Biology subjects

LeBlanc, S. J.

Publications and source records attributed to LeBlanc, S. J..

3 recordsLinked to original sources

bioRxiv2026

Expression of photoactivatable molecules enables FCS in live cells by controlling fluorescence intensity

Cellular systems must act robustly to maintain organismal health, including maintaining biophysical properties that allow for appropriate cellular function, and adapting these properties through changes such as those that occur during cell division. However, we still lack tools to measure many of these physical properties with precision in the living cell. For example, the mechanical properties of the nucleoplasm, the fluid-like substance that fills the nucleus, have not been fully characterized. To investigate these properties, we have turned to the fission yeast Schizosaccharomyces pombe (S. pombe), a well-established, genetically tractable model organism that has been used extensively for studying a variety of cell biophysical processes and structures, including the cytoskeleton and cell division. It is an apt system for studying how the nucleus adapts over the course of the cell cycle, since it undergoes closed mitosis, where the nuclear envelope remains intact during cell division. Studying nucleoplasm properties over the course of closed mitosis may help reveal how nuclear volume, shape, surface area expansion, and chromosome segregation are linked and coordinated. To measure nucleoplasm material properties in S. pombe, we have paired Fluorescence Correlation Spectroscopy (FCS) with a photoswitchable fluorophore, enabling fine control over fluorescent intensity inside live cells. We infer material properties from FCS measurements, while the photoswitchable probe enables confocal imaging in conjunction with these measurements, yielding corresponding information about cellular state and dynamics. Interestingly, we find that nucleoplasm material properties do not vary significantly over the cell cycle. Future studies will use this tool to examine how diverse molecular and genetic perturbations alter nucleoplasmic properties, providing insight into how these properties maintain nuclear function and protect genomic integrity over the cell cycle and during development.

biophysics↗

bioRxiv2024

A practical guide to time-resolved fluorescence microscopy and spectroscopy

Time-correlated single photon counting (TCSPC) coupled with confocal microscopy is a versatile biophysical tool that enables real-time monitoring of biomolecular dynamics across many timescales. With TCSPC, Fluorescence correlation spectroscopy (FCS) and pulsed interleaved excitation-Forster resonance energy transfer (PIE-FRET) are collected simultaneously on diffusing molecules to extract diffusion characteristics and proximity information. This article is a guide to calibrating FCS and PIE-FRET measurements with several biological samples including liposomes, streptavidin-coated quantum dots, proteins, and nucleic acids for reliable determination of diffusion coefficients and FRET efficiency. The FRET efficiency results are also compared to surface-attached single molecules using fluorescence lifetime imaging microscopy (FLIM-FRET). Combining the methods is a powerful approach to revealing mechanistic details of biological processes and pathways.

biophysics↗

bioRxiv2020

Effects of parity, blood progesterone, and non-steroidal anti-inflammatory treatment on the dynamics of the uterine microbiota of healthy postpartum dairy cows

This study evaluated the effects of treatment with meloxicam (a non-steroidal anti-inflammatory drug), parity, and blood progesterone concentration on the dynamics of the uterine microbiome of clinically healthy postpartum dairy cows. Seven primiparous and 9 multiparous postpartum Holstein cows received meloxicam (0.5 mg/kg SC, n = 7 cows) once daily for 4 days (10 to 13 days in milk (DIM)) or were untreated (n = 9 cows). Endometrial cytology samples were collected by cytobrush at 10, 21, and 35 DIM, from which the metagenomic analysis was done using 16S rRNA gene sequence analysis. A radioimmunoassay was used to measure progesterone concentration in blood serum samples at 35 DIM and cows were classified as > 1 ng/mL (n = 10) or [≤] 1 ng/mL (n = 6). Alpha diversity for bacterial genera (Chao1, Shannon-Weiner, and Camargos evenness indices) were not affected by DIM, meloxicam treatment, parity, or progesterone category (P > 0.2). For beta diversity (genera level), principal coordinate analysis (Bray-Curtis) showed differences in microbiome between parity groups (P = 0.01). There was lower overall abundance of Anaerococcus, Bifidobacterium, Corynebacterium, Lactobacillus, Paracoccus, Staphylococcus, and Streptococcus and higher abundance of Bacillus, Fusobacterium, and Novosphingobium in primiparous than multiparous cows (P < 0.05); these patterns were consistent across sampling days. Bray-Curtis dissimilarity did not differ by DIM at sampling, meloxicam treatment, or progesterone category at 35 DIM (P > 0.5). In conclusion, uterine bacterial composition was not different at 10, 21, or 35 DIM, and meloxicam treatment or progesterone category did not affect uterine microbiota in clinically healthy postpartum dairy cows. Primiparous cows presented a different composition of uterine bacteria than multiparous cows. The differences in microbiome associated with parity might be attributable to changes that occur consequent to the first calving, but this hypothesis should be investigated further.

microbiology↗