Journal: Microbiology Spectrum

Article Title: A novel multiplex fluorescent-labeling method for the visualization of mixed-species biofilms in vitro

doi: 10.1128/spectrum.00253-24

Figure Lengend Snippet: Effects of CellTrace labeling on bacteria. ( A ) FC analysis of bacteria labeled with CellTrace. Gram-positive S. gordonii and Gram-negative P. gingivalis were labeled for 1 hour with CellTrace™ CFSE (green) and analyzed using FC. Graphs show CFSE staining expressed as the relative fluorescence intensity ( X -axis) and the relative number of events (counts, Y -axis). The data show three experiments carried out using independent biological replicates and a single population of unstained cells was used as a control. ( B ) Growth curve analysis of CellTrace CFSE-labeled planktonic cultures. Cells were pre-labeled for 1 hour, and growth was measured as optical density. Unlabeled cells were used for comparison. Graphs show mean ± SD of three independent biological replicates (except for P. gingivalis and P. micra where n = 2). Bar charts (inserts) show mean ± SD of CFU counts at experiment start and a later stage of growth for each bacterial species. Labeled and unlabeled cells are shown in green and black, respectively. ( C ) Fluorescence intensity of CellTrace™ CFSE-labeled planktonic cultures. Graphs showing mean ± SD of MFI values over time measured using FC. Two representative Gram positive ( S. gordonii and S. mutans ) and Gram negative ( P. gingivalis and V. parvula ) bacteria are shown.

Article Snippet: Histograms were then used to determine the median fluorescence intensity (MFI) of the whole population and the percentage of CellTrace CFSE-labeled cells within the population using BD Accuri C6Plus software.

Techniques: Labeling, Bacteria, Staining, Fluorescence, Control, Comparison

Journal: Microbiology Spectrum

Article Title: A novel multiplex fluorescent-labeling method for the visualization of mixed-species biofilms in vitro

doi: 10.1128/spectrum.00253-24

Figure Lengend Snippet: CDSM visualization of SS biofilms labeled with CellTrace. Representative CDSM images of SS biofilms related to oral disease models. Biofilms were formed overnight using unlabeled or CellTrace (far red, yellow, CFSE, or violet)-labeled cells. Unlabeled biofilms were post-stained with CellTrace CFSE for comparison. The periodontitis model is represented by S. gordonii, A. odontolyticus, P. gingivalis, and P. micra, whereas the dental caries model is comprised of S. mutans , V. parvula, L. paracasei, and A. naeslundii . The scale bar (10 µm) applies to all panels.

Article Snippet: Histograms were then used to determine the median fluorescence intensity (MFI) of the whole population and the percentage of CellTrace CFSE-labeled cells within the population using BD Accuri C6Plus software.

Techniques: Labeling, Staining, Comparison

Journal: Microbiology Spectrum

Article Title: A novel multiplex fluorescent-labeling method for the visualization of mixed-species biofilms in vitro

doi: 10.1128/spectrum.00253-24

Figure Lengend Snippet: Comparison of surface coverage of SS biofilms labeled with CellTrace CFSE pre- or post-biofilm development ^a

Article Snippet: Histograms were then used to determine the median fluorescence intensity (MFI) of the whole population and the percentage of CellTrace CFSE-labeled cells within the population using BD Accuri C6Plus software.

Techniques: Comparison, Labeling

Journal: Microbiology Spectrum

Article Title: A novel multiplex fluorescent-labeling method for the visualization of mixed-species biofilms in vitro

doi: 10.1128/spectrum.00253-24

Figure Lengend Snippet: Examination of biofilm structure and cell detachment from SS biofilms over time. Gram-positive ( S. gordonii and A. odontolyticus ) and Gram-negative ( P. gingivalis and V. parvula ) bacteria labeled with CellTrace CFSE (green) were used to create SS biofilms. Graphs (left panels) show percentages of CFSE-stained cells present in the starting inocula and biofilm supernatants. Supernatants were extracted and analyzed with FC on days 1, 2, and 3. The right panels show representative Z-plane CDSM images of SS biofilms after 4 hours (day 0) and 1, 2, and 3 days.

Article Snippet: Histograms were then used to determine the median fluorescence intensity (MFI) of the whole population and the percentage of CellTrace CFSE-labeled cells within the population using BD Accuri C6Plus software.

Techniques: Bacteria, Labeling, Staining

Journal: Microbiology Spectrum

Article Title: A novel multiplex fluorescent-labeling method for the visualization of mixed-species biofilms in vitro

doi: 10.1128/spectrum.00253-24

Figure Lengend Snippet: Assessment of CellTrace staining as a marker of bacterial viability in biofilms representative CDSM and CLSM images of overnight SS biofilms stained with CellTrace CFSE (green) or BacLight LIVE/DEAD stain. Biofilms of S. gordonii and P. gingivalis were maintained in PRNM (control) or treated with 70% ethanol or H 2 O 2 for 1 hour. The scale bar (10 µm) applies to all panels.

Article Snippet: Histograms were then used to determine the median fluorescence intensity (MFI) of the whole population and the percentage of CellTrace CFSE-labeled cells within the population using BD Accuri C6Plus software.

Techniques: Staining, Marker, Control

Journal: Microbiology Spectrum

Article Title: A novel multiplex fluorescent-labeling method for the visualization of mixed-species biofilms in vitro

doi: 10.1128/spectrum.00253-24

Figure Lengend Snippet: Visualization of MS biofilms representing a periodontitis model labeled by multiplexing CellTrace dyes. Representative CDSM Z-stack (left) and single-plane (right) images of overnight MS biofilms stained with CellTrace dyes. A unique CellTrace color was used to label four species ( P. gingivalis [red], S. gordonii [yellow], P. micra [green], and A. odontolyticus [violet]). Both multiplex (4-color compilation) and the color-segmented images are shown. The percentage coverage for each species within the biofilm models was calculated using a region of interest (ROI) within the delineated area in the single-plane image. Z-stack images were used to provide information regarding bacterial spatial location and relationships within the overall biofilm.

Article Snippet: Histograms were then used to determine the median fluorescence intensity (MFI) of the whole population and the percentage of CellTrace CFSE-labeled cells within the population using BD Accuri C6Plus software.

Techniques: Labeling, Multiplexing, Staining, Multiplex Assay

Journal: Microbiology Spectrum

Article Title: A novel multiplex fluorescent-labeling method for the visualization of mixed-species biofilms in vitro

doi: 10.1128/spectrum.00253-24

Figure Lengend Snippet: Visualization of MS biofilms representing a dental caries model labeled by multiplexing CellTrace dyes. Representative CDSM Z-stack (left) and single-plane (right) images of overnight MS biofilms stained with CellTrace dyes. A unique CellTrace color was used to label four species ( V. parvula [red], S. mutans [yellow], A. naeslundii [green], and L. paracasei [violet]). Both multiplex (4-color compilation) and the color segmented images are shown. The percentage coverage for each species within the biofilm models was calculated using a region of interest (ROI) within the delineated area in the single-plane image. Z-stack images were used to provide information regarding bacterial spatial location and relationships within the overall biofilm.

Article Snippet: Histograms were then used to determine the median fluorescence intensity (MFI) of the whole population and the percentage of CellTrace CFSE-labeled cells within the population using BD Accuri C6Plus software.

Techniques: Labeling, Multiplexing, Staining, Multiplex Assay

Journal: Microbiology Spectrum

Article Title: A novel multiplex fluorescent-labeling method for the visualization of mixed-species biofilms in vitro

doi: 10.1128/spectrum.00253-24

Figure Lengend Snippet: Assessment of CellTrace durability in MS biofilms. Representative CSDM and CLSM images of MS biofilms. Biofilms were created using the periodontitis model ( P. gingivalis [red], S. gordonii [yellow], P. micra [green], and A. odontolyticus [violet]) and visualized following overnight growth (day 1) and after 4 days through CDSM Z-stack images. Parallel, non-labeled biofilms were also grown, stained with BacLight LIVE/DEAD stain on day 4 and images using CLSM to assess viability. The scale bar (10 µm) applies to all panels.

Article Snippet: Histograms were then used to determine the median fluorescence intensity (MFI) of the whole population and the percentage of CellTrace CFSE-labeled cells within the population using BD Accuri C6Plus software.

Techniques: Labeling, Staining

Journal: Frontiers in Microbiology

Article Title: Phenotype-Independent Isolation of Interspecies Saccharomyces Hybrids by Dual-Dye Fluorescent Staining and Fluorescence-Activated Cell Sorting

doi: 10.3389/fmicb.2019.00871

Figure Lengend Snippet: Intra-species mating of S. cerevisiae strains CEN.PK113-5A ( MATa URA3 his3 -Δ 1 leu2-3,112 trp1-289 ) and IMK439 ( MATα HIS3 TRP1 LEU2 ura3 Δ ::KanMX ). (A) Fluorescence contour plots of unstained CEN.PK113-5A, CEN.PK113-5A stained with CellTrace TM CFSE, IMK439 stained with CellTrace TM Violet, and of the mating culture after 18, 24, and 42 h. The indicated gated areas were used for sorting cells, event rates of each gate are indicated as a percentage of total cell counts. (B) Microscope image (400×) of zygotes sorted from the double-stained population (C+V+) after 42 h of mating. (C) Percentage of cells able to grow on synthetic medium without auxotrophy-complementing supplements in different populations sorted by FACS, as indicated in (A) .

Article Snippet: Fluorescence from CellTrace TM CFSE was imaged through a GFP filter set (Carl Zeiss AG) containing a 470 nm bandpass excitation filter with a bandwidth of 20 and 540 nm emission filter with a bandwidth of 25 nm.

Techniques: Fluorescence, Staining, Microscopy

Journal: Cell reports

Article Title: Regulation of Intronic Polyadenylation by PCF11 Impacts mRNA Expression of Long Genes

doi: 10.1016/j.celrep.2019.02.049

Figure Lengend Snippet:

Article Snippet: After 40~48 hours, cells were analyzed in a BD FACScalibur system (BD Biosciences) with an excitation filter of 488 nm and emission filter of 530 nm for CellTrace CFSE.

Techniques: Recombinant, Modification, Magnetic Beads, Flow Cytometry, Generated, Plasmid Preparation, Clone Assay, Software, Expressing