gfp fluorescence intensity  (Olympus)

Journal: Fly

Article Title: Systematic generation of Drosophila Wnt transgenes enables the characterization of canonical Wnt signaling

doi: 10.1080/19336934.2026.2624185

Figure Lengend Snippet: Optimized pJFRC81-Insulated-GGA-Wnt expression system and validation assays. (A) Diagram illustrating modifications to the pUASattB-cDNA plasmid to generate the optimized pJFRC81-Insulated-GGA-Wnt expression construct. The original pUASattB vector contains a 5× UASt sequence (teal green) upstream of a classical cloning site replaced by the desired cDNA (white). The optimized pJFRC81-Insulated-GGA vector incorporates additional features: insulator sequences (lime green) flanking the transgene, a 10× UASt (teal green), the Syn21 translational enhancer element (grey), and an updated cloning cassette (white). (B) Fold change from qRT-PCR analysis of Wnt expression levels in third-instar larvae (N = 3–4 biological replicates) expressing tub-Gal4, tub-Gal80ts and either UAS-GFP, UAS-Wnt (-Gal4), or UAS-Wnt (+Gal4) after 48 h induction. Statistical significance was assessed using one-way ANOVA for wg, DWnt5, DWnt6, WntD, and DWnt10, and Kruskal – Wallis analysis for DWnt2 and DWnt4 (Graphpad PRISM). Significance is indicated as ns ( p > 0.05), * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. For DWnt2, DWnt4, and DWnt5, one biological replicate in the UAS-GFP control group returned an ‘Undetermined’ Ct value, which could not be included in bar graphs or statistical analysis. The absence of detectable signal in controls further supports the observed upregulation of Wnt expression upon induction. (C) GFP relative fluorescence intensity in Drosophila wing imaginal discs, quantified using Imaris (discs outlined in yellow with FIJI). Expression was compared among: (i) UAS-GFP lines carrying pJFRC81-Insulated-GGA without tub-Gal4, tub-Gal80ts induction (48 h), (ii) UAS-GFP with tub-Gal4, tub-Gal80ts induction (48 h), and (iii) UAS-GFP using pUASattB with tub-Gal4, tub-Gal80ts induction (48 h).

Article Snippet: The absence of detectable signal in controls further supports the observed upregulation of Wnt expression upon induction. (C) GFP relative fluorescence intensity in Drosophila wing imaginal discs, quantified using Imaris (discs outlined in yellow with FIJI).

Techniques: Expressing, Biomarker Discovery, Plasmid Preparation, Construct, Sequencing, Cloning, Quantitative RT-PCR, Control, Fluorescence

Journal: Synthetic and Systems Biotechnology

Article Title: AmXlnR, a transcription factor involved in xylan degradation and pentose catabolism, enhances pullulan production from xylose in Aureobasidium melanogenum

doi: 10.1016/j.synbio.2026.02.007

Figure Lengend Snippet: In vivo G FP expression analysis control of the native and mutated promoters of AmXLN2 , AmBXL2 and AmABF1 . ( a ) Schematic representation of the native promoters and their variants with deleted putative XlnR-binding sites: P XLN2 Δ:P XLN2 with 5′-GGCTGA-3′ deleted; P BXL2 Δ:P BXL2 with 5′-GGTTAA-3′ deleted; P ABF1 Δ:P ABF1 with 5′-GGCTAT-3′ deleted. ( b ) Relative fluorescence intensity, ( c ) relative GFP transcriptional level, and ( d ) bright-field and corresponding fluorescence images of the reporter strains. Three independent replicates were performed for the statistical analysis. ∗ P < 0.05,∗∗ P < 0.01, ns: no significance.

Article Snippet: GFP fluorescence intensity was visualized using an Olympus U-LH100HG fluorescent microscope and quantified using a BioTeK Synergy H1 Hybrid Reader (BioTek Instruments Inc., USA) (485 nm excitation and 520 nm emission).

Techniques: In Vivo, Expressing, Control, Binding Assay, Fluorescence

Journal: bioRxiv

Article Title: From vacuole to cytosol – Disruptive invasion triggers cytosolic release of Salmonella Paratyphi A and subsequent cytosolic motility favors evasion of xenophagy

doi: 10.1101/2022.07.24.501230

Figure Lengend Snippet: A-C . HeLa cells stably expressing LC3B-GFP (green) were pulse-chased with Dextran-Alexa647 (blue) and infected with SPA WT or SPA Δ motAB [P tetA :: motAB ] expressing mCherry (red) at MOI 30. 100 µg x ml -1 cefotaxime and ciprofloxacin were added to infected cells at 2 h p.i. if indicated. At 3-4 h p.i., randomly selected cells were imaged and later assessed for SPA associated with LC3B-positive membranes. A, B. Total counts of SPA with (dashed bars) or without (bars) LC3B decoration, compiled from three independent experiments. B. Cytosolic SPA quantified in A were further scored as motile or non-motile, and assessed for LC3B decoration. C. Percentage of SPA targeted by LC3B-containing membranes calculated from data shown in A and B . Data are means and SD of 195 infected cells with 2,450 bacteria for WT, 171 infected cells with 3,360 bacteria for Δ motAB [P tetA :: motAB ], and 215 infected cells with 1,291 bacteria for WT + cefo/cipro from three independent experiments. Statistical analysis was performed with one-way ANOVA and is indicated as n.s., not significant; *, p < 0.05; **, p < 0.01; ***, p < 0.001. D-F. Representative images showing subpopulations in SPA-infected cells. Arrowheads indicate SPA residing in SCV (blue), cytosolic SPA (red), LC3B-decorated SPA (yellow). Images are shown as single Z-plane from stack ( D ) or as maximum intensity projection ( E, F ). Scale bars: 10 µm.

Article Snippet: For correlation of maximum velocity and LC3B decoration of cytosolic SPA, individual bacteria were analyzed regarding LC3B-GFP fluorescence intensity using ZEISS Efficient Navigation (ZEN) software.

Techniques: Stable Transfection, Expressing, Infection

Journal: bioRxiv

Article Title: From vacuole to cytosol – Disruptive invasion triggers cytosolic release of Salmonella Paratyphi A and subsequent cytosolic motility favors evasion of xenophagy

doi: 10.1101/2022.07.24.501230

Figure Lengend Snippet: HeLa cells expressing LC3B-GFP (green) were pulse-chased with Dextran-Alexa647 (blue) and infected with SPA WT expressing mCherry (red) at MOI 30. Cells were fixed 4 h p.i. during LCI ( A ), and subsequently processed for transmission electron microscopy (TEM) of ultrathin sections. LCI and TEM images were superimposed for correlation ( D ) and details are shown at higher magnification ( E , F ). Arrowheads indicate SPA residing in SCV (blue), cytosolic SPA (red) and SPA targeted by LC3B-positive autophagosomal membrane (yellow). Partial enclosure by autophagosomal membranes is indicated by green arrowheads. Scale bars: 10 µm ( A , B ), 5 µm ( C , D ), 1 µm ( E ), 500 nm ( F ).

Article Snippet: For correlation of maximum velocity and LC3B decoration of cytosolic SPA, individual bacteria were analyzed regarding LC3B-GFP fluorescence intensity using ZEISS Efficient Navigation (ZEN) software.

Techniques: Expressing, Infection, Transmission Assay, Electron Microscopy

Journal: bioRxiv

Article Title: From vacuole to cytosol – Disruptive invasion triggers cytosolic release of Salmonella Paratyphi A and subsequent cytosolic motility favors evasion of xenophagy

doi: 10.1101/2022.07.24.501230

Figure Lengend Snippet: Single bacterial cells after culture in broth media ( A ), or within infected host cells ( B , C ) were registered by LCI. Subsequently, bacterial motility was analyzed by automated ( A , C ) or manual ( B ) tracking analyses. A-C. Maximum velocity of single bacterial cells from tracking SPA grown in culture ( A ), or intracellular SPA ( B ) or L.m. ( C ). For single cell analyses, tracks of 476 SPA WT from O/N culture, 7,058 SPA WT from late log culture, 2,998 SPA WT from micro-aerobic culture were collected from one experiment. For intracellular bacteria, tracks of 16,248 cytosolic SPA WT were collected from three independent experiments, and 42 tracks for L.m. from one experiment. Data are shown as Tukeýs box plots with error bars including data of 1.5 x IQR. Outliers beyond the 1.5 x IQR are shown as dots. Middle line denotes median and “+” mean value. D. Plotting of SPA-associated relative LC3B-GFP intensity against maximum velocity of individual cytosolic SPA. Representative images from measurement of fluorescence intensity are shown as single Z-plane from image stacks. Linear regression and 95% confidence bands were calculated for analyses of 63 cytosolic SPA from two independent experiments.

Article Snippet: For correlation of maximum velocity and LC3B decoration of cytosolic SPA, individual bacteria were analyzed regarding LC3B-GFP fluorescence intensity using ZEISS Efficient Navigation (ZEN) software.

Techniques: Infection, Fluorescence

Journal: bioRxiv

Article Title: From vacuole to cytosol – Disruptive invasion triggers cytosolic release of Salmonella Paratyphi A and subsequent cytosolic motility favors evasion of xenophagy

doi: 10.1101/2022.07.24.501230

Figure Lengend Snippet: HeLa LC3B-GFP cells were infected with SPA WT expressing mCherry at MOI 30. At 3-4 h p.i. cells were imaged and cytosolic SPA were identified (boxes). For single SPA cells LC3B-GFP fluorescence signal intensity was recorded, and velocity was determined by single cell tracking. Three time points of a representative time lapse series are shown, and quantitative data are displayed in . Images shown are single Z-planes from a stack. Scale bar: 10 µm.

Article Snippet: For correlation of maximum velocity and LC3B decoration of cytosolic SPA, individual bacteria were analyzed regarding LC3B-GFP fluorescence intensity using ZEISS Efficient Navigation (ZEN) software.

Techniques: Infection, Expressing, Fluorescence, Single Cell Tracking

Journal: bioRxiv

Article Title: From vacuole to cytosol – Disruptive invasion triggers cytosolic release of Salmonella Paratyphi A and subsequent cytosolic motility favors evasion of xenophagy

doi: 10.1101/2022.07.24.501230

Figure Lengend Snippet: Zipper or trigger invasion with limiting extend of membrane ruffling ( A , B ) primarily lead to an intracellular lifestyle with SCV maturation ( D ). Trigger invasion with extensive membrane ruffling and increased cooperative invasion ( C ) lead to membrane damage and rupture of the nascent SCV ( E ) and a cytosolic lifestyle of SPA. Flagellated and motile cytosolic SPA delay or escape xenophagic capture ( F ). Non-motile cytosolic SPA are captured by LC3B-positive membranes, while flagella-mediated motility permits evasion of xenophagy ( G ).

Article Snippet: For correlation of maximum velocity and LC3B decoration of cytosolic SPA, individual bacteria were analyzed regarding LC3B-GFP fluorescence intensity using ZEISS Efficient Navigation (ZEN) software.

Techniques: