vitro, and as such is of interest for advanced 3D microscopy methods such as light-sheet fluorescence microscopy (LSFM). In a typical LSFM system, the illumination and detection paths are separate and subject to different optical aberrations. To achieve optimal microscope performance, it is necessary to sense and correct these aberrations in both light paths, resulting in a complex microscope system. Here, we show that in an oblique plane microscope (OPM), a type of LSFM with a single primary objective lens, the same deformable mirror can correct both the illumination and fluorescence detection. Besides reducing the complexity, we show that AO in OPM also restores the relative alignment of the light-sheet and focal plane, and that a projection imaging mode can stabilize and improve the wavefront correction in a sensorless AO format. We demonstrate OPM with AO on fluorescent nanospheres and by imaging the vasculature and cancer cells in zebrafish embryos embedded in a glass capillary, restoring diffraction limited resolution and improving the signal strength twofold.

cell biology↗

bioRxiv2024

navigate: an open-source platform for smart light-sheet microscopy.

navigate is a turnkey, open-source software solution designed to enhance light-sheet fluorescence microscopy (LSFM) by integrating smart microscopy techniques into a user-friendly framework. It enables automated, intelligent imaging with a Python-based control system that supports GUI-reconfigurable acquisition routines and the integration of diverse hardware sets. As a comprehensive package, navigate democratizes access to advanced LSFM capabilities, facilitating the development and implementation of smart microscopy workflows without requiring deep programming knowledge or specialized expertise in light-sheet microscopy.

biophysics↗

bioRxiv2020

An enhanced target-enrichment bait set for Hexacorallia provides phylogenomic resolution of the staghorn corals (Acroporidae) and close relatives.

The phylogenetic utility of targeted enrichment methods has been demonstrated in taxa that often have a history of single gene marker development. These genomic capture methods are now being applied to resolve evolutionary relationships from deep to shallow timescales in clades that were previously deficient in molecular marker development and lacking robust morphological characters that reflect evolutionary relationships. Effectively capturing 1000s of loci, however, in a diverse group across a broad time scale requires a bait set that incorporates multiple baits per locus. We redesigned a custom bait set for the cnidarian class Anthozoa to target 1,436 UCE loci and 1,572 exon regions within the subclass Hexacorallia. We test this redesigned bait set on 99 specimens of hard corals (Scleractinia) spanning both the "complex" (Acroporidae, Agariciidae) and "robust" (Fungiidae) clades. With focused sampling in the staghorn coral genus Acropora we explore the ability of capture data to inform the taxonomy of a clade deficient in molecular resolution. A mean of 1850 ({+/-} 298) loci were captured per taxon (955 UCEs, 894 exons). A 75% complete concatenated alignment included 1792 loci (991 UCE, 801 exons) and [~]1.87 million base pairs. Parsimony informative sites varied from 48% for alignments including all three families, to 1.5% among samples within a single Acropora species. Maximum likelihood and Bayesian analyses recover highly resolved topologies and robust molecular relationships not previously found with traditional markers within the Acroporidae. Species level relationships within the Acropora genus do not support traditional morphological groups or morphological phylogenies. Both UCE and exon datasets delineated six well-supported clades within Acropora. The enhanced bait set for Hexacorallia will allow researchers to survey the evolutionary history of important groups of reef building corals where previous molecular marker development has been unsuccessful.

evolutionary biology↗

bioRxiv2017

Universal target-enrichment baits for anthozoan (Cnidaria) phylogenomics: New approaches to long-standing problems

Anthozoans (e.g., corals, anemones) are an ecologically important and diverse group of marine metazoans that occur from shallow to deep waters worldwide. However, our understanding of the evolutionary relationships among the [~]7500 species within this class is hindered by the lack of phylogenetically informative markers that can be reliably sequenced across a diversity of taxa. We designed and tested 16,308 RNA baits to capture 720 Ultraconserved Element loci and 1,071 exon loci. Library preparation and target enrichment was performed on 33 taxa from all orders within the class Anthozoa. Following Illumina sequencing and Trinity assembly, we recovered 1,774 of 1,791 targeted loci. The mean number of loci recovered from each species was 638 {+/-} 222, with more loci recovered from octocorals (783 {+/-} 138 loci) than hexacorals (475 {+/-}187 loci). Phylogenetically informative sites ranged from 26-49% for alignments at differing hierarchical taxonomic levels (e.g., Anthozoa, Octocorallia, Hexacorallia). The percent of variable sites within each of three genera (Acropora, Alcyonium, and Sinularia) for which multiple species were sequenced ranged from 4.7-30%. Maximum likelihood analyses recovered highly resolved trees with topologies matching those supported by other studies, including the monophyly of the order Scleractinia. Our results demonstrate the utility of this target-enrichment approach to resolve phylogenetic relationships from relatively old to recent divergences. Re-designing the baits with improved affinities to capture loci within each sub-class will provide a valuable toolset to address systematic questions and further our understanding of the timing of diversifications in the class Anthozoa.

genomics↗