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biotinylated wisteria floribunda agglutinin  (Vector Laboratories)


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    Vector Laboratories biotinylated wisteria floribunda agglutinin
    Neonatal sevoflurane exposure induces PNNs damage in the hippocampus. (A) Schematic of the experimental protocol for hippocampal extracellular matrix (ECM) analyses. (B) Representative scanning electron micrographs (SEM) of hippocampal ECM ultrastructure in Con and Sev ‑treated groups. Scale bars = 5 µm (5K×), 1.2 µm (20K×), and 500 nm (50K×). (C) Quantification of mean gray value (left) and fiber diameter (right) of hippocampal ECM in Con and Sev groups. (D, E) Wisteria <t>floribunda</t> <t>agglutinin</t> (WFA) staining and quantitative analysis of PNN intensity in the CA2 region at 7, 14, 30, and 60 days post‐exposure (dpe). Scale bar = 50 µm. (F, G) Western blotting and quantification of hapln1 and aggrecan protein levels in the hippocampus of Con and Sev groups. (H, I) Immunofluorescence staining and quantitative analysis of aggrecan in the hippocampus of Con and Sev groups at 60 dpe. Green: aggrecan. Blue: DAPI. Scale bar = 50 µm. (J–M) PNNs in the Con group exhibit bright, well‐organized mesh‐like architecture, whereas PNNs in the Sev group display reduced WFA intensity and a disrupted staining pattern. (J) Representative WFA staining. Purple: WFA. Scale bar = 10 µm. (K) Binary images of the regions in (J). (L) Quantification of black/white pixel ratio. (M) Representative surface intensity profiles of WFA‐positive PNNs. (N) Immunoelectron microscopy of PNNs in the Con and Sev groups. High‐magnification images (right panels) display immunogold labeling within WFA (red arrows) in both groups. Scale bars = 1 µm (left), 500 nm (right). Data represent the mean ± SEM. n = 3 mice per group for SEM (B, C, J–N); n = 6 mice per group for all other experiments. Statistical comparisons were conducted using unpaired Student's t‐test. ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ P < 0.0001.
    Biotinylated Wisteria Floribunda Agglutinin, supplied by Vector Laboratories, used in various techniques. Bioz Stars score: 96/100, based on 355 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/biotinylated+lectins/Biotinylated+Wisteria+Floribunda+Lectin+(WFA%2C+WFL)/pmc13624421-299-26-31
    Average 96 stars, based on 355 article reviews
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    Images

    1) Product Images from "Neonatal Sevoflurane Exposure Induces Long‐Term Cognitive Impairment via Epigenetically Mediated MMP9 Activation and Perineuronal Net Disruption in the Hippocampal CA2"

    Article Title: Neonatal Sevoflurane Exposure Induces Long‐Term Cognitive Impairment via Epigenetically Mediated MMP9 Activation and Perineuronal Net Disruption in the Hippocampal CA2

    Journal: Advanced Science

    doi: 10.1002/advs.78006

    Neonatal sevoflurane exposure induces PNNs damage in the hippocampus. (A) Schematic of the experimental protocol for hippocampal extracellular matrix (ECM) analyses. (B) Representative scanning electron micrographs (SEM) of hippocampal ECM ultrastructure in Con and Sev ‑treated groups. Scale bars = 5 µm (5K×), 1.2 µm (20K×), and 500 nm (50K×). (C) Quantification of mean gray value (left) and fiber diameter (right) of hippocampal ECM in Con and Sev groups. (D, E) Wisteria floribunda agglutinin (WFA) staining and quantitative analysis of PNN intensity in the CA2 region at 7, 14, 30, and 60 days post‐exposure (dpe). Scale bar = 50 µm. (F, G) Western blotting and quantification of hapln1 and aggrecan protein levels in the hippocampus of Con and Sev groups. (H, I) Immunofluorescence staining and quantitative analysis of aggrecan in the hippocampus of Con and Sev groups at 60 dpe. Green: aggrecan. Blue: DAPI. Scale bar = 50 µm. (J–M) PNNs in the Con group exhibit bright, well‐organized mesh‐like architecture, whereas PNNs in the Sev group display reduced WFA intensity and a disrupted staining pattern. (J) Representative WFA staining. Purple: WFA. Scale bar = 10 µm. (K) Binary images of the regions in (J). (L) Quantification of black/white pixel ratio. (M) Representative surface intensity profiles of WFA‐positive PNNs. (N) Immunoelectron microscopy of PNNs in the Con and Sev groups. High‐magnification images (right panels) display immunogold labeling within WFA (red arrows) in both groups. Scale bars = 1 µm (left), 500 nm (right). Data represent the mean ± SEM. n = 3 mice per group for SEM (B, C, J–N); n = 6 mice per group for all other experiments. Statistical comparisons were conducted using unpaired Student's t‐test. ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ P < 0.0001.
    Figure Legend Snippet: Neonatal sevoflurane exposure induces PNNs damage in the hippocampus. (A) Schematic of the experimental protocol for hippocampal extracellular matrix (ECM) analyses. (B) Representative scanning electron micrographs (SEM) of hippocampal ECM ultrastructure in Con and Sev ‑treated groups. Scale bars = 5 µm (5K×), 1.2 µm (20K×), and 500 nm (50K×). (C) Quantification of mean gray value (left) and fiber diameter (right) of hippocampal ECM in Con and Sev groups. (D, E) Wisteria floribunda agglutinin (WFA) staining and quantitative analysis of PNN intensity in the CA2 region at 7, 14, 30, and 60 days post‐exposure (dpe). Scale bar = 50 µm. (F, G) Western blotting and quantification of hapln1 and aggrecan protein levels in the hippocampus of Con and Sev groups. (H, I) Immunofluorescence staining and quantitative analysis of aggrecan in the hippocampus of Con and Sev groups at 60 dpe. Green: aggrecan. Blue: DAPI. Scale bar = 50 µm. (J–M) PNNs in the Con group exhibit bright, well‐organized mesh‐like architecture, whereas PNNs in the Sev group display reduced WFA intensity and a disrupted staining pattern. (J) Representative WFA staining. Purple: WFA. Scale bar = 10 µm. (K) Binary images of the regions in (J). (L) Quantification of black/white pixel ratio. (M) Representative surface intensity profiles of WFA‐positive PNNs. (N) Immunoelectron microscopy of PNNs in the Con and Sev groups. High‐magnification images (right panels) display immunogold labeling within WFA (red arrows) in both groups. Scale bars = 1 µm (left), 500 nm (right). Data represent the mean ± SEM. n = 3 mice per group for SEM (B, C, J–N); n = 6 mice per group for all other experiments. Statistical comparisons were conducted using unpaired Student's t‐test. ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ P < 0.0001.

    Techniques Used: Staining, Analysis, Western Blot, Immunofluorescence, Immuno-Electron Microscopy, Labeling

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    Article Snippet: Biotinylated lectins, including Aleuria aurantia lectin (AAL, 1:5000, Vector Laboratories), Lens culinaris agglutinin (LCA, 1:5000, Vector Laboratories), Sambucus nigra agglutinin (SNA, 1:10000, Vector Laboratories), Maackia amurensis lectin (MAL, 1:3000, Vector Laboratories), Phaseolus vulgaris Erythroagglutinin (PHA-E, 1:3000, Vector Laboratories), Phaseolus vulgaris Leucoagglutinin (PHA-L, 1:3000, Vector Laboratories) and Concanavalin A (ConA, 1:10000, Vector Laboratories) were purchased from Vector Laboratories Inc. (Burlingame, CA, USA) AR TIC LE IN PR ES S (Table 1).

    Article Title: Abnormal glycosylation changes in brain tissue of kainic acid-induced epileptic rats
    Article Snippet: Biotinylated lectins, including Aleuria aurantia lectin (AAL, 1:5000, Vector Laboratories), Lens culinaris agglutinin (LCA, 1:5000, Vector Laboratories), Sambucus nigra agglutinin (SNA, 1:10000, Vector Laboratories), Maackia amurensis lectin (MAL, 1:3000, Vector Laboratories), Phaseolus vulgaris Erythroagglutinin (PHA-E, 1:3000, Vector Laboratories), Phaseolus vulgaris Leucoagglutinin (PHA-L, 1:3000, Vector Laboratories) and Concanavalin A (ConA, 1:10000, Vector Laboratories) were purchased from Vector Laboratories Inc. (Burlingame, CA, USA) (Table ).

    Staining:

    Article Title: MGAT1 knockout in human dendritic cells enhance CD8 + T cell activation
    Article Snippet: .. Briefly, cells were stained with LIVE/DEADTM Fixable Yellow Dead Cell Stain to assess viability, before incubation with a selection of biotinylated lectins from Vector Laboratories: Aleuria aurantia lectin (AAL), Maackia amurensis lectin I and II (MAL-I and MAL-II), Sambucus nigra agglutinin (SNA), Phaseolus vulgaris lectin L (PHA-L) and Peanut agglutinin (PNA). .. Detection was achieved using AF488-conjugated streptavidin (Invitrogen), and both lectins and streptavidin were incubated with the cells on ice for 20 min.

    Article Title: Evidence of an allostatic response by intestinal tissues following induction of joint inflammation
    Article Snippet: Sagittal sections from knee joints were stained with Hematoxylin/Eosin, anti-CD68 (Abcam, Ab283654 ; 1:100) or anti-CD4 Abcam, Ab237722 1:2000). .. Sections of intestinal Swiss rolls were stained using the Alcian Blue Kit (Vector Labs, H-3501), anti-CD68 (1:100), anti-CD4 (1:2000), anti-CD8 (Abcam, Ab237709 , 1:500) or anti-Ki67 (Invitrogen, MA5–14520; 1:100), Invitrogen, MA5–14520) or the following biotinylated lectins (all from Vector Labs), Griffonia Bandeiraea Simplidifolia Lectin I, (GSL I, B-1105–2), Maackia Amurensis Lectin II (MAL II, B-1265–1) and Ulex Europaeus Agglutinin I (UEA, B-1065–2). .. Prior to histochemical staining, antigen retrieval was performed using Tris/EDTA (Abcam, ab93684), and sections were blocked with normal goat serum (Vector Laboratories, S-1000).

    Article Title: Transgenic chicken that expresses a fluorescent protein and SIAT1
    Article Snippet: .. Cells at 80-90% confluence were incubated with biotinylated lectins (Vector Laboratories, MAL II lectin for staining α2,3 residues, SNA lectin for staining α2,6 residues), then incubated with streptavidin-phycoerythrin conjugate, and then fixed in 4% paraformaldehyde for imaging under fluorescence. ..

    Incubation:

    Article Title: MGAT1 knockout in human dendritic cells enhance CD8 + T cell activation
    Article Snippet: .. Briefly, cells were stained with LIVE/DEADTM Fixable Yellow Dead Cell Stain to assess viability, before incubation with a selection of biotinylated lectins from Vector Laboratories: Aleuria aurantia lectin (AAL), Maackia amurensis lectin I and II (MAL-I and MAL-II), Sambucus nigra agglutinin (SNA), Phaseolus vulgaris lectin L (PHA-L) and Peanut agglutinin (PNA). .. Detection was achieved using AF488-conjugated streptavidin (Invitrogen), and both lectins and streptavidin were incubated with the cells on ice for 20 min.

    Article Title: Transgenic chicken that expresses a fluorescent protein and SIAT1
    Article Snippet: .. Cells at 80-90% confluence were incubated with biotinylated lectins (Vector Laboratories, MAL II lectin for staining α2,3 residues, SNA lectin for staining α2,6 residues), then incubated with streptavidin-phycoerythrin conjugate, and then fixed in 4% paraformaldehyde for imaging under fluorescence. ..

    Selection:

    Article Title: MGAT1 knockout in human dendritic cells enhance CD8 + T cell activation
    Article Snippet: .. Briefly, cells were stained with LIVE/DEADTM Fixable Yellow Dead Cell Stain to assess viability, before incubation with a selection of biotinylated lectins from Vector Laboratories: Aleuria aurantia lectin (AAL), Maackia amurensis lectin I and II (MAL-I and MAL-II), Sambucus nigra agglutinin (SNA), Phaseolus vulgaris lectin L (PHA-L) and Peanut agglutinin (PNA). .. Detection was achieved using AF488-conjugated streptavidin (Invitrogen), and both lectins and streptavidin were incubated with the cells on ice for 20 min.

    Microscopy:

    Article Title: Method of isolating and detecting cancer stem cells
    Article Snippet: .. Equipment used: Paraffin blocks, Ice, Microtome, Microtome, Superfrost® blades, Bond Max Automate (Leica Microsystems) with computer, Leica consumables (alcohol, wash buffer, ER1 buffer, dewax buffer, labels, coverslips, tubes), PBS-BSA buffer 5%, Biotinylated Lectins (UEA-1, Jacaline, ABA, ACA, GSL-I and GSL-II) (Vector Lab), Kit Bond Intense R detection (Leica), Leica mounting medium, slides and microscope. ..

    Imaging:

    Article Title: Transgenic chicken that expresses a fluorescent protein and SIAT1
    Article Snippet: .. Cells at 80-90% confluence were incubated with biotinylated lectins (Vector Laboratories, MAL II lectin for staining α2,3 residues, SNA lectin for staining α2,6 residues), then incubated with streptavidin-phycoerythrin conjugate, and then fixed in 4% paraformaldehyde for imaging under fluorescence. ..

    Fluorescence:

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    Vector Laboratories anti α 2 6sa
    Restoration of influenza virus susceptibility in ST6GAL1 −/− / ST3GAL4 −/− Landrace pig cells through complementation. A: Quantitative reverse transcription-PCR (RT-qPCR) analysis of ST6GAL1 and ST3GAL4 gene expression levels in wild-type (WT), knockout (KO), and complemented (KO + ST) cells. B: Immunofluorescent detection of α2,6-linked and α2,3-linked sialylated glycans in wild-type (WT), knockout (KO), and complemented (KO + ST) Landrace pig cells. Red indicates SNA (α2,6) or <t>MAL</t> Ⅱ (α2,3) staining, and blue indicates nuclei. The KO group showed nearly undetectable signals for both SNA and MAL Ⅱ. In contrast, the WT and KO + ST groups exhibited strong fluorescence for both <t>lectins,</t> indicating successful restoration of both α2,6- and α2,3-linked sialylated glycans in complemented cells. Scale bars, 100 μm. C: Cytopathic effects induced by H1N1 influenza virus infection at 0, 24, and 48 h post-infection (HPI). Scale bars, 500 μm. D: Immunofluorescent detection of IAV nucleoprotein (NP) in cells post-infection with H1N1 influenza virus at 48 HPI. NP-Alexa Fluor 488, green; DAPI, blue. Scale bars, 100 μm. E: RT-qPCR analysis of influenza viral M gene expression levels. F: Plaque assay measurement of infectious IAV (H1N1) virus titers (lg PFU/mL). Data are presented as mean ± standard deviation (A, E, and F); n = 3 independent cell culture experiments conducted on separate days with identical cell batches. Statistical significance was determined by one-way ANOVA followed by Tukey's multiple comparisons test. * P < 0.05, ** P < 0.01, and *** P < 0.001.
    Anti α 2 6sa, supplied by Vector Laboratories, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/biotinylated+lectins/Biotinylated+Sambucus+Nigra+Lectin+(SNA%2C+EBL)/10__1016_slash_j__gendis__2026__102499-79-58-59
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    biotinylated vicia villosa lectin (vvl, vva)  (Vector Laboratories)
    95
    Vector Laboratories biotinylated vicia villosa lectin (vvl, vva)
    Restoration of influenza virus susceptibility in ST6GAL1 −/− / ST3GAL4 −/− Landrace pig cells through complementation. A: Quantitative reverse transcription-PCR (RT-qPCR) analysis of ST6GAL1 and ST3GAL4 gene expression levels in wild-type (WT), knockout (KO), and complemented (KO + ST) cells. B: Immunofluorescent detection of α2,6-linked and α2,3-linked sialylated glycans in wild-type (WT), knockout (KO), and complemented (KO + ST) Landrace pig cells. Red indicates SNA (α2,6) or <t>MAL</t> Ⅱ (α2,3) staining, and blue indicates nuclei. The KO group showed nearly undetectable signals for both SNA and MAL Ⅱ. In contrast, the WT and KO + ST groups exhibited strong fluorescence for both <t>lectins,</t> indicating successful restoration of both α2,6- and α2,3-linked sialylated glycans in complemented cells. Scale bars, 100 μm. C: Cytopathic effects induced by H1N1 influenza virus infection at 0, 24, and 48 h post-infection (HPI). Scale bars, 500 μm. D: Immunofluorescent detection of IAV nucleoprotein (NP) in cells post-infection with H1N1 influenza virus at 48 HPI. NP-Alexa Fluor 488, green; DAPI, blue. Scale bars, 100 μm. E: RT-qPCR analysis of influenza viral M gene expression levels. F: Plaque assay measurement of infectious IAV (H1N1) virus titers (lg PFU/mL). Data are presented as mean ± standard deviation (A, E, and F); n = 3 independent cell culture experiments conducted on separate days with identical cell batches. Statistical significance was determined by one-way ANOVA followed by Tukey's multiple comparisons test. * P < 0.05, ** P < 0.01, and *** P < 0.001.
    Biotinylated Vicia Villosa Lectin (Vvl, Vva), supplied by Vector Laboratories, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    marker isolectin b4  (Vector Laboratories)
    95
    Vector Laboratories marker isolectin b4
    Restoration of influenza virus susceptibility in ST6GAL1 −/− / ST3GAL4 −/− Landrace pig cells through complementation. A: Quantitative reverse transcription-PCR (RT-qPCR) analysis of ST6GAL1 and ST3GAL4 gene expression levels in wild-type (WT), knockout (KO), and complemented (KO + ST) cells. B: Immunofluorescent detection of α2,6-linked and α2,3-linked sialylated glycans in wild-type (WT), knockout (KO), and complemented (KO + ST) Landrace pig cells. Red indicates SNA (α2,6) or <t>MAL</t> Ⅱ (α2,3) staining, and blue indicates nuclei. The KO group showed nearly undetectable signals for both SNA and MAL Ⅱ. In contrast, the WT and KO + ST groups exhibited strong fluorescence for both <t>lectins,</t> indicating successful restoration of both α2,6- and α2,3-linked sialylated glycans in complemented cells. Scale bars, 100 μm. C: Cytopathic effects induced by H1N1 influenza virus infection at 0, 24, and 48 h post-infection (HPI). Scale bars, 500 μm. D: Immunofluorescent detection of IAV nucleoprotein (NP) in cells post-infection with H1N1 influenza virus at 48 HPI. NP-Alexa Fluor 488, green; DAPI, blue. Scale bars, 100 μm. E: RT-qPCR analysis of influenza viral M gene expression levels. F: Plaque assay measurement of infectious IAV (H1N1) virus titers (lg PFU/mL). Data are presented as mean ± standard deviation (A, E, and F); n = 3 independent cell culture experiments conducted on separate days with identical cell batches. Statistical significance was determined by one-way ANOVA followed by Tukey's multiple comparisons test. * P < 0.05, ** P < 0.01, and *** P < 0.001.
    Marker Isolectin B4, supplied by Vector Laboratories, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/biotinylated+lectins/Biotinylated+Griffonia+Simplicifolia+Lectin+I+(GSL+I)+isolectin+B4/pmc13133793-44-48-54
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    Image Search Results


    Neonatal sevoflurane exposure induces PNNs damage in the hippocampus. (A) Schematic of the experimental protocol for hippocampal extracellular matrix (ECM) analyses. (B) Representative scanning electron micrographs (SEM) of hippocampal ECM ultrastructure in Con and Sev ‑treated groups. Scale bars = 5 µm (5K×), 1.2 µm (20K×), and 500 nm (50K×). (C) Quantification of mean gray value (left) and fiber diameter (right) of hippocampal ECM in Con and Sev groups. (D, E) Wisteria floribunda agglutinin (WFA) staining and quantitative analysis of PNN intensity in the CA2 region at 7, 14, 30, and 60 days post‐exposure (dpe). Scale bar = 50 µm. (F, G) Western blotting and quantification of hapln1 and aggrecan protein levels in the hippocampus of Con and Sev groups. (H, I) Immunofluorescence staining and quantitative analysis of aggrecan in the hippocampus of Con and Sev groups at 60 dpe. Green: aggrecan. Blue: DAPI. Scale bar = 50 µm. (J–M) PNNs in the Con group exhibit bright, well‐organized mesh‐like architecture, whereas PNNs in the Sev group display reduced WFA intensity and a disrupted staining pattern. (J) Representative WFA staining. Purple: WFA. Scale bar = 10 µm. (K) Binary images of the regions in (J). (L) Quantification of black/white pixel ratio. (M) Representative surface intensity profiles of WFA‐positive PNNs. (N) Immunoelectron microscopy of PNNs in the Con and Sev groups. High‐magnification images (right panels) display immunogold labeling within WFA (red arrows) in both groups. Scale bars = 1 µm (left), 500 nm (right). Data represent the mean ± SEM. n = 3 mice per group for SEM (B, C, J–N); n = 6 mice per group for all other experiments. Statistical comparisons were conducted using unpaired Student's t‐test. ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ P < 0.0001.

    Journal: Advanced Science

    Article Title: Neonatal Sevoflurane Exposure Induces Long‐Term Cognitive Impairment via Epigenetically Mediated MMP9 Activation and Perineuronal Net Disruption in the Hippocampal CA2

    doi: 10.1002/advs.78006

    Figure Lengend Snippet: Neonatal sevoflurane exposure induces PNNs damage in the hippocampus. (A) Schematic of the experimental protocol for hippocampal extracellular matrix (ECM) analyses. (B) Representative scanning electron micrographs (SEM) of hippocampal ECM ultrastructure in Con and Sev ‑treated groups. Scale bars = 5 µm (5K×), 1.2 µm (20K×), and 500 nm (50K×). (C) Quantification of mean gray value (left) and fiber diameter (right) of hippocampal ECM in Con and Sev groups. (D, E) Wisteria floribunda agglutinin (WFA) staining and quantitative analysis of PNN intensity in the CA2 region at 7, 14, 30, and 60 days post‐exposure (dpe). Scale bar = 50 µm. (F, G) Western blotting and quantification of hapln1 and aggrecan protein levels in the hippocampus of Con and Sev groups. (H, I) Immunofluorescence staining and quantitative analysis of aggrecan in the hippocampus of Con and Sev groups at 60 dpe. Green: aggrecan. Blue: DAPI. Scale bar = 50 µm. (J–M) PNNs in the Con group exhibit bright, well‐organized mesh‐like architecture, whereas PNNs in the Sev group display reduced WFA intensity and a disrupted staining pattern. (J) Representative WFA staining. Purple: WFA. Scale bar = 10 µm. (K) Binary images of the regions in (J). (L) Quantification of black/white pixel ratio. (M) Representative surface intensity profiles of WFA‐positive PNNs. (N) Immunoelectron microscopy of PNNs in the Con and Sev groups. High‐magnification images (right panels) display immunogold labeling within WFA (red arrows) in both groups. Scale bars = 1 µm (left), 500 nm (right). Data represent the mean ± SEM. n = 3 mice per group for SEM (B, C, J–N); n = 6 mice per group for all other experiments. Statistical comparisons were conducted using unpaired Student's t‐test. ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ P < 0.0001.

    Article Snippet: Serial coronal brain sections were blocked in PBS containing 3% BSA and 0.3% Triton X‐100, then incubated overnight at room temperature with the following primary antibodies: biotinylated Wisteria floribunda agglutinin (WFA; Vector Laboratories, 1:300), rabbit anti‐Aggrecan (Merck‐Millipore, MABT83, 1:200), rabbit anti‐NeuN (Merck Millipore, ABN78, 1:600), rabbit anti‐GFAP (GeneTex, GTX108711, 1:200), mouse anti‐CC1 (Sigma–Aldrich, OP80, 1:100), rabbit anti‐Iba1 (WAKO, Lek0542, 1:200), rabbit anti‐Camk2 (GeneTex, GTX135117, 1:200), mouse anti‐CaMKIIα (Abcam, ab22609, 1:200), goat anti‐BDNF (Vector Laboratories, EB08117, 1:500), rabbit anti‐VGlut2 (Abcam, ab216463, 1:200), chicken anti‐MAP2 (Abcam, ab5392, 1:500), and rabbit anti‐MMP9 (Abclonal, A25299, 1:200).After PBS washes, sections were incubated for 3 h at room temperature in the dark with corresponding Alexa Fluor 488‐ or 594‐conjugated secondary antibodies (donkey anti‐rabbit, donkey anti‐mouse, donkey anti‐chicken, and donkey anti‐goat; Jackson ImmunoResearch, 1:500) or Alexa Fluor 488‐conjugated streptavidin (Cell Signaling Technology, 89067S, 1:500).

    Techniques: Staining, Analysis, Western Blot, Immunofluorescence, Immuno-Electron Microscopy, Labeling

    Immunofluorescent detection of α2,6-linked and α2,3-linked sialylated glycans in wild-type and ST6GAL1 −/− /ST3GAL4 −/− pig fibroblasts/respiratory tissues. A: Quantitative reverse transcription-PCR analysis of ST6GAL1 and ST3GAL4 gene expression levels in ST6GAL1 −/− / ST3GAL4 −/− knockout (KO) and wild-type (WT) pig fibroblasts. Data are presented as mean ± standard deviation (SD); n = 3 independent biological replicates, representing fibroblasts isolated from three separate pigs for each genotype group. Statistical significance between WT and KO groups was assessed by unpaired two-sided Student's t -test, with *** P < 0.001. B: Immunofluorescent detection of α2,6-linked and α2,3-linked sialylated glycans in WT and KO pig fibroblasts. C and D: Immunofluorescent detection of α2,6-linked and α2,3-linked sialylated glycans in WT and KO pig respiratory tract. Panels show the upper respiratory tract including larynx, trachea, bronchus (C), and lung tissue (D). Red indicates SNA (α2,6) or MAL Ⅱ (α2,3) staining, and blue indicates nuclei. White arrows indicate sialylated glycan-positive regions in WT samples (B and C). In panel D, WT pigs exhibited a strong α2,6-linked sialylated glycan signal (white arrow) in bronchiolar epithelium, lacking an α2,3-linked sialylated glycan signal (red arrow); both sialylated glycans were present in WT alveoli (green arrows), but absent in KO pig alveoli; weak signals of both sialylated glycans were detected in the bronchiolar epithelium of KO pigs (yellow arrows). Scale bars, 100 μm (B) and 500 μm (C and D).

    Journal: Journal of Biomedical Research

    Article Title: Generation of ST6GAL1/ST3GAL4 double-knockout Landrace pigs with reduced respiratory sialylated glycans and decreased influenza A virus susceptibility of derived fibroblasts in vitro

    doi: 10.7555/JBR.39.20250300

    Figure Lengend Snippet: Immunofluorescent detection of α2,6-linked and α2,3-linked sialylated glycans in wild-type and ST6GAL1 −/− /ST3GAL4 −/− pig fibroblasts/respiratory tissues. A: Quantitative reverse transcription-PCR analysis of ST6GAL1 and ST3GAL4 gene expression levels in ST6GAL1 −/− / ST3GAL4 −/− knockout (KO) and wild-type (WT) pig fibroblasts. Data are presented as mean ± standard deviation (SD); n = 3 independent biological replicates, representing fibroblasts isolated from three separate pigs for each genotype group. Statistical significance between WT and KO groups was assessed by unpaired two-sided Student's t -test, with *** P < 0.001. B: Immunofluorescent detection of α2,6-linked and α2,3-linked sialylated glycans in WT and KO pig fibroblasts. C and D: Immunofluorescent detection of α2,6-linked and α2,3-linked sialylated glycans in WT and KO pig respiratory tract. Panels show the upper respiratory tract including larynx, trachea, bronchus (C), and lung tissue (D). Red indicates SNA (α2,6) or MAL Ⅱ (α2,3) staining, and blue indicates nuclei. White arrows indicate sialylated glycan-positive regions in WT samples (B and C). In panel D, WT pigs exhibited a strong α2,6-linked sialylated glycan signal (white arrow) in bronchiolar epithelium, lacking an α2,3-linked sialylated glycan signal (red arrow); both sialylated glycans were present in WT alveoli (green arrows), but absent in KO pig alveoli; weak signals of both sialylated glycans were detected in the bronchiolar epithelium of KO pigs (yellow arrows). Scale bars, 100 μm (B) and 500 μm (C and D).

    Article Snippet: Cells and tissues were then incubated with biotinylated lectins MAL II (1∶200, Cat. #B-1265-1, Vector Laboratories, Burlingame, CA, USA) or SNA (1∶200, Cat. #B-1305-2, Vector Laboratories) for 1 h, followed by three washes with PBS containing 0.05% Tween-20 (PBST).

    Techniques: Reverse Transcription, Analysis, Gene Expression, Knock-Out, Standard Deviation, Isolation, Tissue, Staining, Glycoproteomics

    Restoration of influenza virus susceptibility in ST6GAL1 −/− / ST3GAL4 −/− Landrace pig cells through complementation. A: Quantitative reverse transcription-PCR (RT-qPCR) analysis of ST6GAL1 and ST3GAL4 gene expression levels in wild-type (WT), knockout (KO), and complemented (KO + ST) cells. B: Immunofluorescent detection of α2,6-linked and α2,3-linked sialylated glycans in wild-type (WT), knockout (KO), and complemented (KO + ST) Landrace pig cells. Red indicates SNA (α2,6) or MAL Ⅱ (α2,3) staining, and blue indicates nuclei. The KO group showed nearly undetectable signals for both SNA and MAL Ⅱ. In contrast, the WT and KO + ST groups exhibited strong fluorescence for both lectins, indicating successful restoration of both α2,6- and α2,3-linked sialylated glycans in complemented cells. Scale bars, 100 μm. C: Cytopathic effects induced by H1N1 influenza virus infection at 0, 24, and 48 h post-infection (HPI). Scale bars, 500 μm. D: Immunofluorescent detection of IAV nucleoprotein (NP) in cells post-infection with H1N1 influenza virus at 48 HPI. NP-Alexa Fluor 488, green; DAPI, blue. Scale bars, 100 μm. E: RT-qPCR analysis of influenza viral M gene expression levels. F: Plaque assay measurement of infectious IAV (H1N1) virus titers (lg PFU/mL). Data are presented as mean ± standard deviation (A, E, and F); n = 3 independent cell culture experiments conducted on separate days with identical cell batches. Statistical significance was determined by one-way ANOVA followed by Tukey's multiple comparisons test. * P < 0.05, ** P < 0.01, and *** P < 0.001.

    Journal: Journal of Biomedical Research

    Article Title: Generation of ST6GAL1/ST3GAL4 double-knockout Landrace pigs with reduced respiratory sialylated glycans and decreased influenza A virus susceptibility of derived fibroblasts in vitro

    doi: 10.7555/JBR.39.20250300

    Figure Lengend Snippet: Restoration of influenza virus susceptibility in ST6GAL1 −/− / ST3GAL4 −/− Landrace pig cells through complementation. A: Quantitative reverse transcription-PCR (RT-qPCR) analysis of ST6GAL1 and ST3GAL4 gene expression levels in wild-type (WT), knockout (KO), and complemented (KO + ST) cells. B: Immunofluorescent detection of α2,6-linked and α2,3-linked sialylated glycans in wild-type (WT), knockout (KO), and complemented (KO + ST) Landrace pig cells. Red indicates SNA (α2,6) or MAL Ⅱ (α2,3) staining, and blue indicates nuclei. The KO group showed nearly undetectable signals for both SNA and MAL Ⅱ. In contrast, the WT and KO + ST groups exhibited strong fluorescence for both lectins, indicating successful restoration of both α2,6- and α2,3-linked sialylated glycans in complemented cells. Scale bars, 100 μm. C: Cytopathic effects induced by H1N1 influenza virus infection at 0, 24, and 48 h post-infection (HPI). Scale bars, 500 μm. D: Immunofluorescent detection of IAV nucleoprotein (NP) in cells post-infection with H1N1 influenza virus at 48 HPI. NP-Alexa Fluor 488, green; DAPI, blue. Scale bars, 100 μm. E: RT-qPCR analysis of influenza viral M gene expression levels. F: Plaque assay measurement of infectious IAV (H1N1) virus titers (lg PFU/mL). Data are presented as mean ± standard deviation (A, E, and F); n = 3 independent cell culture experiments conducted on separate days with identical cell batches. Statistical significance was determined by one-way ANOVA followed by Tukey's multiple comparisons test. * P < 0.05, ** P < 0.01, and *** P < 0.001.

    Article Snippet: Cells and tissues were then incubated with biotinylated lectins MAL II (1∶200, Cat. #B-1265-1, Vector Laboratories, Burlingame, CA, USA) or SNA (1∶200, Cat. #B-1305-2, Vector Laboratories) for 1 h, followed by three washes with PBS containing 0.05% Tween-20 (PBST).

    Techniques: Virus, Reverse Transcription, Quantitative RT-PCR, Analysis, Gene Expression, Knock-Out, Staining, Fluorescence, Infection, Plaque Assay, Standard Deviation, Cell Culture

    Restoration of influenza virus susceptibility in ST6GAL1 −/− / ST3GAL4 −/− Landrace pig cells through complementation. A: Quantitative reverse transcription-PCR (RT-qPCR) analysis of ST6GAL1 and ST3GAL4 gene expression levels in wild-type (WT), knockout (KO), and complemented (KO + ST) cells. B: Immunofluorescent detection of α2,6-linked and α2,3-linked sialylated glycans in wild-type (WT), knockout (KO), and complemented (KO + ST) Landrace pig cells. Red indicates SNA (α2,6) or MAL Ⅱ (α2,3) staining, and blue indicates nuclei. The KO group showed nearly undetectable signals for both SNA and MAL Ⅱ. In contrast, the WT and KO + ST groups exhibited strong fluorescence for both lectins, indicating successful restoration of both α2,6- and α2,3-linked sialylated glycans in complemented cells. Scale bars, 100 μm. C: Cytopathic effects induced by H1N1 influenza virus infection at 0, 24, and 48 h post-infection (HPI). Scale bars, 500 μm. D: Immunofluorescent detection of IAV nucleoprotein (NP) in cells post-infection with H1N1 influenza virus at 48 HPI. NP-Alexa Fluor 488, green; DAPI, blue. Scale bars, 100 μm. E: RT-qPCR analysis of influenza viral M gene expression levels. F: Plaque assay measurement of infectious IAV (H1N1) virus titers (lg PFU/mL). Data are presented as mean ± standard deviation (A, E, and F); n = 3 independent cell culture experiments conducted on separate days with identical cell batches. Statistical significance was determined by one-way ANOVA followed by Tukey's multiple comparisons test. * P < 0.05, ** P < 0.01, and *** P < 0.001.

    Journal: Journal of Biomedical Research

    Article Title: Generation of ST6GAL1/ST3GAL4 double-knockout Landrace pigs with reduced respiratory sialylated glycans and decreased influenza A virus susceptibility of derived fibroblasts in vitro

    doi: 10.7555/JBR.39.20250300

    Figure Lengend Snippet: Restoration of influenza virus susceptibility in ST6GAL1 −/− / ST3GAL4 −/− Landrace pig cells through complementation. A: Quantitative reverse transcription-PCR (RT-qPCR) analysis of ST6GAL1 and ST3GAL4 gene expression levels in wild-type (WT), knockout (KO), and complemented (KO + ST) cells. B: Immunofluorescent detection of α2,6-linked and α2,3-linked sialylated glycans in wild-type (WT), knockout (KO), and complemented (KO + ST) Landrace pig cells. Red indicates SNA (α2,6) or MAL Ⅱ (α2,3) staining, and blue indicates nuclei. The KO group showed nearly undetectable signals for both SNA and MAL Ⅱ. In contrast, the WT and KO + ST groups exhibited strong fluorescence for both lectins, indicating successful restoration of both α2,6- and α2,3-linked sialylated glycans in complemented cells. Scale bars, 100 μm. C: Cytopathic effects induced by H1N1 influenza virus infection at 0, 24, and 48 h post-infection (HPI). Scale bars, 500 μm. D: Immunofluorescent detection of IAV nucleoprotein (NP) in cells post-infection with H1N1 influenza virus at 48 HPI. NP-Alexa Fluor 488, green; DAPI, blue. Scale bars, 100 μm. E: RT-qPCR analysis of influenza viral M gene expression levels. F: Plaque assay measurement of infectious IAV (H1N1) virus titers (lg PFU/mL). Data are presented as mean ± standard deviation (A, E, and F); n = 3 independent cell culture experiments conducted on separate days with identical cell batches. Statistical significance was determined by one-way ANOVA followed by Tukey's multiple comparisons test. * P < 0.05, ** P < 0.01, and *** P < 0.001.

    Article Snippet: Cells and tissues were then incubated with biotinylated lectins MAL II (1∶200, Cat. #B-1265-1, Vector Laboratories, Burlingame, CA, USA) or SNA (1∶200, Cat. #B-1305-2, Vector Laboratories) for 1 h, followed by three washes with PBS containing 0.05% Tween-20 (PBST).

    Techniques: Virus, Reverse Transcription, Quantitative RT-PCR, Analysis, Gene Expression, Knock-Out, Staining, Fluorescence, Infection, Plaque Assay, Standard Deviation, Cell Culture