Journal: Poultry Science

Article Title: Duck intestinal organoids: A novel model for waterfowl parvovirus infection and immune responses investigation

doi: 10.1016/j.psj.2026.106446

Figure Lengend Snippet: Development of duck intestinal organoids. (A) Graphical representation for isolation of duck intestinal crypts. (B) Culture of intestinal organoids at 3 days; scale bar: 100 μm (Left) and 20 μm (Right) (C) Duck intestinal organoids were subjected to IFA staining for absorptive enterocytes (Villin), stem cells (Lgr5), goblet cells (MUC2) and Paneth cells (LYZ); scale bar: 20 μm.

Article Snippet: Subsequently, complete intestinal organoid growth medium (OGM; Stem Cell, Canada, 06010) supplemented with 10 μM Y-27632 (an ATP-competitive inhibitor of Rho-associated kinases; CST, USA, 72302) and 0.5% duck serum was overlaid for organoid culture and maintenance.

Techniques: Isolation, Staining

Journal: Poultry Science

Article Title: Duck intestinal organoids: A novel model for waterfowl parvovirus infection and immune responses investigation

doi: 10.1016/j.psj.2026.106446

Figure Lengend Snippet: Duck intestinal organoids monolayer is susceptible to NGPV. (A) Establishment of duck intestinal organoids monolayer; scale bar: 100 μm. (B-C) Duck intestinal organoids monolayer was inoculated with NGPV and then collected at 24, 36 and 48 h for viral load detected by RT-qPCR and viral titer measured by TCID 50 . (D) NGPV and mock-infected monolayer was stained with NGPV VP3 protein (green) and DAPI for nucleus (Blue); scale bar: 50 μm. Results are presented as mean ± SD of data from three independent experiments ***, P ≤ 0.001, determined by two-tailed Student’s t-test.

Article Snippet: Subsequently, complete intestinal organoid growth medium (OGM; Stem Cell, Canada, 06010) supplemented with 10 μM Y-27632 (an ATP-competitive inhibitor of Rho-associated kinases; CST, USA, 72302) and 0.5% duck serum was overlaid for organoid culture and maintenance.

Techniques: Quantitative RT-PCR, Infection, Staining, Two Tailed Test

Journal: Poultry Science

Article Title: Duck intestinal organoids: A novel model for waterfowl parvovirus infection and immune responses investigation

doi: 10.1016/j.psj.2026.106446

Figure Lengend Snippet: Innate immune responses are activated by NGPV in duck intestinal organoids monolayer. (A–F) Duck intestinal organoids monolayer was inoculated with NGPV and then collected at 24, 36 and 48 h. Transcription levels of TNF-α (A), IL-1β (B), IL-6 (C), IFN-α (D), IFN-β (E) and MX (F) at the indicated times post-NGPV infection were evaluated by RT-qPCR. (G-I) The protein levels of TNF-α (G), IL-6 (H) and IFN-β (I) at 36 hpi were detected by ELISA. Results are presented as mean ± SD of data from three independent experiments *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001, determined by two-tailed Student’s t-test.

Article Snippet: Subsequently, complete intestinal organoid growth medium (OGM; Stem Cell, Canada, 06010) supplemented with 10 μM Y-27632 (an ATP-competitive inhibitor of Rho-associated kinases; CST, USA, 72302) and 0.5% duck serum was overlaid for organoid culture and maintenance.

Techniques: Infection, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Two Tailed Test

Journal: Poultry Science

Article Title: Duck intestinal organoids: A novel model for waterfowl parvovirus infection and immune responses investigation

doi: 10.1016/j.psj.2026.106446

Figure Lengend Snippet: Duck apical-out intestinal organoids support NGPV replication. (A) An illustrative schematic diagram depicting the establishment of duck apical-out intestinal organoids was constructed using BioRender.com. (B) Duck apical-out organoids were stained with ZO-1 and Villin respectively; scale bar: 20 μm. (C-D) Duck apical-out organoids were infected with NGPV and then collected at 24, 36 and 48 h for viral load detected by RT-qPCR and viral titer determined by TCID 50 . Results are presented as mean ± SD of data from three independent experiments ***, P ≤ 0.001, determined by two-tailed Student’s t-test.

Article Snippet: Subsequently, complete intestinal organoid growth medium (OGM; Stem Cell, Canada, 06010) supplemented with 10 μM Y-27632 (an ATP-competitive inhibitor of Rho-associated kinases; CST, USA, 72302) and 0.5% duck serum was overlaid for organoid culture and maintenance.

Techniques: Construct, Staining, Infection, Quantitative RT-PCR, Two Tailed Test

Journal: Poultry Science

Article Title: Duck intestinal organoids: A novel model for waterfowl parvovirus infection and immune responses investigation

doi: 10.1016/j.psj.2026.106446

Figure Lengend Snippet: Antiviral responses after infection of duck apical-out intestinal organoids by NGPV. (A-F) Apical-out organoids were was infected with NGPV and then collected at 24, 36 and 48 h. Transcription levels of TNF-α (A), IL-1β (B), IL-6 (C), IFN-α (D), IFN-β (E) and MX (F) at the indicated times post-NGPV infection were evaluated by RT-qPCR. (G-I) The protein levels of TNF-α (G), IL-6 (H) and IFN-β (I) at 36 hpi were determined by ELISA. Results are presented as mean ± SD of data from three independent experiments *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001, determined by two-tailed Student’s t-test.

Article Snippet: Subsequently, complete intestinal organoid growth medium (OGM; Stem Cell, Canada, 06010) supplemented with 10 μM Y-27632 (an ATP-competitive inhibitor of Rho-associated kinases; CST, USA, 72302) and 0.5% duck serum was overlaid for organoid culture and maintenance.

Techniques: Infection, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Two Tailed Test

Journal: Cell host & microbe

Article Title: Fiber- and acetate-mediated modulation of MHC-II expression on intestinal epithelium protects from Clostridioides difficile infection

doi: 10.1016/j.chom.2024.12.017

Figure Lengend Snippet:

Article Snippet: IntestiCult Intestinal Organoid Growth Medium , STEMCELL Technologies , Cat # 06005.

Techniques: Control, Bacteria, Recombinant, Red Blood Cell Lysis, Cell Recovery, Membrane, Electron Microscopy, Staining, Enzyme-linked Immunosorbent Assay, DNA Purification, Software, Microscopy

Journal: bioRxiv

Article Title: Fluorescence Lifetime Imaging Microscopy (FLIM) visualizes internalization and biological impact of nanoplastics in live intestinal organoids

doi: 10.1101/2025.01.27.635069

Figure Lengend Snippet: A: Representative 3D confocal images of WGA-Alexa Fluor 488 and phalloidin–Texas Red (F-actin labeling) co-stained organoids with apical-out (AO), apical-basal (AB) and basal-out (BO) topology. Scale bar is 100 µm. B: Large field mosaic scan of PFA-fixed pig intestinal organoids after 18 h of polarity reversion, co-stained with fluorescent WGA and phalloidin. Left: representative images of AO, AB and BO organoids, indicated on mosaic image ( right ). Scale bar is 100 µm. C: Comparison of the yield of AO, AB and BO topology, estimated with WGA staining and F-actin labeling, respectively (AO organoids: 93.1% / 95.5%, BO organoids 0.53% / 0.55%, analyzed from 4 mosaic scanned images, see table ST3). D: Total quantification of topology, observed with WGA and Nile Red labeling for polarity reverted organoids: AO with lipid droplets (AO LDs) and AO without lipid droplets (AO no-LDs), AB and BO, quantified from mosaic scanned image. E: Lipid droplets display a characteristic distribution in AO organoids, contrasting with BO. F: Co-staining with WGA and Nile Red reveals organoid structure in apical-basal organoid (AB). Scale bar is 50 µm.

Article Snippet: Lipidure TM -CM5206 (AMS.52000034GB1G, Amsbio, UK), Matrigel growth factor-reduced (734-0269, VWR, Belgium), human intestinal organoid growth medium (STEMCELL technologies, 06010, Belgium), mouse intestinal organoid growth medium (STEMCELL technologies, 06005, Belgium), DMEM high glucose GlutaMax TM Supplement media (61965026, Gibco, Belgium), 0.5 M EDTA solution (15575020, Invitrogen, Belgium), 24-well Tissue Culture plates (734-2325, VWR, Belgium), sodium valproate (P4543, Sigma-Aldrich,Ireland), CHIR99021 (SML1046, Sigma-Aldrich,Ireland), PBS (18912-014, Gibco, Belgium).

Techniques: Labeling, Staining, Comparison

Journal: bioRxiv

Article Title: Fluorescence Lifetime Imaging Microscopy (FLIM) visualizes internalization and biological impact of nanoplastics in live intestinal organoids

doi: 10.1101/2025.01.27.635069

Figure Lengend Snippet: A: Scheme of experimental workflow. B: Representative confocal fluorescence images and corresponding phasor plots of the pig small intestinal organoid incubated with NP A-D (10 µg/ mL, 18 h), co-stained with Nile Red. NP A, B and D displayed signals on both fluorescent intensity images and phasor plots, whereas type C showed no signal, similar to control. C: Different types of uptake of NP type D uptake (10 µg/mL NPD, 20% laser power) into organoids, co-stained with WGA, with respect to the topology and size. Left: partial NP D uptake in AO, showing high pixel signal on a phasor plot. Middle: organoid size-dependent NP D uptake in AO. ROI 1: enhanced NP D pixel signal in a small AO, ROI 2: reduced NP D pixel signal in larger organoid. Right: Homogeneous NP D distribution in BO with strong signal on a phasor plot. D: Topology-dependent uptake of NP D (magenta) in apical-basal organoid (AB), co-stained with WGA (green) and Nile Red (yellow). Left: 3D reconstruction shows distinct NP D distribution in AO and BO regions. Right: representative fluorescence image of AB organoid taken from 3D reconstruction. AO and BO regions defined by WGA and Nile Red signals, with corresponding NP type D phasor FLIM plots of these regions (bottom right).

Article Snippet: Lipidure TM -CM5206 (AMS.52000034GB1G, Amsbio, UK), Matrigel growth factor-reduced (734-0269, VWR, Belgium), human intestinal organoid growth medium (STEMCELL technologies, 06010, Belgium), mouse intestinal organoid growth medium (STEMCELL technologies, 06005, Belgium), DMEM high glucose GlutaMax TM Supplement media (61965026, Gibco, Belgium), 0.5 M EDTA solution (15575020, Invitrogen, Belgium), 24-well Tissue Culture plates (734-2325, VWR, Belgium), sodium valproate (P4543, Sigma-Aldrich,Ireland), CHIR99021 (SML1046, Sigma-Aldrich,Ireland), PBS (18912-014, Gibco, Belgium).

Techniques: Fluorescence, Incubation, Staining, Control

Journal: bioRxiv

Article Title: Fluorescence Lifetime Imaging Microscopy (FLIM) visualizes internalization and biological impact of nanoplastics in live intestinal organoids

doi: 10.1101/2025.01.27.635069

Figure Lengend Snippet: A, B: Representative images of NP D uptake (10 µg/mL, 18 h) in pig intestinal organoids in respect to their apical-basal topology with high loading in both AO and BO (A) and high loading in BO and zero at AO (B). Scale bar is 50 µm. C, D: Comparison of NP D uptake in AO vs. BO organoids as a function of loading concentration (0-1 µg/mL, left panel and 1-50 µg/mL, right panel) on a widefield fluorescence microscope (intensity-based approach, C), and confocal FLIM microscope (phasor FLIM event counting approach, D). FLIM events were counted from the phasor plots reconstructed in the napari phasor plugin from the exported list of G and S coordinates. Results of one of the two independent experimental replicates are shown. Both C and D data were produced from the same samples. The box charts represent 25, median and 75 percentiles with dots corresponding to individual intensity or event count square ROI square ROI normalized values. Attribution of organoids to AO and BO-topology groups was done based on WGA staining. BO group also includes AB organoids. Statistical comparison between AO and BO groups over a range of loading concentrations was performed by Mann-Whitney test (lines represent detected statistical difference, p <0.05).

Article Snippet: Lipidure TM -CM5206 (AMS.52000034GB1G, Amsbio, UK), Matrigel growth factor-reduced (734-0269, VWR, Belgium), human intestinal organoid growth medium (STEMCELL technologies, 06010, Belgium), mouse intestinal organoid growth medium (STEMCELL technologies, 06005, Belgium), DMEM high glucose GlutaMax TM Supplement media (61965026, Gibco, Belgium), 0.5 M EDTA solution (15575020, Invitrogen, Belgium), 24-well Tissue Culture plates (734-2325, VWR, Belgium), sodium valproate (P4543, Sigma-Aldrich,Ireland), CHIR99021 (SML1046, Sigma-Aldrich,Ireland), PBS (18912-014, Gibco, Belgium).

Techniques: Comparison, Concentration Assay, Fluorescence, Microscopy, Produced, Staining, MANN-WHITNEY

Journal: bioRxiv

Article Title: Fluorescence Lifetime Imaging Microscopy (FLIM) visualizes internalization and biological impact of nanoplastics in live intestinal organoids

doi: 10.1101/2025.01.27.635069

Figure Lengend Snippet: A: Representative 3D confocal images of WGA-Alexa Fluor 488 and phalloidin–Texas Red (F-actin labeling) co-stained organoids with apical-out (AO), apical-basal (AB) and basal-out (BO) topology. Scale bar is 100 µm. B: Large field mosaic scan of PFA-fixed pig intestinal organoids after 18 h of polarity reversion, co-stained with fluorescent WGA and phalloidin. Left: representative images of AO, AB and BO organoids, indicated on mosaic image ( right ). Scale bar is 100 µm. C: Comparison of the yield of AO, AB and BO topology, estimated with WGA staining and F-actin labeling, respectively (AO organoids: 93.1% / 95.5%, BO organoids 0.53% / 0.55%, analyzed from 4 mosaic scanned images, see table ST3). D: Total quantification of topology, observed with WGA and Nile Red labeling for polarity reverted organoids: AO with lipid droplets (AO LDs) and AO without lipid droplets (AO no-LDs), AB and BO, quantified from mosaic scanned image. E: Lipid droplets display a characteristic distribution in AO organoids, contrasting with BO. F: Co-staining with WGA and Nile Red reveals organoid structure in apical-basal organoid (AB). Scale bar is 50 µm.

Article Snippet: Lipidure TM -CM5206 (AMS.52000034GB1G, Amsbio, UK), Matrigel growth factor-reduced (734-0269, VWR, Belgium), human intestinal organoid growth medium (STEMCELL technologies, 06010, Belgium), mouse intestinal organoid growth medium (STEMCELL technologies, 06005, Belgium), DMEM high glucose GlutaMax TM Supplement media (61965026, Gibco, Belgium), 0.5 M EDTA solution (15575020, Invitrogen, Belgium), 24-well Tissue Culture plates (734-2325, VWR, Belgium), sodium valproate (P4543, Sigma-Aldrich,Ireland), CHIR99021 (SML1046, Sigma-Aldrich,Ireland), PBS (18912-014, Gibco, Belgium).

Techniques: Labeling, Staining, Comparison

Journal: bioRxiv

Article Title: Fluorescence Lifetime Imaging Microscopy (FLIM) visualizes internalization and biological impact of nanoplastics in live intestinal organoids

doi: 10.1101/2025.01.27.635069

Figure Lengend Snippet: A: Scheme of experimental workflow. B: Representative confocal fluorescence images and corresponding phasor plots of the pig small intestinal organoid incubated with NP A-D (10 µg/ mL, 18 h), co-stained with Nile Red. NP A, B and D displayed signals on both fluorescent intensity images and phasor plots, whereas type C showed no signal, similar to control. C: Different types of uptake of NP type D uptake (10 µg/mL NPD, 20% laser power) into organoids, co-stained with WGA, with respect to the topology and size. Left: partial NP D uptake in AO, showing high pixel signal on a phasor plot. Middle: organoid size-dependent NP D uptake in AO. ROI 1: enhanced NP D pixel signal in a small AO, ROI 2: reduced NP D pixel signal in larger organoid. Right: Homogeneous NP D distribution in BO with strong signal on a phasor plot. D: Topology-dependent uptake of NP D (magenta) in apical-basal organoid (AB), co-stained with WGA (green) and Nile Red (yellow). Left: 3D reconstruction shows distinct NP D distribution in AO and BO regions. Right: representative fluorescence image of AB organoid taken from 3D reconstruction. AO and BO regions defined by WGA and Nile Red signals, with corresponding NP type D phasor FLIM plots of these regions (bottom right).

Article Snippet: Lipidure TM -CM5206 (AMS.52000034GB1G, Amsbio, UK), Matrigel growth factor-reduced (734-0269, VWR, Belgium), human intestinal organoid growth medium (STEMCELL technologies, 06010, Belgium), mouse intestinal organoid growth medium (STEMCELL technologies, 06005, Belgium), DMEM high glucose GlutaMax TM Supplement media (61965026, Gibco, Belgium), 0.5 M EDTA solution (15575020, Invitrogen, Belgium), 24-well Tissue Culture plates (734-2325, VWR, Belgium), sodium valproate (P4543, Sigma-Aldrich,Ireland), CHIR99021 (SML1046, Sigma-Aldrich,Ireland), PBS (18912-014, Gibco, Belgium).

Techniques: Fluorescence, Incubation, Staining, Control

Journal: bioRxiv

Article Title: Fluorescence Lifetime Imaging Microscopy (FLIM) visualizes internalization and biological impact of nanoplastics in live intestinal organoids

doi: 10.1101/2025.01.27.635069

Figure Lengend Snippet: A, B: Representative images of NP D uptake (10 µg/mL, 18 h) in pig intestinal organoids in respect to their apical-basal topology with high loading in both AO and BO (A) and high loading in BO and zero at AO (B). Scale bar is 50 µm. C, D: Comparison of NP D uptake in AO vs. BO organoids as a function of loading concentration (0-1 µg/mL, left panel and 1-50 µg/mL, right panel) on a widefield fluorescence microscope (intensity-based approach, C), and confocal FLIM microscope (phasor FLIM event counting approach, D). FLIM events were counted from the phasor plots reconstructed in the napari phasor plugin from the exported list of G and S coordinates. Results of one of the two independent experimental replicates are shown. Both C and D data were produced from the same samples. The box charts represent 25, median and 75 percentiles with dots corresponding to individual intensity or event count square ROI square ROI normalized values. Attribution of organoids to AO and BO-topology groups was done based on WGA staining. BO group also includes AB organoids. Statistical comparison between AO and BO groups over a range of loading concentrations was performed by Mann-Whitney test (lines represent detected statistical difference, p <0.05).

Article Snippet: Lipidure TM -CM5206 (AMS.52000034GB1G, Amsbio, UK), Matrigel growth factor-reduced (734-0269, VWR, Belgium), human intestinal organoid growth medium (STEMCELL technologies, 06010, Belgium), mouse intestinal organoid growth medium (STEMCELL technologies, 06005, Belgium), DMEM high glucose GlutaMax TM Supplement media (61965026, Gibco, Belgium), 0.5 M EDTA solution (15575020, Invitrogen, Belgium), 24-well Tissue Culture plates (734-2325, VWR, Belgium), sodium valproate (P4543, Sigma-Aldrich,Ireland), CHIR99021 (SML1046, Sigma-Aldrich,Ireland), PBS (18912-014, Gibco, Belgium).

Techniques: Comparison, Concentration Assay, Fluorescence, Microscopy, Produced, Staining, MANN-WHITNEY