biotin conjugated concanavalin a  (Vector Laboratories)


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    Name:
    Unconjugated Concanavalin A Con A
    Description:
    Concanavalin A Con A recognizes α linked mannose present as part of a core oligosaccharide in many serum and membrane glycoproteins At neutral and alkaline pH Con A exists as a tetramer of four identical subunits below pH 5 6 Con A dissociates into active dimers of 52 kDa Acetylation succinylation or other derivatizations can also produce stable forms with dimeric structures See succinylated Con A Nicks in the sequence are often present in the purest preparations due to hydrolytic damage within the seeds Con A requires calcium or manganese ions at each of its four saccharide binding sites Although these divalent metal ions are bound tightly to the polypeptide structure buffers which can bind calcium such as phosphate generally should be avoided in diluting Con A since a gradual loss in activity may occur Inhibiting Eluting Sugar mixture of 200 mM α methylmannoside 200 mM α methylglucoside
    Catalog Number:
    L-1000
    Price:
    None
    Category:
    Proteins
    Size:
    500 mg
    Buy from Supplier


    Structured Review

    Vector Laboratories biotin conjugated concanavalin a
    Unconjugated Concanavalin A Con A
    Concanavalin A Con A recognizes α linked mannose present as part of a core oligosaccharide in many serum and membrane glycoproteins At neutral and alkaline pH Con A exists as a tetramer of four identical subunits below pH 5 6 Con A dissociates into active dimers of 52 kDa Acetylation succinylation or other derivatizations can also produce stable forms with dimeric structures See succinylated Con A Nicks in the sequence are often present in the purest preparations due to hydrolytic damage within the seeds Con A requires calcium or manganese ions at each of its four saccharide binding sites Although these divalent metal ions are bound tightly to the polypeptide structure buffers which can bind calcium such as phosphate generally should be avoided in diluting Con A since a gradual loss in activity may occur Inhibiting Eluting Sugar mixture of 200 mM α methylmannoside 200 mM α methylglucoside
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    Images

    1) Product Images from "The Metabolic Chemical Reporter 6‑Azido-6-deoxy-glucose Further Reveals the Substrate Promiscuity of O‑GlcNAc Transferase and Catalyzes the Discovery of Intracellular Protein Modification by O‑Glucose"

    Article Title: The Metabolic Chemical Reporter 6‑Azido-6-deoxy-glucose Further Reveals the Substrate Promiscuity of O‑GlcNAc Transferase and Catalyzes the Discovery of Intracellular Protein Modification by O‑Glucose

    Journal: Journal of the American Chemical Society

    doi: 10.1021/jacs.7b13488

    A significant fraction of 6AzGlc-dependent labeling is O-linked. (A) Known O -GlcNAcylated proteins are labeled by 6AzGlc. H1299 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO for 16 h, followed by CuAAC with a cleavable alkyne-biotin tag. After enrichment on streptavidin beads, the labeled proteins were eluted and visualized by Western blotting. The nonglycosylated protein β -actin is a negative control. (B) A notable fraction of 6AzGlc-dependent signal is sensitive to β -elimination. NIH3T3 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO vehicle for 16 h, followed by CuAAC with alkyne-biotin, separation by SDS-PAGE and transfer to a PVDF membrane. The indicated membranes were then treated for 24 h with either H 2 O or 55 mM NaOH before analysis by streptavidin or Western blotting. (C) 6AzGlc is not incorporated into N-linked glycans. NIH3T3 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO vehicle for 16 h. The corresponding cell lysates were then incubated with either PNGase-F or H 2 O vehicle as indicated before CuAAC with alkyne TAMRA and analysis by in-gel fluorescence. A fraction of the treated lysate was separated before CuAAC and analyzed by Lectin blotting with Concanavalin A (ConA).
    Figure Legend Snippet: A significant fraction of 6AzGlc-dependent labeling is O-linked. (A) Known O -GlcNAcylated proteins are labeled by 6AzGlc. H1299 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO for 16 h, followed by CuAAC with a cleavable alkyne-biotin tag. After enrichment on streptavidin beads, the labeled proteins were eluted and visualized by Western blotting. The nonglycosylated protein β -actin is a negative control. (B) A notable fraction of 6AzGlc-dependent signal is sensitive to β -elimination. NIH3T3 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO vehicle for 16 h, followed by CuAAC with alkyne-biotin, separation by SDS-PAGE and transfer to a PVDF membrane. The indicated membranes were then treated for 24 h with either H 2 O or 55 mM NaOH before analysis by streptavidin or Western blotting. (C) 6AzGlc is not incorporated into N-linked glycans. NIH3T3 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO vehicle for 16 h. The corresponding cell lysates were then incubated with either PNGase-F or H 2 O vehicle as indicated before CuAAC with alkyne TAMRA and analysis by in-gel fluorescence. A fraction of the treated lysate was separated before CuAAC and analyzed by Lectin blotting with Concanavalin A (ConA).

    Techniques Used: Labeling, Western Blot, Negative Control, SDS Page, Incubation, Fluorescence

    2) Product Images from "Glycan Profiling Shows Unvaried N-Glycomes in MSC Clones with Distinct Differentiation Potentials"

    Article Title: Glycan Profiling Shows Unvaried N-Glycomes in MSC Clones with Distinct Differentiation Potentials

    Journal: Frontiers in Cell and Developmental Biology

    doi: 10.3389/fcell.2016.00052

    N -glycan profile of MSCs does not correlate with differentiation capacity. (A) Osteogenic capacities of Y101 and Y202 hTERT-MSC lines. For details see Figure 2A . (B) N -glycan profiles of the Y101 and Y202 hTERT-MSC lines following 24 h culture in basal medium. Representative MALDI-TOF spectra as in Figure 1 , but glycan assignments are omitted for clarity. (C) Heat map display comparing the N -glycan profiles of Y101, Y202 hTERT-MSCs, and osteoblasts derived from the Y101 hTERT-MSCs. Averaged (Y101, Y202 n = 5, osteoblasts n = 3) normalized peak intensities were compared. Each row in the heat map display represents a single glycan structure. Glycans are grouped by type: oligomannose (O), hybrid (H), or complex (C), and sorted by abundance in the Y101 profile within each type. The cells' colors denote glycan abundance as indicated in the legend. (D) Y101 or Y202 cells were grown for 0 or 8 days in basal or osteogenic (osteo) medium, then single cell suspensions stained with ConA-FITC and DAPI and analyzed by flow cytometry. The median intensity of the FITC fluorescence of live cells is shown with SEM from three independent replicates. (E) Y101 cells were cultured in basal medium containing 10 μg/mL swainsonine for 48 h before addition of osteogenic medium containing swainonsine at the same concentration. Mineral deposition and ALP activity were tested by ALP/von Kossa staining 0, 7, 14, and 21 days after addition of osteogenic medium. *** P
    Figure Legend Snippet: N -glycan profile of MSCs does not correlate with differentiation capacity. (A) Osteogenic capacities of Y101 and Y202 hTERT-MSC lines. For details see Figure 2A . (B) N -glycan profiles of the Y101 and Y202 hTERT-MSC lines following 24 h culture in basal medium. Representative MALDI-TOF spectra as in Figure 1 , but glycan assignments are omitted for clarity. (C) Heat map display comparing the N -glycan profiles of Y101, Y202 hTERT-MSCs, and osteoblasts derived from the Y101 hTERT-MSCs. Averaged (Y101, Y202 n = 5, osteoblasts n = 3) normalized peak intensities were compared. Each row in the heat map display represents a single glycan structure. Glycans are grouped by type: oligomannose (O), hybrid (H), or complex (C), and sorted by abundance in the Y101 profile within each type. The cells' colors denote glycan abundance as indicated in the legend. (D) Y101 or Y202 cells were grown for 0 or 8 days in basal or osteogenic (osteo) medium, then single cell suspensions stained with ConA-FITC and DAPI and analyzed by flow cytometry. The median intensity of the FITC fluorescence of live cells is shown with SEM from three independent replicates. (E) Y101 cells were cultured in basal medium containing 10 μg/mL swainsonine for 48 h before addition of osteogenic medium containing swainonsine at the same concentration. Mineral deposition and ALP activity were tested by ALP/von Kossa staining 0, 7, 14, and 21 days after addition of osteogenic medium. *** P

    Techniques Used: Derivative Assay, Staining, Flow Cytometry, Cytometry, Fluorescence, Cell Culture, Concentration Assay, ALP Assay, Activity Assay

    3) Product Images from "Arginase as a Mediator of Diabetic Retinopathy"

    Article Title: Arginase as a Mediator of Diabetic Retinopathy

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2013.00173

    Prevention of diabetes-induced increase in leukocyte adhesion by arginase deletion . Wild type controls, wild type diabetic or arginase AI +/− AII −/− (KO) diabetic mice were perfused through left ventricle with Concanavalin A to label leukocytes attached to the vascular endothelium. The number of attached leukocytes was significantly increased in the wildtype diabetic mice as compared to non-diabetic controls and arginase KO significantly blunted this effect (* p
    Figure Legend Snippet: Prevention of diabetes-induced increase in leukocyte adhesion by arginase deletion . Wild type controls, wild type diabetic or arginase AI +/− AII −/− (KO) diabetic mice were perfused through left ventricle with Concanavalin A to label leukocytes attached to the vascular endothelium. The number of attached leukocytes was significantly increased in the wildtype diabetic mice as compared to non-diabetic controls and arginase KO significantly blunted this effect (* p

    Techniques Used: Mouse Assay

    4) Product Images from "Morniga-G, a T/Tn-Specific Lectin, Induces Leukemic Cell Death via Caspase and DR5 Receptor-Dependent Pathways"

    Article Title: Morniga-G, a T/Tn-Specific Lectin, Induces Leukemic Cell Death via Caspase and DR5 Receptor-Dependent Pathways

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms20010230

    Morniga-G activates healthy human lymphocytes and induces cell death in leukemia cells. PBMCs from healthy donors were cultured for 3 days: ( A ) in the presence of increasing concentrations of Morniga-M (MorM), Morniga-G (MorG), and Con A and proliferative index (from the [ 3 H]-thymidine incorporation) was calculated; and ( B ) in the presence of lectins at concentrations triggering maximal proliferation. CD25 expression was evaluated using flow cytometry, in CD3+ T lymphocytes, in CD19+ CD3- B lymphocytes, and in CD56+ CD3- NK lymphocytes. Values are means ± SD of three experiments performed with three to four different healthy donors. ( C ) Resting PBMCs and Jurkat A3 leukemic cells were incubated with anti-Tn mouse monoclonal antibody + PE-conjugated anti-mouse antibody (αTn + antiα-PE) or FITC-conjugated Morniga G (MorG-FITC) and analyzed using cytofluorimetry. Healthy peripheral lymphocytes (PBLs) were analyzed in a gate corresponding to lymphocytes as defined by the size and granularity parameters. Autofluorescence: purple histograms, dashed green histogram: fluorescent positive cells. ( D ) PBMCs of healthy donors and Jurkat A3 leukemic cells were cultured for 24 h with different concentrations of MorG, then cell viability was evaluated in 3-(4,5-dimethylthiazol)-2-5-diphenyl terazolium bromide MTT reduction assays (mean values ± SD of four independent experiments).
    Figure Legend Snippet: Morniga-G activates healthy human lymphocytes and induces cell death in leukemia cells. PBMCs from healthy donors were cultured for 3 days: ( A ) in the presence of increasing concentrations of Morniga-M (MorM), Morniga-G (MorG), and Con A and proliferative index (from the [ 3 H]-thymidine incorporation) was calculated; and ( B ) in the presence of lectins at concentrations triggering maximal proliferation. CD25 expression was evaluated using flow cytometry, in CD3+ T lymphocytes, in CD19+ CD3- B lymphocytes, and in CD56+ CD3- NK lymphocytes. Values are means ± SD of three experiments performed with three to four different healthy donors. ( C ) Resting PBMCs and Jurkat A3 leukemic cells were incubated with anti-Tn mouse monoclonal antibody + PE-conjugated anti-mouse antibody (αTn + antiα-PE) or FITC-conjugated Morniga G (MorG-FITC) and analyzed using cytofluorimetry. Healthy peripheral lymphocytes (PBLs) were analyzed in a gate corresponding to lymphocytes as defined by the size and granularity parameters. Autofluorescence: purple histograms, dashed green histogram: fluorescent positive cells. ( D ) PBMCs of healthy donors and Jurkat A3 leukemic cells were cultured for 24 h with different concentrations of MorG, then cell viability was evaluated in 3-(4,5-dimethylthiazol)-2-5-diphenyl terazolium bromide MTT reduction assays (mean values ± SD of four independent experiments).

    Techniques Used: Cell Culture, Expressing, Flow Cytometry, Cytometry, Incubation, MTT Assay

    5) Product Images from "SYTO11 staining versus FISH staining: a comparison of two methods to stain Wolbachia pipientis in cell cultures"

    Article Title: SYTO11 staining versus FISH staining: a comparison of two methods to stain Wolbachia pipientis in cell cultures

    Journal: Letters in applied microbiology

    doi: 10.1111/j.1472-765X.2010.02986.x

    FISH staining versus SYTO11 staining: numerical data. Number of infected cells and infection scores as counted on preparations of C7-10R and C7-10B cells seeded on concanavalin A covered slips. The number of infected cells is expressed as percentage. The error bars represent the standard deviation.
    Figure Legend Snippet: FISH staining versus SYTO11 staining: numerical data. Number of infected cells and infection scores as counted on preparations of C7-10R and C7-10B cells seeded on concanavalin A covered slips. The number of infected cells is expressed as percentage. The error bars represent the standard deviation.

    Techniques Used: Fluorescence In Situ Hybridization, Staining, Infection, Standard Deviation

    FISH staining versus SYTO11 staining of C7-10R cells seeded on concanavalin A covered slips. A and C: FISH staining of Wolbachia w Ri in C7-10R cell line. B and D: SYTO11 staining of w Ri in C7-10R. Wolbachia are clearly distinguishable as bright fluorescent particles in the cytoplasm of infected cells. E and F: background staining of cell structures in C7-10. The time of exposure in A, B, C and D was 200 milliseconds, and 1200 milliseconds in E and F. H, M, L: high, medium, low cell infection scores. Scale bars: 10 μm
    Figure Legend Snippet: FISH staining versus SYTO11 staining of C7-10R cells seeded on concanavalin A covered slips. A and C: FISH staining of Wolbachia w Ri in C7-10R cell line. B and D: SYTO11 staining of w Ri in C7-10R. Wolbachia are clearly distinguishable as bright fluorescent particles in the cytoplasm of infected cells. E and F: background staining of cell structures in C7-10. The time of exposure in A, B, C and D was 200 milliseconds, and 1200 milliseconds in E and F. H, M, L: high, medium, low cell infection scores. Scale bars: 10 μm

    Techniques Used: Fluorescence In Situ Hybridization, Staining, Infection

    6) Product Images from "Engineering of spectator glycocalyx structures to evaluate molecular interactions at crowded cellular boundaries."

    Article Title: Engineering of spectator glycocalyx structures to evaluate molecular interactions at crowded cellular boundaries.

    Journal: Faraday discussions

    doi: 10.1039/c9fd00024k

    Association of ConA and SNA lectins with glycocalyx-remodeled RBCs. (A) Binding of fluorescein (FL)-labeled lectins to remodeled cells were assessed via flow cytometry. The presence of spectator glycopolymers 5 at the surface of RBCs attenuates both the initial rate as well as saturation binding of ConA to cell surface glycans. (B) Glycopolymers 5 have no effect on the initial rates of SNA binding but inhibit lectin association near saturation. Relative initial rates were calculated from the linear regions of lectin binding curves and normalized to control cells without polymer treatment. Δ MFI max corresponds to the change in median fluorescence intensity of cells at saturation lectin binding. (ANOVA, Tukey’s multiple comparisons test; p ***
    Figure Legend Snippet: Association of ConA and SNA lectins with glycocalyx-remodeled RBCs. (A) Binding of fluorescein (FL)-labeled lectins to remodeled cells were assessed via flow cytometry. The presence of spectator glycopolymers 5 at the surface of RBCs attenuates both the initial rate as well as saturation binding of ConA to cell surface glycans. (B) Glycopolymers 5 have no effect on the initial rates of SNA binding but inhibit lectin association near saturation. Relative initial rates were calculated from the linear regions of lectin binding curves and normalized to control cells without polymer treatment. Δ MFI max corresponds to the change in median fluorescence intensity of cells at saturation lectin binding. (ANOVA, Tukey’s multiple comparisons test; p ***

    Techniques Used: Binding Assay, Labeling, Flow Cytometry, Fluorescence

    7) Product Images from "Transcriptomic analyses reveal comprehensive responses of insect hemocytes to mycopathogen Beauveria bassiana, and fungal virulence-related cell wall protein assists pathogen to evade host cellular defense"

    Article Title: Transcriptomic analyses reveal comprehensive responses of insect hemocytes to mycopathogen Beauveria bassiana, and fungal virulence-related cell wall protein assists pathogen to evade host cellular defense

    Journal: Virulence

    doi: 10.1080/21505594.2020.1827886

    BbCwp protects fungus from host recognition. (a) Disruption of BbCwp resulted in an enhanced hemocyte encapsulation. Fungal strain was labeled by expressing the mCherry gene. Conidial suspension (5 µl, 10 5 conidia/ml) was injected into host. After an incubation of 3 h at 25°C, hemocyte encapsulation was examined under a fluorescence microscope. (b) Conidial lectin-binding pattern. Lectins included concanavalin A (ConA), Galanthus nivalis lectin (GNL), peanut agglutinin (PNA), and wheat germ agglutinin (WGA). Δ BbCwp mutant strain displayed a significant increase in fluorescence intensity of WGA. (c) Relative expression levels of β-1, 3-glucan recognition protein genes ( βGRP ). In G. mellonella , there are 11 βGRP genes. Comparative analyses between the wild type/Δ BbCwp were performed at different time points during infection process. Gene disruption led to a significant up-regulation of all tested genes at 1 d post infection. Tukey’s HSD was used to determine the statistical significance using a threshold of P
    Figure Legend Snippet: BbCwp protects fungus from host recognition. (a) Disruption of BbCwp resulted in an enhanced hemocyte encapsulation. Fungal strain was labeled by expressing the mCherry gene. Conidial suspension (5 µl, 10 5 conidia/ml) was injected into host. After an incubation of 3 h at 25°C, hemocyte encapsulation was examined under a fluorescence microscope. (b) Conidial lectin-binding pattern. Lectins included concanavalin A (ConA), Galanthus nivalis lectin (GNL), peanut agglutinin (PNA), and wheat germ agglutinin (WGA). Δ BbCwp mutant strain displayed a significant increase in fluorescence intensity of WGA. (c) Relative expression levels of β-1, 3-glucan recognition protein genes ( βGRP ). In G. mellonella , there are 11 βGRP genes. Comparative analyses between the wild type/Δ BbCwp were performed at different time points during infection process. Gene disruption led to a significant up-regulation of all tested genes at 1 d post infection. Tukey’s HSD was used to determine the statistical significance using a threshold of P

    Techniques Used: Labeling, Expressing, Injection, Incubation, Fluorescence, Microscopy, Binding Assay, Whole Genome Amplification, Mutagenesis, Infection

    8) Product Images from "Ischaemia-induced retinal neovascularisation and diabetic retinopathy in mice with conditional knockout of hypoxia-inducible factor-1 in retinal Müller cells"

    Article Title: Ischaemia-induced retinal neovascularisation and diabetic retinopathy in mice with conditional knockout of hypoxia-inducible factor-1 in retinal Müller cells

    Journal: Diabetologia

    doi: 10.1007/s00125-011-2081-0

    Decreased leucostasis in a Hif-1α KO mouse model of type 1 diabetes. a Non-diabetic mice, ( b ) diabetic wild-type (WT) mice and ( c ) diabetic Hif-1α KO mice at 2 months after onset of streptozotocin-induced diabetes were used for leucostasis assay. Adherent leucocytes (arrow) were stained with FITC-conjugated concanavalin-A in the retinal vasculature after removal of circulating leucocytes by perfusion. The retinal vasculature and leucocytes were visualised in retinal flat mounts under fluorescence microscope. Scale bar 50 μm. d Adherent leucocytes in the retinal vasculature were counted and averaged, showing that diabetic (DM) Hif-1α KO mice had significantly fewer leucocytes than diabetic WT mice. Values are mean±SEM; n =5, ** p
    Figure Legend Snippet: Decreased leucostasis in a Hif-1α KO mouse model of type 1 diabetes. a Non-diabetic mice, ( b ) diabetic wild-type (WT) mice and ( c ) diabetic Hif-1α KO mice at 2 months after onset of streptozotocin-induced diabetes were used for leucostasis assay. Adherent leucocytes (arrow) were stained with FITC-conjugated concanavalin-A in the retinal vasculature after removal of circulating leucocytes by perfusion. The retinal vasculature and leucocytes were visualised in retinal flat mounts under fluorescence microscope. Scale bar 50 μm. d Adherent leucocytes in the retinal vasculature were counted and averaged, showing that diabetic (DM) Hif-1α KO mice had significantly fewer leucocytes than diabetic WT mice. Values are mean±SEM; n =5, ** p

    Techniques Used: Mouse Assay, Staining, Fluorescence, Microscopy

    9) Product Images from "Role of IL-6 in Angiotensin II-Induced Retinal Vascular Inflammation"

    Article Title: Role of IL-6 in Angiotensin II-Induced Retinal Vascular Inflammation

    Journal: Investigative Ophthalmology & Visual Science

    doi: 10.1167/iovs.09-3375

    Suppression of angiotensin II–induced increases in retinal vascular leukostasis in IL-6–deficient mice. ( A ) Flatmount images of concanavalin A–labeled retinas showing adherent leukocytes ( arrows ) within the retinal vessels of wild-type
    Figure Legend Snippet: Suppression of angiotensin II–induced increases in retinal vascular leukostasis in IL-6–deficient mice. ( A ) Flatmount images of concanavalin A–labeled retinas showing adherent leukocytes ( arrows ) within the retinal vessels of wild-type

    Techniques Used: Mouse Assay, Labeling

    IL-6 induced leukostasis in both wild-type and IL-6–deficient mice. ( A ) Flatmount images of concanavalin A–labeled retinas showing adherent leukocytes ( arrows ) within the retinal vessels of wild-type (WT) and IL-6–deficient (IL-6ko)
    Figure Legend Snippet: IL-6 induced leukostasis in both wild-type and IL-6–deficient mice. ( A ) Flatmount images of concanavalin A–labeled retinas showing adherent leukocytes ( arrows ) within the retinal vessels of wild-type (WT) and IL-6–deficient (IL-6ko)

    Techniques Used: Mouse Assay, Labeling

    10) Product Images from "Rapid CB1 cannabinoid receptor desensitization defines the time course of ERK1/2 MAP kinase signaling"

    Article Title: Rapid CB1 cannabinoid receptor desensitization defines the time course of ERK1/2 MAP kinase signaling

    Journal: Neuropharmacology

    doi: 10.1016/j.neuropharm.2007.06.005

    S426A/S430A CB 1 receptor internalization and pharmacological blockade of CB 1 receptor internalization with Concanavalin A
    Figure Legend Snippet: S426A/S430A CB 1 receptor internalization and pharmacological blockade of CB 1 receptor internalization with Concanavalin A

    Techniques Used:

    11) Product Images from "PPARα activation directly upregulates thrombomodulin in the diabetic retina"

    Article Title: PPARα activation directly upregulates thrombomodulin in the diabetic retina

    Journal: Scientific Reports

    doi: 10.1038/s41598-020-67579-1

    Pemafibrate inhibits retinal vascular leukostasis by upregulating TM. ( a ) Fluorescence microscopy images showing retinal adherent vascular leukocytes in diabetic rats. STZ-induced diabetic rats were treated with pemafibrate or vehicle and intravitreally injected with control siRNA or siRNA targeting Thbd . The retinal vascular adherent leukocytes were stained with FITC-conjugated concanavalin-A. Representative images of retinal flat mounts from nondiabetic rats (upper left), untreated diabetic rats (upper right), diabetic rats treated with pemafibrate and control siRNA (lower left), and diabetic rats treated with pemafibrate and si Thbd (lower right) are shown. Arrows indicate adherent leukocytes. Scale bar = 50 μm. ( b ) Quantification of adherent leukocytes in retinal images (n = 6 per group). (Results are expressed as mean ± SD. *p
    Figure Legend Snippet: Pemafibrate inhibits retinal vascular leukostasis by upregulating TM. ( a ) Fluorescence microscopy images showing retinal adherent vascular leukocytes in diabetic rats. STZ-induced diabetic rats were treated with pemafibrate or vehicle and intravitreally injected with control siRNA or siRNA targeting Thbd . The retinal vascular adherent leukocytes were stained with FITC-conjugated concanavalin-A. Representative images of retinal flat mounts from nondiabetic rats (upper left), untreated diabetic rats (upper right), diabetic rats treated with pemafibrate and control siRNA (lower left), and diabetic rats treated with pemafibrate and si Thbd (lower right) are shown. Arrows indicate adherent leukocytes. Scale bar = 50 μm. ( b ) Quantification of adherent leukocytes in retinal images (n = 6 per group). (Results are expressed as mean ± SD. *p

    Techniques Used: Fluorescence, Microscopy, Injection, Staining

    12) Product Images from "Death and Transfiguration in Static Staphylococcus epidermidis Cultures"

    Article Title: Death and Transfiguration in Static Staphylococcus epidermidis Cultures

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0100002

    Early stage structures. A–C. Camera pictures, MH strain: floating and sedimented macroscopic aggregates of a 1-day culture. D. Scanning electron microscopy (SEM) picture, MH strain, 1-day culture: bacteria (arrow 1) and matrix elements (arrow 2) form small aggregates. E. SEM picture, 3-day culture, MH strain: a fibrous sheet (region 1) and short strands (arrow 2) form larger aggregates. F. SEM picture, 5-day culture, MH strain: aggregates consist of solid sheets (region 1) and subjacent a network of strands (region 2). G. SEM picture, 5-day culture, MH strain: predominantly parallel orientation of the strand network. H. SEM picture, 5-day culture: bacteria-associated strands (arrow 1), cross-connected by fibers (arrow 2). I. Confocal laser scanning microscopy (cLSM) picture, MH strain, 10-day culture: strands and associated particulate material (arrows 1, 2) are stained with Concanavalin A; bacteria are labeled with Syto59 (arrow 3). J. SEM picture, 10-day culture, MH strain: almost solid, parallel strands with cross-connecting fibers.
    Figure Legend Snippet: Early stage structures. A–C. Camera pictures, MH strain: floating and sedimented macroscopic aggregates of a 1-day culture. D. Scanning electron microscopy (SEM) picture, MH strain, 1-day culture: bacteria (arrow 1) and matrix elements (arrow 2) form small aggregates. E. SEM picture, 3-day culture, MH strain: a fibrous sheet (region 1) and short strands (arrow 2) form larger aggregates. F. SEM picture, 5-day culture, MH strain: aggregates consist of solid sheets (region 1) and subjacent a network of strands (region 2). G. SEM picture, 5-day culture, MH strain: predominantly parallel orientation of the strand network. H. SEM picture, 5-day culture: bacteria-associated strands (arrow 1), cross-connected by fibers (arrow 2). I. Confocal laser scanning microscopy (cLSM) picture, MH strain, 10-day culture: strands and associated particulate material (arrows 1, 2) are stained with Concanavalin A; bacteria are labeled with Syto59 (arrow 3). J. SEM picture, 10-day culture, MH strain: almost solid, parallel strands with cross-connecting fibers.

    Techniques Used: Electron Microscopy, Confocal Laser Scanning Microscopy, Staining, Labeling

    Specific staining, time-course and development model. A. cLSM picture, 14-day culture, strain #49134: the EUB338 probe labels the compartment structure (red), while the bacteria are double-stained by the EUB338 probe and Syto59 (yellow). B. cLSM picture, 14-day culture, MH strain: Concanavalin A stains the compartment structure, but not the bacteria (arrow 1). C. cLSM picture, 14-day culture, strain #49134: the Sypro protein stain (green) and the Syto59 nucleic acid stain label both the bacteria (yellow) but not the compartment structure. D. cLSM picture, 14-day culture, strain #49134: the lipophilic stain Nile Red (green) does not label compartment walls, only diffuse biofilm matrix. E. SEM picture, 28-day culture, MH strain: huge aggregates with intact bacteria and strands continued to be present in long-time cultures. F. SEM picture, MH strain, overnight grown colony on agar. G. The model depicts the main development stages and their timely appearances in the context with the CFU time-course (MH strain).
    Figure Legend Snippet: Specific staining, time-course and development model. A. cLSM picture, 14-day culture, strain #49134: the EUB338 probe labels the compartment structure (red), while the bacteria are double-stained by the EUB338 probe and Syto59 (yellow). B. cLSM picture, 14-day culture, MH strain: Concanavalin A stains the compartment structure, but not the bacteria (arrow 1). C. cLSM picture, 14-day culture, strain #49134: the Sypro protein stain (green) and the Syto59 nucleic acid stain label both the bacteria (yellow) but not the compartment structure. D. cLSM picture, 14-day culture, strain #49134: the lipophilic stain Nile Red (green) does not label compartment walls, only diffuse biofilm matrix. E. SEM picture, 28-day culture, MH strain: huge aggregates with intact bacteria and strands continued to be present in long-time cultures. F. SEM picture, MH strain, overnight grown colony on agar. G. The model depicts the main development stages and their timely appearances in the context with the CFU time-course (MH strain).

    Techniques Used: Staining, Confocal Laser Scanning Microscopy

    13) Product Images from "Retinal Vascular Endothelial Growth Factor Induces Intercellular Adhesion Molecule-1 and Endothelial Nitric Oxide Synthase Expression and Initiates Early Diabetic Retinal Leukocyte Adhesion in Vivo"

    Article Title: Retinal Vascular Endothelial Growth Factor Induces Intercellular Adhesion Molecule-1 and Endothelial Nitric Oxide Synthase Expression and Initiates Early Diabetic Retinal Leukocyte Adhesion in Vivo

    Journal: The American Journal of Pathology

    doi:

    Lectin staining with Concanavalin A.  A:  Retinal venule of a diabetic animal with adherent leukocytes after perfusion removal of nonadherent blood elements. Adherent leukocyte density after lectin staining in arterioles ( B ), venules ( C ), and capillaries ( D ). Compared to the diabetic animals, the retinae of nondiabetic animals showed two- to threefold more adherent leukocytes in the arterioles, venules, and capillaries, respectively ( P
    Figure Legend Snippet: Lectin staining with Concanavalin A. A: Retinal venule of a diabetic animal with adherent leukocytes after perfusion removal of nonadherent blood elements. Adherent leukocyte density after lectin staining in arterioles ( B ), venules ( C ), and capillaries ( D ). Compared to the diabetic animals, the retinae of nondiabetic animals showed two- to threefold more adherent leukocytes in the arterioles, venules, and capillaries, respectively ( P

    Techniques Used: Staining

    14) Product Images from "Choroidal neovascularization is inhibited via an intraocular decrease of inflammatory cells in mice lacking complement component C3"

    Article Title: Choroidal neovascularization is inhibited via an intraocular decrease of inflammatory cells in mice lacking complement component C3

    Journal: Scientific Reports

    doi: 10.1038/srep15702

    Lesion size after laser photocoagulation. At 7 days after laser injury, the mice were perfused transcardially with PBS and then with FITC-conjugated concanavalin A (20 μg/mL in PBS) to label the choroidal neovascularization (CNVs). The eyes were removed and retinal pigment epithelium (RPE)-choroid flatmounts were prepared. Measurements of CNV lesion area were carried out using ImageJ. The outline of the CNV was drawn around the contour of the lesion and then the total lesion area was measured. The average area obtained from 4 lesions in each eye was used for analysis. ( A ) Lesion size was significantly smaller in C3 −/− mice than in wild-type (WT) mice. ( B ) The CNV in a representative WT mouse (white arrows). ( C ) The CNV lesion in a representative C3 −/− mouse is smaller than that in the WT mouse shown in B. Scale bars, 100 μm. * P
    Figure Legend Snippet: Lesion size after laser photocoagulation. At 7 days after laser injury, the mice were perfused transcardially with PBS and then with FITC-conjugated concanavalin A (20 μg/mL in PBS) to label the choroidal neovascularization (CNVs). The eyes were removed and retinal pigment epithelium (RPE)-choroid flatmounts were prepared. Measurements of CNV lesion area were carried out using ImageJ. The outline of the CNV was drawn around the contour of the lesion and then the total lesion area was measured. The average area obtained from 4 lesions in each eye was used for analysis. ( A ) Lesion size was significantly smaller in C3 −/− mice than in wild-type (WT) mice. ( B ) The CNV in a representative WT mouse (white arrows). ( C ) The CNV lesion in a representative C3 −/− mouse is smaller than that in the WT mouse shown in B. Scale bars, 100 μm. * P

    Techniques Used: Mouse Assay

    15) Product Images from "Morniga-G, a T/Tn-Specific Lectin, Induces Leukemic Cell Death via Caspase and DR5 Receptor-Dependent Pathways"

    Article Title: Morniga-G, a T/Tn-Specific Lectin, Induces Leukemic Cell Death via Caspase and DR5 Receptor-Dependent Pathways

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms20010230

    Morniga-G activates healthy human lymphocytes and induces cell death in leukemia cells. PBMCs from healthy donors were cultured for 3 days: ( A ) in the presence of increasing concentrations of Morniga-M (MorM), Morniga-G (MorG), and Con A and proliferative index (from the [ 3 H]-thymidine incorporation) was calculated; and ( B ) in the presence of lectins at concentrations triggering maximal proliferation. CD25 expression was evaluated using flow cytometry, in CD3+ T lymphocytes, in CD19+ CD3- B lymphocytes, and in CD56+ CD3- NK lymphocytes. Values are means ± SD of three experiments performed with three to four different healthy donors. ( C ) Resting PBMCs and Jurkat A3 leukemic cells were incubated with anti-Tn mouse monoclonal antibody + PE-conjugated anti-mouse antibody (αTn + antiα-PE) or FITC-conjugated Morniga G (MorG-FITC) and analyzed using cytofluorimetry. Healthy peripheral lymphocytes (PBLs) were analyzed in a gate corresponding to lymphocytes as defined by the size and granularity parameters. Autofluorescence: purple histograms, dashed green histogram: fluorescent positive cells. ( D ) PBMCs of healthy donors and Jurkat A3 leukemic cells were cultured for 24 h with different concentrations of MorG, then cell viability was evaluated in 3-(4,5-dimethylthiazol)-2-5-diphenyl terazolium bromide MTT reduction assays (mean values ± SD of four independent experiments).
    Figure Legend Snippet: Morniga-G activates healthy human lymphocytes and induces cell death in leukemia cells. PBMCs from healthy donors were cultured for 3 days: ( A ) in the presence of increasing concentrations of Morniga-M (MorM), Morniga-G (MorG), and Con A and proliferative index (from the [ 3 H]-thymidine incorporation) was calculated; and ( B ) in the presence of lectins at concentrations triggering maximal proliferation. CD25 expression was evaluated using flow cytometry, in CD3+ T lymphocytes, in CD19+ CD3- B lymphocytes, and in CD56+ CD3- NK lymphocytes. Values are means ± SD of three experiments performed with three to four different healthy donors. ( C ) Resting PBMCs and Jurkat A3 leukemic cells were incubated with anti-Tn mouse monoclonal antibody + PE-conjugated anti-mouse antibody (αTn + antiα-PE) or FITC-conjugated Morniga G (MorG-FITC) and analyzed using cytofluorimetry. Healthy peripheral lymphocytes (PBLs) were analyzed in a gate corresponding to lymphocytes as defined by the size and granularity parameters. Autofluorescence: purple histograms, dashed green histogram: fluorescent positive cells. ( D ) PBMCs of healthy donors and Jurkat A3 leukemic cells were cultured for 24 h with different concentrations of MorG, then cell viability was evaluated in 3-(4,5-dimethylthiazol)-2-5-diphenyl terazolium bromide MTT reduction assays (mean values ± SD of four independent experiments).

    Techniques Used: Cell Culture, Expressing, Flow Cytometry, Cytometry, Incubation, MTT Assay

    16) Product Images from "Adenosine deaminase deficiency increases thymic apoptosis and causes defective T cell receptor signaling"

    Article Title: Adenosine deaminase deficiency increases thymic apoptosis and causes defective T cell receptor signaling

    Journal: Journal of Clinical Investigation

    doi:

    ADA deficiency is accompanied with defects in TCR signaling pathways in vivo and in vitro. ( a ) Partial phosphorylation of CD3 ζ chain is notably reduced in nonactivated ex vivo thymocytes from ADA –/– mice, whereas ZAP-70 levels are shown to be the same in parallel samples of immunoprecipitates from ex vivo thymocytes harvested from ADA –/– and ADA +/+ littermates. ( b ) Intracellular Ca 2+ mobilization upon ConA stimulation is inhibited by adenosine in normal thymocytes in the presence of the ADA inhibitor EHNA. Thymocytes from ADA +/+ or ADA –/– mice were used for isolation of ZAP-70 immunocomplexes followed by immunoblotting analysis of TCR ζ chain phosphorylation with an anti phosphotyrosine mAb as described in Methods. Anti ZAP-70 mAb’s were used in control immunoblotting. For measurements of calcium flux ADA +/+ thymocytes were preloaded with indo-1 and analyzed on a FACSVantage flow cytometer as described in Methods. The percentages of cells that increase intracellular calcium after stimulation with Concanavalin A are shown on the graphs. Adenosine (100 μΜ) alone or in combination with EHNA (10 μM) was added a minute before the ConA stimulation. Arrow indicates time of injection of T cell–activating stimuli.
    Figure Legend Snippet: ADA deficiency is accompanied with defects in TCR signaling pathways in vivo and in vitro. ( a ) Partial phosphorylation of CD3 ζ chain is notably reduced in nonactivated ex vivo thymocytes from ADA –/– mice, whereas ZAP-70 levels are shown to be the same in parallel samples of immunoprecipitates from ex vivo thymocytes harvested from ADA –/– and ADA +/+ littermates. ( b ) Intracellular Ca 2+ mobilization upon ConA stimulation is inhibited by adenosine in normal thymocytes in the presence of the ADA inhibitor EHNA. Thymocytes from ADA +/+ or ADA –/– mice were used for isolation of ZAP-70 immunocomplexes followed by immunoblotting analysis of TCR ζ chain phosphorylation with an anti phosphotyrosine mAb as described in Methods. Anti ZAP-70 mAb’s were used in control immunoblotting. For measurements of calcium flux ADA +/+ thymocytes were preloaded with indo-1 and analyzed on a FACSVantage flow cytometer as described in Methods. The percentages of cells that increase intracellular calcium after stimulation with Concanavalin A are shown on the graphs. Adenosine (100 μΜ) alone or in combination with EHNA (10 μM) was added a minute before the ConA stimulation. Arrow indicates time of injection of T cell–activating stimuli.

    Techniques Used: In Vivo, In Vitro, Ex Vivo, Mouse Assay, Isolation, Flow Cytometry, Cytometry, Injection

    17) Product Images from "Complete B Cell Deficiency Reduces Allograft Inflammation and Intragraft Macrophages a Rat Kidney Transplant Model"

    Article Title: Complete B Cell Deficiency Reduces Allograft Inflammation and Intragraft Macrophages a Rat Kidney Transplant Model

    Journal: Transplantation

    doi: 10.1097/TP.0000000000002010

    Splenocytes from wild type and B cell deficient rats produced IFN-γ in response to activation in vitro A, Low levels of IFN-γ producing cells were present in media treated splenocytes from wild type and B −/− rats. Stimulation with the T-cell activator Concanavalin A (ConA) resulted in a significant increase in IFN-γ producing cells compared to media treated controls. *** P
    Figure Legend Snippet: Splenocytes from wild type and B cell deficient rats produced IFN-γ in response to activation in vitro A, Low levels of IFN-γ producing cells were present in media treated splenocytes from wild type and B −/− rats. Stimulation with the T-cell activator Concanavalin A (ConA) resulted in a significant increase in IFN-γ producing cells compared to media treated controls. *** P

    Techniques Used: Produced, Activation Assay, In Vitro

    18) Product Images from "Rabbit CD200R binds host CD200 but not CD200-like proteins from poxviruses"

    Article Title: Rabbit CD200R binds host CD200 but not CD200-like proteins from poxviruses

    Journal: Virology

    doi: 10.1016/j.virol.2015.10.026

    Flow cytometry showing rabbit CD200R coated beads do not bind myxoma infected cells. (A) RK13 cell lines infected with GFP tagged wild type (WT vMyx GFP) or GFP tagged M141 knock out (M141 KO vMyx GFP) virus were harvested 24 hpi. Both WT vMyx GFP (red) and M141 KO vMyx GFP (blue) infections generated comparable numbers of virus positive cells as shown by GFP expression in flow cytometry. Gray shaded area refers to uninfected cells. The gate indicates the GFP high population of the infected cells used for the following bead binding analyses. (B) Uninfected (C) WT vMyx GFP infected and (D) M141 KO vMyx GFP infected cells were tested for binding to nile red fluorescent beads coated with biotinylated rCD4 d3+4 chimeric proteins of rabbit CD200R (blue). Beads coated with biotinylated rCD4 d3+4 only (gray shaded) were used as negative control whereas, biotinylated concanavalin A coated beads were used as positive control for the bead binding assay. Flow cytometry plots are representatives of three independent experiments.
    Figure Legend Snippet: Flow cytometry showing rabbit CD200R coated beads do not bind myxoma infected cells. (A) RK13 cell lines infected with GFP tagged wild type (WT vMyx GFP) or GFP tagged M141 knock out (M141 KO vMyx GFP) virus were harvested 24 hpi. Both WT vMyx GFP (red) and M141 KO vMyx GFP (blue) infections generated comparable numbers of virus positive cells as shown by GFP expression in flow cytometry. Gray shaded area refers to uninfected cells. The gate indicates the GFP high population of the infected cells used for the following bead binding analyses. (B) Uninfected (C) WT vMyx GFP infected and (D) M141 KO vMyx GFP infected cells were tested for binding to nile red fluorescent beads coated with biotinylated rCD4 d3+4 chimeric proteins of rabbit CD200R (blue). Beads coated with biotinylated rCD4 d3+4 only (gray shaded) were used as negative control whereas, biotinylated concanavalin A coated beads were used as positive control for the bead binding assay. Flow cytometry plots are representatives of three independent experiments.

    Techniques Used: Flow Cytometry, Cytometry, Infection, Knock-Out, Generated, Expressing, Binding Assay, Negative Control, Positive Control

    19) Product Images from "Photogenerated Lectin Sensors Produced by Thiol-Ene/Yne Photo-Click Chemistry in Aqueous Solution"

    Article Title: Photogenerated Lectin Sensors Produced by Thiol-Ene/Yne Photo-Click Chemistry in Aqueous Solution

    Journal: Biosensors & bioelectronics

    doi: 10.1016/j.bios.2012.01.001

    Illustration of referenced binding curves of triplicate injections of Con A and RCA-I to mannose-/galactose-surfaces on a) alkyne-based surfaces and b) alkene-based surfaces.
    Figure Legend Snippet: Illustration of referenced binding curves of triplicate injections of Con A and RCA-I to mannose-/galactose-surfaces on a) alkyne-based surfaces and b) alkene-based surfaces.

    Techniques Used: Binding Assay

    20) Product Images from "Glycosylated gold nanoparticle libraries for label-free multiplexed lectin biosensing gold nanoparticle libraries for label-free multiplexed lectin biosensing †Electronic supplementary information (ESI) available. See DOI: 10.1039/c5tb01994jClick here for additional data file."

    Article Title: Glycosylated gold nanoparticle libraries for label-free multiplexed lectin biosensing gold nanoparticle libraries for label-free multiplexed lectin biosensing †Electronic supplementary information (ESI) available. See DOI: 10.1039/c5tb01994jClick here for additional data file.

    Journal: Journal of Materials Chemistry. B, Materials for Biology and Medicine

    doi: 10.1039/c5tb01994j

    Dose-dependent binding isotherms of Man, Gal, Glc, ManNAc, GalNAc, GlcNAc and Fuc functionalised particles with (A) Con A (B) RCA 120  (C) SBA (D) WGA (E) UEA.
    Figure Legend Snippet: Dose-dependent binding isotherms of Man, Gal, Glc, ManNAc, GalNAc, GlcNAc and Fuc functionalised particles with (A) Con A (B) RCA 120 (C) SBA (D) WGA (E) UEA.

    Techniques Used: Binding Assay, Gas Chromatography, Whole Genome Amplification

    Lectin discrimination. (A) Absorbance at 700 nm of Man, Gal, Glc, ManNAc, GalNAc and GlcNAc functionalised particles with 6.25 μg mL –1  Con A, RCA 120 , SBA, WGA, PNA, UEA. Each is the average of 5 measurements. (B) Linear discriminant plot showing excellent segregation between lectins.
    Figure Legend Snippet: Lectin discrimination. (A) Absorbance at 700 nm of Man, Gal, Glc, ManNAc, GalNAc and GlcNAc functionalised particles with 6.25 μg mL –1 Con A, RCA 120 , SBA, WGA, PNA, UEA. Each is the average of 5 measurements. (B) Linear discriminant plot showing excellent segregation between lectins.

    Techniques Used: Gas Chromatography, Whole Genome Amplification

    21) Product Images from "Leukostasis and pigment epithelium-derived factor in rat models of diabetic retinopathy"

    Article Title: Leukostasis and pigment epithelium-derived factor in rat models of diabetic retinopathy

    Journal: Molecular Vision

    doi:

    Adherent leukocytes in the retinal vasculature. Confocal fluorescence photographs of retinal vasculature and adherent leukocytes in Sprague-Dawley (SD) rats as control ( A , D , G ), streptozotocin-induced diabetic rats (STZ), which were spontaneously diabetic Torii (SDT) rats: ( B , E , H ), and SDT rats ( C , F , I ). The retinal vasculature and adherent leukocytes were stained green by FITC-conjugated concanavalin A lectin ( A , B , C ), and the adherent leukocytes were labeled with phycoerythrin-conjugated anti-rat CD45 antibody ( D , E , F ). The adherent leukocytes appear yellow in the merges images ( G , H , I ). Scale bar equals 20 μm. J shows the Index of retinal leukostasis. The adherent leukocytes were significantly increased in STZ rats compared to that of control SD rats in short term experiments (control vs STZ, p
    Figure Legend Snippet: Adherent leukocytes in the retinal vasculature. Confocal fluorescence photographs of retinal vasculature and adherent leukocytes in Sprague-Dawley (SD) rats as control ( A , D , G ), streptozotocin-induced diabetic rats (STZ), which were spontaneously diabetic Torii (SDT) rats: ( B , E , H ), and SDT rats ( C , F , I ). The retinal vasculature and adherent leukocytes were stained green by FITC-conjugated concanavalin A lectin ( A , B , C ), and the adherent leukocytes were labeled with phycoerythrin-conjugated anti-rat CD45 antibody ( D , E , F ). The adherent leukocytes appear yellow in the merges images ( G , H , I ). Scale bar equals 20 μm. J shows the Index of retinal leukostasis. The adherent leukocytes were significantly increased in STZ rats compared to that of control SD rats in short term experiments (control vs STZ, p

    Techniques Used: Fluorescence, Staining, Labeling

    Confocal fluorescence microscopy for detection of adherent monocytes to human umbilical vein endothelial cells. Monocytes and nuclei of human umbilical vein endothelial cells (HUVECs) were labeled in green with Calcein-AM ( A ). The cell surfaces of the monocytes and HUVECs were labeled red with rhodamine-conjugated Con-A lectin ( B ). Adherent monocytes appeared yellow in the merged images obtained by combining FITC and rhodamine images ( C ). Arrows show monocytes. D - G show merged images of HUVECs and adherent monocytes. HUVECs were treated with phosphate buffered saline as a control ( D ), vascular endothelial growth factor (VEGF; E ), pigment epithelial growth factor (PEDF; F ), and both VEGF and PEDF ( G ). H shows quantification of adherent monocytes. Administration of PEDF did not significantly alter the number of adherent monocytes compared to that of controls. Alternatively, when VEGF was added, the number of adherent monocytes was significantly increased (p
    Figure Legend Snippet: Confocal fluorescence microscopy for detection of adherent monocytes to human umbilical vein endothelial cells. Monocytes and nuclei of human umbilical vein endothelial cells (HUVECs) were labeled in green with Calcein-AM ( A ). The cell surfaces of the monocytes and HUVECs were labeled red with rhodamine-conjugated Con-A lectin ( B ). Adherent monocytes appeared yellow in the merged images obtained by combining FITC and rhodamine images ( C ). Arrows show monocytes. D - G show merged images of HUVECs and adherent monocytes. HUVECs were treated with phosphate buffered saline as a control ( D ), vascular endothelial growth factor (VEGF; E ), pigment epithelial growth factor (PEDF; F ), and both VEGF and PEDF ( G ). H shows quantification of adherent monocytes. Administration of PEDF did not significantly alter the number of adherent monocytes compared to that of controls. Alternatively, when VEGF was added, the number of adherent monocytes was significantly increased (p

    Techniques Used: Fluorescence, Microscopy, Labeling

    22) Product Images from "Glycoprofiling as a novel tool in serological assays of systemic sclerosis: A comparative study with three bioanalytical methods"

    Article Title: Glycoprofiling as a novel tool in serological assays of systemic sclerosis: A comparative study with three bioanalytical methods

    Journal: Analytica chimica acta

    doi: 10.1016/j.aca.2014.10.029

    AFM images (1x1 μm) of unmodified microarray slide (upper image with R q =0.5 nm) and the same slide after the sample spotting and incubation with a lectin (bottom image with R q =2.4 nm). Isolated islets of the proteins (INV, Con A and streptavidin
    Figure Legend Snippet: AFM images (1x1 μm) of unmodified microarray slide (upper image with R q =0.5 nm) and the same slide after the sample spotting and incubation with a lectin (bottom image with R q =2.4 nm). Isolated islets of the proteins (INV, Con A and streptavidin

    Techniques Used: Microarray, Incubation, Isolation

    23) Product Images from "Therapeutic Effects of a Novel Agonist of Peroxisome Proliferator-Activated Receptor Alpha for the Treatment of Diabetic Retinopathy"

    Article Title: Therapeutic Effects of a Novel Agonist of Peroxisome Proliferator-Activated Receptor Alpha for the Treatment of Diabetic Retinopathy

    Journal: Investigative Ophthalmology & Visual Science

    doi: 10.1167/iovs.16-21402

    Effects of Y-0452 on retinal vascular inflammation and leakage in diabetic rats. Rats with 1 month of STZ-induced diabetes received daily intraperitoneal injections of Y-0452 (10 mg/kg/d) or the same volume of the vehicle (Veh) as a control for 3 weeks. Retinal vascular endothelial cells and adherent leukocytes were stained with FITC–concanavalin-A and visualized under a fluorescence microscope. (A–C) Representative images of retinal adherent leukocytes in nondiabetic and STZ-induced diabetic rats ([A] nondiabetic, [B] STZ+Veh, [C] STZ+Y-0452). (D) Quantification of retinal adherent leukocytes was performed from ×40 magnification images. Adherent leukocytes were counted in six random fields per retina (white arrows indicate adherent leukocytes). (E) Retinal vascular leakage was measured using Evans blue as a tracer and normalized by total retinal protein concentrations (n = 6; *P
    Figure Legend Snippet: Effects of Y-0452 on retinal vascular inflammation and leakage in diabetic rats. Rats with 1 month of STZ-induced diabetes received daily intraperitoneal injections of Y-0452 (10 mg/kg/d) or the same volume of the vehicle (Veh) as a control for 3 weeks. Retinal vascular endothelial cells and adherent leukocytes were stained with FITC–concanavalin-A and visualized under a fluorescence microscope. (A–C) Representative images of retinal adherent leukocytes in nondiabetic and STZ-induced diabetic rats ([A] nondiabetic, [B] STZ+Veh, [C] STZ+Y-0452). (D) Quantification of retinal adherent leukocytes was performed from ×40 magnification images. Adherent leukocytes were counted in six random fields per retina (white arrows indicate adherent leukocytes). (E) Retinal vascular leakage was measured using Evans blue as a tracer and normalized by total retinal protein concentrations (n = 6; *P

    Techniques Used: Staining, Fluorescence, Microscopy

    24) Product Images from "Glycan heterogeneity on gold nanoparticles increases lectin discrimination capacity in label-free multiplexed bioassays †"

    Article Title: Glycan heterogeneity on gold nanoparticles increases lectin discrimination capacity in label-free multiplexed bioassays †

    Journal: The Analyst

    doi: 10.1039/c6an00549g

    (A) The LDA model generated to discriminate between each lectin, Con A, RCA 120  and SBA. In the model each point represents a sample of that lectin and the ellipse represents one standard deviation from the average. (B) Correct reassignment percentages of each lectin. (C) The LDA model produced to discriminate between both lectin and concentration. Each points represents a sample of that lectin binding to all of the nanoparticles and the ellipse represents one standard deviation from the average response. (D) The LDA model produced for only those samples where the lectin concentration was  > 100 nM.
    Figure Legend Snippet: (A) The LDA model generated to discriminate between each lectin, Con A, RCA 120 and SBA. In the model each point represents a sample of that lectin and the ellipse represents one standard deviation from the average. (B) Correct reassignment percentages of each lectin. (C) The LDA model produced to discriminate between both lectin and concentration. Each points represents a sample of that lectin binding to all of the nanoparticles and the ellipse represents one standard deviation from the average response. (D) The LDA model produced for only those samples where the lectin concentration was > 100 nM.

    Techniques Used: Generated, Standard Deviation, Produced, Concentration Assay, Binding Assay

    Analysis of lectin–glycoAuNP interactions. (A) Photograph of 96-well plate (background removed) with 11 different glycosylated gold nanoparticles from 100% mannosamine functionality to 100% galactose functionality after incubation with a serial dilution of SBA. Dose-dependent binding isotherms of each glycoAuNP with (B) SBA (C) RCA 120  (D) Con A.
    Figure Legend Snippet: Analysis of lectin–glycoAuNP interactions. (A) Photograph of 96-well plate (background removed) with 11 different glycosylated gold nanoparticles from 100% mannosamine functionality to 100% galactose functionality after incubation with a serial dilution of SBA. Dose-dependent binding isotherms of each glycoAuNP with (B) SBA (C) RCA 120 (D) Con A.

    Techniques Used: Incubation, Serial Dilution, Binding Assay

    (A) The LDA model generated to discriminate between Con A, RCA 120  and SBA using binding to only 100% mannose and 100% galactose particles. In the model each point represents a sample of that lectin and the ellipse represents one standard deviation from the average. (B) Correct reassignment percentages of each lectin.
    Figure Legend Snippet: (A) The LDA model generated to discriminate between Con A, RCA 120 and SBA using binding to only 100% mannose and 100% galactose particles. In the model each point represents a sample of that lectin and the ellipse represents one standard deviation from the average. (B) Correct reassignment percentages of each lectin.

    Techniques Used: Generated, Binding Assay, Standard Deviation

    25) Product Images from "Core 2 mucin-type O-glycan inhibits EPEC or EHEC O157:H7 invasion into HT-29 epithelial cells"

    Article Title: Core 2 mucin-type O-glycan inhibits EPEC or EHEC O157:H7 invasion into HT-29 epithelial cells

    Journal: Gut Pathogens

    doi: 10.1186/s13099-015-0078-9

    Lectin histochemical staining in HT-29, HT-29-OBN, HT-29-Gal, and HT-29-Gal-OBN cells. HT-29, HT-29-OBN, HT-29-Gal, or HT-29-Gal-OBN cells were stained with FITC-labeled MAA, PNA, DBA, UEA-I, GSAII or ConA lectins and were observed under fluorescence microscopy. ×40 objective
    Figure Legend Snippet: Lectin histochemical staining in HT-29, HT-29-OBN, HT-29-Gal, and HT-29-Gal-OBN cells. HT-29, HT-29-OBN, HT-29-Gal, or HT-29-Gal-OBN cells were stained with FITC-labeled MAA, PNA, DBA, UEA-I, GSAII or ConA lectins and were observed under fluorescence microscopy. ×40 objective

    Techniques Used: Staining, Labeling, Fluorescence, Microscopy

    26) Product Images from "Epstein-Barr virus activates F-box protein FBXO2 to limit viral infectivity by targeting glycoprotein B for degradation"

    Article Title: Epstein-Barr virus activates F-box protein FBXO2 to limit viral infectivity by targeting glycoprotein B for degradation

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1007208

    N -linked high-mannose glycosylation of EBV gB is required for FBXO2 interaction. (A)  Analysis of glycan modifications of gB. gB-SFB purified from CNE2 cells stably expressing the protein was treated with endoglycosidase H (Endo H) or PNGase H or a Deglycosylation Mix before electrophoresis on 8% polyacrylamide gels. The sizes of molecular mass markers (M) are shown in kDa.  (B)  CNE2 cells stably expressing gB-SFB were treated with tunicamycin at different concentrations as indicated overnight before harvest. Con A agarose was used to enrich  N -linked glycoproteins. TCL: total cell lysates.  (C-D)  Inhibition of N-linked glycosylation of gB by tunicamycin abrogates the gB-FBXO2 association. HEK293T cells transfected with plasmids encoding Myc-FBXO2 and gB-SFB were treated with 0.25 μg/mL tunicamycin or DMSO for 24 h, and the cells were harvested and subjected to immunoprecipitation by S-protein agarose  (C)  or anti-Myc agarose  (D) .  (E)  Sequential pull-down of glycosylated gB by GST-FBXO2 followed by Con A agarose pull-down. Eight percent of the TCL of cells stably expressing gB was loaded as input; F-T: 8% of the flow-through.  (F)  Schematic diagram of glycosylation sites on gB and its N-to-Q mutants. The signal peptide (SP), furin cleavage site and transmembrane (TM) domain of gB are indicated.  (G)  Ribbon diagrams of the monomeric EBV gB structure, illustrated with the PyMol program using PDB 3FVC as the template. The glycosylation sites as shown in  (F)  are colored in magenta.  (H-I)  Con A agarose pull-down of gB single  (H)  or multiple  (I)  N-to-Q mutants.  (J-K)  Co-IP of FBXO2 with glycosylation-defective gB mutants. HEK293T cells were co-transfected with Myc-FBXO2 and gB N-to-Q mutants as indicated, and the cell lysates were subjected to immunoprecipitation by S-protein agarose  (J)  or anti-Myc agarose  (K)  and immunoblotted with anti-Myc and anti-FLAG antibodies.
    Figure Legend Snippet: N -linked high-mannose glycosylation of EBV gB is required for FBXO2 interaction. (A) Analysis of glycan modifications of gB. gB-SFB purified from CNE2 cells stably expressing the protein was treated with endoglycosidase H (Endo H) or PNGase H or a Deglycosylation Mix before electrophoresis on 8% polyacrylamide gels. The sizes of molecular mass markers (M) are shown in kDa. (B) CNE2 cells stably expressing gB-SFB were treated with tunicamycin at different concentrations as indicated overnight before harvest. Con A agarose was used to enrich N -linked glycoproteins. TCL: total cell lysates. (C-D) Inhibition of N-linked glycosylation of gB by tunicamycin abrogates the gB-FBXO2 association. HEK293T cells transfected with plasmids encoding Myc-FBXO2 and gB-SFB were treated with 0.25 μg/mL tunicamycin or DMSO for 24 h, and the cells were harvested and subjected to immunoprecipitation by S-protein agarose (C) or anti-Myc agarose (D) . (E) Sequential pull-down of glycosylated gB by GST-FBXO2 followed by Con A agarose pull-down. Eight percent of the TCL of cells stably expressing gB was loaded as input; F-T: 8% of the flow-through. (F) Schematic diagram of glycosylation sites on gB and its N-to-Q mutants. The signal peptide (SP), furin cleavage site and transmembrane (TM) domain of gB are indicated. (G) Ribbon diagrams of the monomeric EBV gB structure, illustrated with the PyMol program using PDB 3FVC as the template. The glycosylation sites as shown in (F) are colored in magenta. (H-I) Con A agarose pull-down of gB single (H) or multiple (I) N-to-Q mutants. (J-K) Co-IP of FBXO2 with glycosylation-defective gB mutants. HEK293T cells were co-transfected with Myc-FBXO2 and gB N-to-Q mutants as indicated, and the cell lysates were subjected to immunoprecipitation by S-protein agarose (J) or anti-Myc agarose (K) and immunoblotted with anti-Myc and anti-FLAG antibodies.

    Techniques Used: Purification, Stable Transfection, Expressing, Electrophoresis, Inhibition, Transfection, Immunoprecipitation, Flow Cytometry, Co-Immunoprecipitation Assay

    FBXO2 ubiquitinates and degrades  N -linked glycosylated gB. (A)  FBXO2 reduces the level of glycosylated gB. HEK293T cells were co-transfected with gB, along with the FL, N-terminal or C-terminal FBXO2 constructs. 48 hrs later, the glycosylated proteins were enriched by Con A agarose pull-down, and the bound proteins and total cell lysates were immunoblotted with anti-FLAG and anti-Myc antibodies.  (B)  FBXO2 did not affect the total or glycosylated levels of the glycosylation-defective 7NQ gB mutant. The experiments were conducted as in  (A) .  (C)  CNE2 cells stably expressing gB were transfected with three siRNAs targeting FBXO2 or a scramble control siRNA, and 72 h later, the cells were harvested and subjected to Con A pull-down and WB.  (D)  Effects of MG132 and bafilomycin A1 on the levels of total and glycosylated gB.  (E)  FBXO2 ubiquitinates gB  in vivo . HEK293T cells were transfected with His-ubiquitin (Ub), gB-SFB and Myc-FBXO2 or empty vector. The cells were treated with or without 20 μM MG132 for 6 h before harvest. 48 hrs post-transfection, the cells were lysed in 6 M guanidine-HCl buffer, the His-tagged ubiquitinated proteins were enriched by Ni-NTA (nickel-nitrilotriacetic acid) pull-down, and the bound proteins were eluted by SDS loading buffer and immunoblotted with antibodies as indicated.  (F)  HEK293T cells stably expressing gB were transfected with empty vector or Myc-FBXO2, and 36 h later the cells were treated with 50 μg/mL cycloheximide (CHX) to block protein synthesis and collected at the indicated time points. (top) Total cell lysates were analyzed by WB; (bottom) the degradation of gB was calculated by determining the relative quantification of gB from WB results. The mean value for vector-transfected cells at 0 h was normalized to a relative protein level of 100% (n = 3).
    Figure Legend Snippet: FBXO2 ubiquitinates and degrades N -linked glycosylated gB. (A) FBXO2 reduces the level of glycosylated gB. HEK293T cells were co-transfected with gB, along with the FL, N-terminal or C-terminal FBXO2 constructs. 48 hrs later, the glycosylated proteins were enriched by Con A agarose pull-down, and the bound proteins and total cell lysates were immunoblotted with anti-FLAG and anti-Myc antibodies. (B) FBXO2 did not affect the total or glycosylated levels of the glycosylation-defective 7NQ gB mutant. The experiments were conducted as in (A) . (C) CNE2 cells stably expressing gB were transfected with three siRNAs targeting FBXO2 or a scramble control siRNA, and 72 h later, the cells were harvested and subjected to Con A pull-down and WB. (D) Effects of MG132 and bafilomycin A1 on the levels of total and glycosylated gB. (E) FBXO2 ubiquitinates gB in vivo . HEK293T cells were transfected with His-ubiquitin (Ub), gB-SFB and Myc-FBXO2 or empty vector. The cells were treated with or without 20 μM MG132 for 6 h before harvest. 48 hrs post-transfection, the cells were lysed in 6 M guanidine-HCl buffer, the His-tagged ubiquitinated proteins were enriched by Ni-NTA (nickel-nitrilotriacetic acid) pull-down, and the bound proteins were eluted by SDS loading buffer and immunoblotted with antibodies as indicated. (F) HEK293T cells stably expressing gB were transfected with empty vector or Myc-FBXO2, and 36 h later the cells were treated with 50 μg/mL cycloheximide (CHX) to block protein synthesis and collected at the indicated time points. (top) Total cell lysates were analyzed by WB; (bottom) the degradation of gB was calculated by determining the relative quantification of gB from WB results. The mean value for vector-transfected cells at 0 h was normalized to a relative protein level of 100% (n = 3).

    Techniques Used: Transfection, Construct, Mutagenesis, Stable Transfection, Expressing, Western Blot, In Vivo, Plasmid Preparation, Blocking Assay

    27) Product Images from "An organophosphonate strategy for functionalizing silicon photonic biosensors"

    Article Title: An organophosphonate strategy for functionalizing silicon photonic biosensors

    Journal: Langmuir

    doi: 10.1021/la2043153

    Binding response of microrings functionalized with  2  and  3  to (a) Con A and (b) RCA 60  . (c) Reproducible binding response of the microrings functionalized with mannose to Con A after urea regeneration. 1. HEPES buffer; 2. 500 nM Con A in HEPES; 3. 8 M
    Figure Legend Snippet: Binding response of microrings functionalized with 2 and 3 to (a) Con A and (b) RCA 60 . (c) Reproducible binding response of the microrings functionalized with mannose to Con A after urea regeneration. 1. HEPES buffer; 2. 500 nM Con A in HEPES; 3. 8 M

    Techniques Used: Binding Assay

    28) Product Images from "In contrast to Western diet, a plant-based, high-fat, low-sugar diet does not exacerbate retinal endothelial injury in streptozotocin-induced diabetes"

    Article Title: In contrast to Western diet, a plant-based, high-fat, low-sugar diet does not exacerbate retinal endothelial injury in streptozotocin-induced diabetes

    Journal: The FASEB Journal

    doi: 10.1096/fj.201900462R

    Retinal leukostasis and endothelial damage in animals on semipurified diets. Normal and STZ-induced diabetic animals were fed the indicated semipurified diets for 2 wk. The diets were designed to be high in fats, mimicking the composition of plant-based and animal-based sources with and without added cholesterol (0.1% of the total diet). At the end of the period, animals were perfused with FITC-conjugated ConA lectin; subsequently, retinal flat mounts were generated for histologic examinations. A ) Quantitation of the number of firmly adhering leukocytes. B ) Composite micrographs of representative sections of retinas from indicated groups. C ) PI-stained injured or dead endothelial cells per retina in STZ-induced diabetic rats and nondiabetic controls fed indicated diets. D ) Representative micrographs showing PI-positive cells in retinal flat mounts. Scale bar, 200 µm. * P
    Figure Legend Snippet: Retinal leukostasis and endothelial damage in animals on semipurified diets. Normal and STZ-induced diabetic animals were fed the indicated semipurified diets for 2 wk. The diets were designed to be high in fats, mimicking the composition of plant-based and animal-based sources with and without added cholesterol (0.1% of the total diet). At the end of the period, animals were perfused with FITC-conjugated ConA lectin; subsequently, retinal flat mounts were generated for histologic examinations. A ) Quantitation of the number of firmly adhering leukocytes. B ) Composite micrographs of representative sections of retinas from indicated groups. C ) PI-stained injured or dead endothelial cells per retina in STZ-induced diabetic rats and nondiabetic controls fed indicated diets. D ) Representative micrographs showing PI-positive cells in retinal flat mounts. Scale bar, 200 µm. * P

    Techniques Used: Generated, Quantitation Assay, Staining

    29) Product Images from "A defucosylated anti-PD-L1 monoclonal antibody 13-mG2a-f exerts antitumor effects in mouse xenograft models of oral squamous cell carcinoma"

    Article Title: A defucosylated anti-PD-L1 monoclonal antibody 13-mG2a-f exerts antitumor effects in mouse xenograft models of oral squamous cell carcinoma

    Journal: Biochemistry and Biophysics Reports

    doi: 10.1016/j.bbrep.2020.100801

    Confirmation of defucosylation of 13-mG 2a -f by enzyme-linked immunosorbent assay (ELISA) using lectins. (A) L 1 Mab-13 and 13-mG 2a -f were immobilized and incubated with biotin-labeled concanavalin A (Con A), followed by peroxidase-conjugated streptavidin. OD 655 was measured as a function of Con A concentration. (B) L 1 Mab-13 and 13-mG 2a -f were immobilized and incubated with biotin-labeled Aleuria aurantia lectin (AAL), followed by peroxidase-conjugated streptavidin. OD 655 was measured as a function of AAL concentration. (C) L 1 Mab-13 and 13-mG 2a -f were immobilized and incubated with biotin-labeled Pholiota squarrosa lectin (PhoSL), followed by peroxidase-conjugated streptavidin. OD 655 was measured as a function of PhoSL concentration.
    Figure Legend Snippet: Confirmation of defucosylation of 13-mG 2a -f by enzyme-linked immunosorbent assay (ELISA) using lectins. (A) L 1 Mab-13 and 13-mG 2a -f were immobilized and incubated with biotin-labeled concanavalin A (Con A), followed by peroxidase-conjugated streptavidin. OD 655 was measured as a function of Con A concentration. (B) L 1 Mab-13 and 13-mG 2a -f were immobilized and incubated with biotin-labeled Aleuria aurantia lectin (AAL), followed by peroxidase-conjugated streptavidin. OD 655 was measured as a function of AAL concentration. (C) L 1 Mab-13 and 13-mG 2a -f were immobilized and incubated with biotin-labeled Pholiota squarrosa lectin (PhoSL), followed by peroxidase-conjugated streptavidin. OD 655 was measured as a function of PhoSL concentration.

    Techniques Used: Enzyme-linked Immunosorbent Assay, Incubation, Labeling, Concentration Assay

    30) Product Images from "Adiponectin Suppresses Pathological Microvessel Formation in Retina Through Modulation of Tumor Necrosis Factor-α Expression"

    Article Title: Adiponectin Suppresses Pathological Microvessel Formation in Retina Through Modulation of Tumor Necrosis Factor-α Expression

    Journal: Circulation research

    doi: 10.1161/CIRCRESAHA.109.194506

    Exacerbated retinal leukostasis in APN-KO mice. A, Representative photographs of retinal vasculature and adherent leukocytes (arrows) labeled with FITC-conjugated Con A lectin in WT (left) and APN-KO (right) mice at P15. B, Quantitative analysis of leukocyte
    Figure Legend Snippet: Exacerbated retinal leukostasis in APN-KO mice. A, Representative photographs of retinal vasculature and adherent leukocytes (arrows) labeled with FITC-conjugated Con A lectin in WT (left) and APN-KO (right) mice at P15. B, Quantitative analysis of leukocyte

    Techniques Used: Mouse Assay, Labeling

    31) Product Images from "Inhibition of Nucleotide Sugar Transport in Trypanosoma brucei Alters Surface Glycosylation *"

    Article Title: Inhibition of Nucleotide Sugar Transport in Trypanosoma brucei Alters Surface Glycosylation *

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M113.453597

    KO of  tbnst4  causes defects in LacNAc- N -glycosylation. A  and  B,  isolated sVSG from WT ( lanes 1  and  4 ) and two dKO clonal cell lines (A6 an d B1,  lanes 2  and  3  or  5  and  6 ) were separated by SDS-PAGE and subjected to Western blotting, which was detected with biotinylated ECL ( A ) or ConA ( B ) lectins with ( right panels ) or without ( left panels ) the corresponding sugar inhibitors for each of lectins and subsequently, reacted with HRP-conjugated streptavidin.  C  and  D , the same procedure as  A  and  B  was applied except that sVSG-depleted cell lysates were used for Western blotting. ECL ( C ) and Con A ( D ) blots are indicated. 1 × 10 6  and 7 × 10 6  cell equivalents per lane were loaded for sVSG and sVSG-depleted cell lysates, respectively. VSG was used as loading controls.
    Figure Legend Snippet: KO of tbnst4 causes defects in LacNAc- N -glycosylation. A and B, isolated sVSG from WT ( lanes 1 and 4 ) and two dKO clonal cell lines (A6 an d B1, lanes 2 and 3 or 5 and 6 ) were separated by SDS-PAGE and subjected to Western blotting, which was detected with biotinylated ECL ( A ) or ConA ( B ) lectins with ( right panels ) or without ( left panels ) the corresponding sugar inhibitors for each of lectins and subsequently, reacted with HRP-conjugated streptavidin. C and D , the same procedure as A and B was applied except that sVSG-depleted cell lysates were used for Western blotting. ECL ( C ) and Con A ( D ) blots are indicated. 1 × 10 6 and 7 × 10 6 cell equivalents per lane were loaded for sVSG and sVSG-depleted cell lysates, respectively. VSG was used as loading controls.

    Techniques Used: Isolation, SDS Page, Western Blot

    32) Product Images from "High-mannose intercellular adhesion molecule-1 enhances CD16+ monocyte adhesion to the endothelium"

    Article Title: High-mannose intercellular adhesion molecule-1 enhances CD16+ monocyte adhesion to the endothelium

    Journal: American Journal of Physiology - Heart and Circulatory Physiology

    doi: 10.1152/ajpheart.00306.2019

    High-mannose (HM)-intercellular adhesion molecule-1 (ICAM-1) enhances CD16 + monocyte adhesion under flow. A : Western blot of ICAM-1 from COS-1 cells transfected with or without human ICAM-1 and grown in the presence or absence of Kifunensine (Kif). B : surface concanavalin A (ConA) reactive epitopes were immunoprecipitated from COS-1 cells grown in the presence or absence of Kif, and ICAM-1 levels were determined by Western blotting. WCL, whole cell lysate. C : COS-1 cells transfected with or without human ICAM-1 and grown in the presence or absence of Kif were subjected to a proximity-ligation assay (PLA) for HM- and α 2,6 -sialylated ICAM-1. Shown are representative images from each time point. Red puncta represents positive PLA signal, and blue staining is DAPI nuclear stain. SNA, Sambucus nigra . D : quantification of PLA signal from COS-1 cells. P ≤ 0.05 compared with wild-type (WT)- Hippeastrum hybrid amaryllis (HHL) signal (*) and WT + Kif SNA signal (#). Data are means ± SE, n = 3 with an average of 2 fields/ n . E : COS-1 cells treated as above were exposed to isolated C16 + and CD16 − monocytes under flow, and adhesion was measured over 2 min. Data are means ± SE, n = 3. Each symbol represents a single experiment with two replicates per experiment. P ≤ 0.05 compared with empty vector (EV) (*) and compared with WT alone by 1-way ANOVA with Tukey’s posttest within monocyte groups ($). F : monocyte rolling velocities from E were calculated as described above. Data are means ± SE, n = 3 or 4. P ≤ 0.05 compared with EV (*) and compared with WT alone by 1-way ANOVA with Tukey’s posttest within monocyte groups ($). G : COS-1 cells were subjected to monocyte adhesion under flow as described above. Before adhesion assay, some cells were treated with the lectins HHL or ConA to block HM/hybrid N -glycans on the cell surface. Data are means ± SE, n = 3 or 4 with two replicates per experiment. P ≤ 0.05 compared with WT alone (*) and WT + Kif (#) by 1-way ANOVA with Tukey’s posttest within monocyte groups. Broken line represents average of nontransfected cells. H : PLA images from WT and EV COS-1 cells subject to PLA for ICAM-1 and HHL from C collected with longer exposure times. I : velocities calculated from G . Data are means ± SE, n = 4. P ≤ 0.05 compared with WT (*) and WT + Kif (#) by 1-way ANOVA and Tukey’s posttest within monocyte groups. Broken lines represent average of nontransfected cells. J : following native monocyte adhesion to COS-1 cells under flow, cells were fixed and stained for CD16 + and CD16 − markers. Data are means ± SE, n = 3. P ≤ 0.05 compared with baseline (*) and WT ($) by 1-way ANOVA with Tukey’s posttest within monocyte groups.
    Figure Legend Snippet: High-mannose (HM)-intercellular adhesion molecule-1 (ICAM-1) enhances CD16 + monocyte adhesion under flow. A : Western blot of ICAM-1 from COS-1 cells transfected with or without human ICAM-1 and grown in the presence or absence of Kifunensine (Kif). B : surface concanavalin A (ConA) reactive epitopes were immunoprecipitated from COS-1 cells grown in the presence or absence of Kif, and ICAM-1 levels were determined by Western blotting. WCL, whole cell lysate. C : COS-1 cells transfected with or without human ICAM-1 and grown in the presence or absence of Kif were subjected to a proximity-ligation assay (PLA) for HM- and α 2,6 -sialylated ICAM-1. Shown are representative images from each time point. Red puncta represents positive PLA signal, and blue staining is DAPI nuclear stain. SNA, Sambucus nigra . D : quantification of PLA signal from COS-1 cells. P ≤ 0.05 compared with wild-type (WT)- Hippeastrum hybrid amaryllis (HHL) signal (*) and WT + Kif SNA signal (#). Data are means ± SE, n = 3 with an average of 2 fields/ n . E : COS-1 cells treated as above were exposed to isolated C16 + and CD16 − monocytes under flow, and adhesion was measured over 2 min. Data are means ± SE, n = 3. Each symbol represents a single experiment with two replicates per experiment. P ≤ 0.05 compared with empty vector (EV) (*) and compared with WT alone by 1-way ANOVA with Tukey’s posttest within monocyte groups ($). F : monocyte rolling velocities from E were calculated as described above. Data are means ± SE, n = 3 or 4. P ≤ 0.05 compared with EV (*) and compared with WT alone by 1-way ANOVA with Tukey’s posttest within monocyte groups ($). G : COS-1 cells were subjected to monocyte adhesion under flow as described above. Before adhesion assay, some cells were treated with the lectins HHL or ConA to block HM/hybrid N -glycans on the cell surface. Data are means ± SE, n = 3 or 4 with two replicates per experiment. P ≤ 0.05 compared with WT alone (*) and WT + Kif (#) by 1-way ANOVA with Tukey’s posttest within monocyte groups. Broken line represents average of nontransfected cells. H : PLA images from WT and EV COS-1 cells subject to PLA for ICAM-1 and HHL from C collected with longer exposure times. I : velocities calculated from G . Data are means ± SE, n = 4. P ≤ 0.05 compared with WT (*) and WT + Kif (#) by 1-way ANOVA and Tukey’s posttest within monocyte groups. Broken lines represent average of nontransfected cells. J : following native monocyte adhesion to COS-1 cells under flow, cells were fixed and stained for CD16 + and CD16 − markers. Data are means ± SE, n = 3. P ≤ 0.05 compared with baseline (*) and WT ($) by 1-way ANOVA with Tukey’s posttest within monocyte groups.

    Techniques Used: Western Blot, Transfection, Immunoprecipitation, Proximity Ligation Assay, Staining, Isolation, Plasmid Preparation, Cell Adhesion Assay, Blocking Assay

    CD16 + monocyte adhesion is dependent on endothelial high-mannose (HM) epitopes and intercellular adhesion molecule-1 (ICAM-1). A : human umbilical vein endothelial cells (HUVECs) were treated with 10 ng/mL TNF-α for 4 h before monocyte adhesion assay. Some cells were pretreated with Kifunensine (Kif) to form HM epitopes on the cell surface, and then monocyte adhesion under flow was measured. Before adhesion assay, some cells were treated with the lectins Hippeastrum hybrid amaryllis (HHL) or concanavalin A (ConA) to block HM/hybrid N -glycans on the cell surface, or with denatured lectins to assess nonspecific binding effects. Each symbol represents the average of an independent experiment. Data are means ± SE, n = 3 or 4. P ≤ 0.05 compared with control (*), TNF-α alone ($), and TNF-α + Kif (#) by 1-way ANOVA with Tukey’s posttest within monocyte groups. B : HUVECs were treated as described, and some cells were treated with anti-ICAM-1 blocking antibodies that recognize domains 1 (D1) or 3 (D3) of ICAM-1 before monocyte adhesion assay. Each symbol represents the average of an independent experiment. Data are means ± SE, n = 3 or 4. P ≤ 0.05 compared with control (*), TNF-α ($), TNF-α + IgG control ( ), TNF-α + Kif (#), and TNF-α + Kif + IgG control (%) by 1-way ANOVA with Tukey’s posttest within monocyte and cell treatment groups.
    Figure Legend Snippet: CD16 + monocyte adhesion is dependent on endothelial high-mannose (HM) epitopes and intercellular adhesion molecule-1 (ICAM-1). A : human umbilical vein endothelial cells (HUVECs) were treated with 10 ng/mL TNF-α for 4 h before monocyte adhesion assay. Some cells were pretreated with Kifunensine (Kif) to form HM epitopes on the cell surface, and then monocyte adhesion under flow was measured. Before adhesion assay, some cells were treated with the lectins Hippeastrum hybrid amaryllis (HHL) or concanavalin A (ConA) to block HM/hybrid N -glycans on the cell surface, or with denatured lectins to assess nonspecific binding effects. Each symbol represents the average of an independent experiment. Data are means ± SE, n = 3 or 4. P ≤ 0.05 compared with control (*), TNF-α alone ($), and TNF-α + Kif (#) by 1-way ANOVA with Tukey’s posttest within monocyte groups. B : HUVECs were treated as described, and some cells were treated with anti-ICAM-1 blocking antibodies that recognize domains 1 (D1) or 3 (D3) of ICAM-1 before monocyte adhesion assay. Each symbol represents the average of an independent experiment. Data are means ± SE, n = 3 or 4. P ≤ 0.05 compared with control (*), TNF-α ($), TNF-α + IgG control ( ), TNF-α + Kif (#), and TNF-α + Kif + IgG control (%) by 1-way ANOVA with Tukey’s posttest within monocyte and cell treatment groups.

    Techniques Used: Cell Adhesion Assay, Blocking Assay, Binding Assay

    TNF-α forms endothelial high-mannose (HM)-intercellular adhesion molecule-1 (ICAM-1) in a time-dependent manner. Human umbilical vein endothelial cells (HUVECs) were treated with 10 ng/mL TNF-α for 0, 4, or 18 h, and either lysates were collected for Western blot analyses or cells were processed for proximity-ligation assay (PLA).  A : representative Western blot for ICAM-1 expression. The 100-kDa band represents the fully glycosylated complex ICAM-1, and the 75-kDa band represents the hypoglycosylated high-mannose ICAM-1. Kif, Kifunensine; Swain, swainsonine.  B : quantification of HM-ICAM-1 as a percentage of total ICAM-1. Data are means ± SE,  n  = 3.  C : HUVECs were treated as above and subject to a PLA for HM/hybrid, HM, α 2,6 -sialylated, and α 2,3 -sialylated ICAM-1. Shown are representative images from each time point. Red puncta represents positive PLA signal, blue staining is DAPI nuclear stain. ConA, concanavalin A; HHL,  Hippeastrum hybrid amaryllis ; SNA,  Sambucus nigra ; MAL-II, maackia amurensis lectin II.  D : PLA staining controls where one PLA reagent was left out ( left , no anti-ICAM-1;  right , no avidin).  E : quantification of PLA puncta. For each replicate, puncta were counted in three fields and averaged. Each symbol represents an independent replicate. Data are means ± SE,  n  = 4. * P  ≤ 0.05 compared with respective time control by 1-way ANOVA with Tukey’s posttest.
    Figure Legend Snippet: TNF-α forms endothelial high-mannose (HM)-intercellular adhesion molecule-1 (ICAM-1) in a time-dependent manner. Human umbilical vein endothelial cells (HUVECs) were treated with 10 ng/mL TNF-α for 0, 4, or 18 h, and either lysates were collected for Western blot analyses or cells were processed for proximity-ligation assay (PLA). A : representative Western blot for ICAM-1 expression. The 100-kDa band represents the fully glycosylated complex ICAM-1, and the 75-kDa band represents the hypoglycosylated high-mannose ICAM-1. Kif, Kifunensine; Swain, swainsonine. B : quantification of HM-ICAM-1 as a percentage of total ICAM-1. Data are means ± SE, n = 3. C : HUVECs were treated as above and subject to a PLA for HM/hybrid, HM, α 2,6 -sialylated, and α 2,3 -sialylated ICAM-1. Shown are representative images from each time point. Red puncta represents positive PLA signal, blue staining is DAPI nuclear stain. ConA, concanavalin A; HHL, Hippeastrum hybrid amaryllis ; SNA, Sambucus nigra ; MAL-II, maackia amurensis lectin II. D : PLA staining controls where one PLA reagent was left out ( left , no anti-ICAM-1; right , no avidin). E : quantification of PLA puncta. For each replicate, puncta were counted in three fields and averaged. Each symbol represents an independent replicate. Data are means ± SE, n = 4. * P ≤ 0.05 compared with respective time control by 1-way ANOVA with Tukey’s posttest.

    Techniques Used: Western Blot, Proximity Ligation Assay, Expressing, Staining, Avidin-Biotin Assay

    33) Product Images from "A defucosylated anti-CD44 monoclonal antibody 5-mG2a-f exerts antitumor effects in mouse xenograft models of oral squamous cell carcinoma"

    Article Title: A defucosylated anti-CD44 monoclonal antibody 5-mG2a-f exerts antitumor effects in mouse xenograft models of oral squamous cell carcinoma

    Journal: Oncology Reports

    doi: 10.3892/or.2020.7735

    Confirmation of defucosylation of 5-mG 2a -f by enzyme-linked immunosorbent assay (ELISA) using lectins. (A) C 44 Mab-5 and 5-mG 2a -f were immobilized and incubated with biotin-labeled concanavalin A (Con A), followed by peroxidase-conjugated streptavidin. The enzymatic reaction was produced using a 1-Step Ultra TMB-ELISA. (B) C 44 Mab-5 and 5-mG 2a -f were immobilized and incubated with biotin-labeled Aleuria aurantia lectin (AAL), followed by peroxidase-conjugated streptavidin. The enzymatic reaction was produced using a 1-Step Ultra TMB-ELISA. (C) C 44 Mab-5 and 5-mG 2a -f were immobilized and incubated with biotin-labeled Pholiota squarrosa lectin (PhoSL), followed by peroxidase-conjugated streptavidin. The enzymatic reaction was produced using a 1-Step Ultra TMB-ELISA.
    Figure Legend Snippet: Confirmation of defucosylation of 5-mG 2a -f by enzyme-linked immunosorbent assay (ELISA) using lectins. (A) C 44 Mab-5 and 5-mG 2a -f were immobilized and incubated with biotin-labeled concanavalin A (Con A), followed by peroxidase-conjugated streptavidin. The enzymatic reaction was produced using a 1-Step Ultra TMB-ELISA. (B) C 44 Mab-5 and 5-mG 2a -f were immobilized and incubated with biotin-labeled Aleuria aurantia lectin (AAL), followed by peroxidase-conjugated streptavidin. The enzymatic reaction was produced using a 1-Step Ultra TMB-ELISA. (C) C 44 Mab-5 and 5-mG 2a -f were immobilized and incubated with biotin-labeled Pholiota squarrosa lectin (PhoSL), followed by peroxidase-conjugated streptavidin. The enzymatic reaction was produced using a 1-Step Ultra TMB-ELISA.

    Techniques Used: Enzyme-linked Immunosorbent Assay, Incubation, Labeling, Produced

    34) Product Images from "Enterococcus hirae biofilm formation on hospital material surfaces and effect of new biocides"

    Article Title: Enterococcus hirae biofilm formation on hospital material surfaces and effect of new biocides

    Journal: Environmental Health and Preventive Medicine

    doi: 10.1186/s12199-017-0670-3

    Optical microscope fluorescence representative images of E. hirae ATCC 10541 biofilms at 48 h on polystyrene at 20 °C ( a ) and 37 °C ( b ) on stainless steel coupon at 20 °C ( c ) and 37 °C ( d ) by Con-A staining for detecting α glucans in matrix formation. Original magnification ×1000
    Figure Legend Snippet: Optical microscope fluorescence representative images of E. hirae ATCC 10541 biofilms at 48 h on polystyrene at 20 °C ( a ) and 37 °C ( b ) on stainless steel coupon at 20 °C ( c ) and 37 °C ( d ) by Con-A staining for detecting α glucans in matrix formation. Original magnification ×1000

    Techniques Used: Microscopy, Fluorescence, Staining

    35) Product Images from "GnT1IP-L specifically inhibits MGAT1 in the Golgi via its luminal domain"

    Article Title: GnT1IP-L specifically inhibits MGAT1 in the Golgi via its luminal domain

    Journal: eLife

    doi: 10.7554/eLife.08916

    The TM and cytoplasmic domain of GnT1IP-L does not inhibit MGAT1. ( A ) Lectin-resistance test of cloned CHO cells stably expressing GnT1IP-L/MGAT1-Myc compared to CHO cells and Lec1 CHO cells that lack MGAT1 (n = 2). ( B ) The same cloned GnT1IP-L transfectant lines were compared to CHO and Lec1 cells for resistance to Con A (n = 2). ( C ) Western analysis of CHO cell lysates from the cloned transfectants in ( A ) and ( B ). * non-specific band shows equal loading. DOI: http://dx.doi.org/10.7554/eLife.08916.007
    Figure Legend Snippet: The TM and cytoplasmic domain of GnT1IP-L does not inhibit MGAT1. ( A ) Lectin-resistance test of cloned CHO cells stably expressing GnT1IP-L/MGAT1-Myc compared to CHO cells and Lec1 CHO cells that lack MGAT1 (n = 2). ( B ) The same cloned GnT1IP-L transfectant lines were compared to CHO and Lec1 cells for resistance to Con A (n = 2). ( C ) Western analysis of CHO cell lysates from the cloned transfectants in ( A ) and ( B ). * non-specific band shows equal loading. DOI: http://dx.doi.org/10.7554/eLife.08916.007

    Techniques Used: Clone Assay, Stable Transfection, Expressing, Transfection, Western Blot

    36) Product Images from "N-Glycosylation Regulates the Trafficking and Surface Mobility of GluN3A-Containing NMDA Receptors"

    Article Title: N-Glycosylation Regulates the Trafficking and Surface Mobility of GluN3A-Containing NMDA Receptors

    Journal: Frontiers in Molecular Neuroscience

    doi: 10.3389/fnmol.2018.00188

    N -glycosylation regulates the mobility of GluN3A-containing NMDARs at the cell surface of hippocampal neurons. (A,C) Representative trajectories of quantum dot (QD)-labeled GFP-GluN3A (GluN3A) subunits in hippocampal neurons treated with 200 μg/ml DMM (A) or with the lectin AAL (20 μg/ml) (C) . (B,D) Comparison of the cumulative distribution of the diffusion coefficients measured for QD-labeled GluN3A subunits in hippocampal neurons treated for two days with 200 μg/ml DMM, 100 μg/ml swainsonine ( n = 208–314 trajectories per condition; B ), or incubated after QD labeling with 20 μg/ml of AAL, conA, or WGA ( n = 84–131 trajectories per condition; D ). The diffusion coefficients in fixed cells (shown in D ) were obtained from an independent experiment ( n = 49 trajectories). Pairwise comparisons of the distributions show the following statistically significant differences: DMM vs. control ( p
    Figure Legend Snippet: N -glycosylation regulates the mobility of GluN3A-containing NMDARs at the cell surface of hippocampal neurons. (A,C) Representative trajectories of quantum dot (QD)-labeled GFP-GluN3A (GluN3A) subunits in hippocampal neurons treated with 200 μg/ml DMM (A) or with the lectin AAL (20 μg/ml) (C) . (B,D) Comparison of the cumulative distribution of the diffusion coefficients measured for QD-labeled GluN3A subunits in hippocampal neurons treated for two days with 200 μg/ml DMM, 100 μg/ml swainsonine ( n = 208–314 trajectories per condition; B ), or incubated after QD labeling with 20 μg/ml of AAL, conA, or WGA ( n = 84–131 trajectories per condition; D ). The diffusion coefficients in fixed cells (shown in D ) were obtained from an independent experiment ( n = 49 trajectories). Pairwise comparisons of the distributions show the following statistically significant differences: DMM vs. control ( p

    Techniques Used: Labeling, Diffusion-based Assay, Incubation, Whole Genome Amplification

    37) Product Images from "Signal Peptide-Binding Drug as a Selective Inhibitor of Co-Translational Protein Translocation"

    Article Title: Signal Peptide-Binding Drug as a Selective Inhibitor of Co-Translational Protein Translocation

    Journal: PLoS Biology

    doi: 10.1371/journal.pbio.1002011

    CADA specifically inhibits the biogenesis of human CD4. (A, B) CADA inhibits the biosynthesis of CD4. CD4 + .CHO cells were washed and kept in methionine and cysteine-free medium in the presence or absence of 16 µM CADA for 45 min before exposure to [ 35 S]methionine/cysteine (Met/Cys) for 30 min. Pulsed-labelled cells were then washed, lysed, and analyzed directly (A) or incubated in normal medium for up to 4 h (chase) in the presence or absence of 16 µM CADA (B). At specified time points cell lysates were immunoprecipitated for CD4. The flow through fraction (FT) of the CD4-immunoprecipitated samples is also presented. Note that the weaker CD4 bands in the control samples at longer chase time points are the result of the high turnover of hCD4 in CHO cells. Molecular mass is in kDa. (C–F) CD4 negative and stably CD4-YFP transfected CHO cells were pretreated with CADA (5 µM) or DMSO for 1 h before starvation in Met/Cys free medium with CADA, DMSO, or 50 µg/ml CHX. Cells were pulsed for 30 min, washed, and incubated in fresh medium without serum for 90 min. After collection of supernatant proteins (Media) cells were first permeabilized with digitonin buffer to obtain the cytosolic cell fraction before lysis in NP-40 buffer to collect the membrane proteins. Membrane fractions were further incubated with Concanavalin A (ConA) agarose beads (Glycosylated). Molecular mass is in kDa. (D) Quantification of 35 S incorporation in (C) by scintillation counting ( n = 4). NS, not significant; * p
    Figure Legend Snippet: CADA specifically inhibits the biogenesis of human CD4. (A, B) CADA inhibits the biosynthesis of CD4. CD4 + .CHO cells were washed and kept in methionine and cysteine-free medium in the presence or absence of 16 µM CADA for 45 min before exposure to [ 35 S]methionine/cysteine (Met/Cys) for 30 min. Pulsed-labelled cells were then washed, lysed, and analyzed directly (A) or incubated in normal medium for up to 4 h (chase) in the presence or absence of 16 µM CADA (B). At specified time points cell lysates were immunoprecipitated for CD4. The flow through fraction (FT) of the CD4-immunoprecipitated samples is also presented. Note that the weaker CD4 bands in the control samples at longer chase time points are the result of the high turnover of hCD4 in CHO cells. Molecular mass is in kDa. (C–F) CD4 negative and stably CD4-YFP transfected CHO cells were pretreated with CADA (5 µM) or DMSO for 1 h before starvation in Met/Cys free medium with CADA, DMSO, or 50 µg/ml CHX. Cells were pulsed for 30 min, washed, and incubated in fresh medium without serum for 90 min. After collection of supernatant proteins (Media) cells were first permeabilized with digitonin buffer to obtain the cytosolic cell fraction before lysis in NP-40 buffer to collect the membrane proteins. Membrane fractions were further incubated with Concanavalin A (ConA) agarose beads (Glycosylated). Molecular mass is in kDa. (D) Quantification of 35 S incorporation in (C) by scintillation counting ( n = 4). NS, not significant; * p

    Techniques Used: Incubation, Immunoprecipitation, Flow Cytometry, Stable Transfection, Transfection, Lysis

    38) Product Images from "Capture of microtubule plus-ends at the actin cortex promotes axophilic neuronal migration by enhancing microtubule tension in the leading process"

    Article Title: Capture of microtubule plus-ends at the actin cortex promotes axophilic neuronal migration by enhancing microtubule tension in the leading process

    Journal: Frontiers in Cellular Neuroscience

    doi: 10.3389/fncel.2014.00400

    Microtubule +TIPs lose association with cortical actin after blocking calcium release . (A) Triple-staining against EB1 (green), F-actin (red), and GnRH (blue) in a control-treated cell with EB1 and F-actin imaged with STED microscopy. (B) Zoomed image of the box in (A) . Arrows indicate examples of super-resolution co-localization, showing several EB1 puncta associated with the actin cortex in the leading process. (C,D) Low (C) and high magnification ( D , of region boxed in C ) of triple-stained neurons treated with 2-APB. Blocking calcium release reduces the number of EB1 puncta associated with the actin cortex (arrows, D ). Scale bars, 5 μm. (E) No differences were detected in raw EB1 staining fluorescence (n.s., p = 0.227, t -test, Cohen's d = 0.53, n = 10 control and 12 2-APB treated GnRH neurons). (F) Fewer EB1 puncta were observed in the leading process actin cortex of 2-APB treated GnRH neurons compared with vehicle controls ( *** p = 0.0006, t -test, Cohen's d = 1.75, n = 10 controls and 12 treated GnRH neurons, N = 3 explants for both conditions). (G) Fraction of time microtubules spent moving toward the distal growth cone was reduced by the inhibitor of cortical actin flow, Concanavalin A ( n = 7 cells from N = 3 explants; ** p = 0.0026, Fisher's exact test).
    Figure Legend Snippet: Microtubule +TIPs lose association with cortical actin after blocking calcium release . (A) Triple-staining against EB1 (green), F-actin (red), and GnRH (blue) in a control-treated cell with EB1 and F-actin imaged with STED microscopy. (B) Zoomed image of the box in (A) . Arrows indicate examples of super-resolution co-localization, showing several EB1 puncta associated with the actin cortex in the leading process. (C,D) Low (C) and high magnification ( D , of region boxed in C ) of triple-stained neurons treated with 2-APB. Blocking calcium release reduces the number of EB1 puncta associated with the actin cortex (arrows, D ). Scale bars, 5 μm. (E) No differences were detected in raw EB1 staining fluorescence (n.s., p = 0.227, t -test, Cohen's d = 0.53, n = 10 control and 12 2-APB treated GnRH neurons). (F) Fewer EB1 puncta were observed in the leading process actin cortex of 2-APB treated GnRH neurons compared with vehicle controls ( *** p = 0.0006, t -test, Cohen's d = 1.75, n = 10 controls and 12 treated GnRH neurons, N = 3 explants for both conditions). (G) Fraction of time microtubules spent moving toward the distal growth cone was reduced by the inhibitor of cortical actin flow, Concanavalin A ( n = 7 cells from N = 3 explants; ** p = 0.0026, Fisher's exact test).

    Techniques Used: Blocking Assay, Staining, Microscopy, Fluorescence, Flow Cytometry

    39) Product Images from "Antiangiogenic and Antineuroinflammatory Effects of Kallistatin Through Interactions With the Canonical Wnt Pathway"

    Article Title: Antiangiogenic and Antineuroinflammatory Effects of Kallistatin Through Interactions With the Canonical Wnt Pathway

    Journal: Diabetes

    doi: 10.2337/db12-1710

    Decreased retinal neuroinflammation and retinal vascular leakage in diabetic kallistatin-TG mice. Akita mice and Akita×kallistatin-TG mice at 5 months of age were used for leukostasis assay. A : Adherent leukocytes (arrow) were stained with FITC-conjugated concanavalin-A in the retinal vasculature. The retinal vasculature and leukocytes were visualized in retinal flat mounts under a fluorescence microscope. Scale bar: 50 μm. B : Adherent leukocytes in the retinal vasculature were counted in age-matched WT, kallistatin-TG, Akita, and Akita×kallistatin-TG mice. n = 7–10. C : Retinal levels of soluble ICAM-1 were measured by ELISA in age-matched WT, kallistatin-TG, Akita, and Akita×kallistatin-TG mice and expressed as percentages of the respective WT control. D : Retinal vascular permeability of WT, kallistatin-TG, Akita, and Akita×kallistatin-TG mice was measured using Evans blue as a tracer, normalized by retinal protein concentrations, and expressed as a percentage of the permeability in WT control. E : Retinal levels of VEGF-A were measured by ELISA in WT, kallistatin-TG, Akita, and Akita×kallistatin-TG mice and expressed as percentages of the respective WT control. All values are mean ± SD. * P
    Figure Legend Snippet: Decreased retinal neuroinflammation and retinal vascular leakage in diabetic kallistatin-TG mice. Akita mice and Akita×kallistatin-TG mice at 5 months of age were used for leukostasis assay. A : Adherent leukocytes (arrow) were stained with FITC-conjugated concanavalin-A in the retinal vasculature. The retinal vasculature and leukocytes were visualized in retinal flat mounts under a fluorescence microscope. Scale bar: 50 μm. B : Adherent leukocytes in the retinal vasculature were counted in age-matched WT, kallistatin-TG, Akita, and Akita×kallistatin-TG mice. n = 7–10. C : Retinal levels of soluble ICAM-1 were measured by ELISA in age-matched WT, kallistatin-TG, Akita, and Akita×kallistatin-TG mice and expressed as percentages of the respective WT control. D : Retinal vascular permeability of WT, kallistatin-TG, Akita, and Akita×kallistatin-TG mice was measured using Evans blue as a tracer, normalized by retinal protein concentrations, and expressed as a percentage of the permeability in WT control. E : Retinal levels of VEGF-A were measured by ELISA in WT, kallistatin-TG, Akita, and Akita×kallistatin-TG mice and expressed as percentages of the respective WT control. All values are mean ± SD. * P

    Techniques Used: Mouse Assay, Staining, Fluorescence, Microscopy, Enzyme-linked Immunosorbent Assay, Permeability

    Related Articles

    Whole Genome Amplification:

    Article Title: Transcriptomic analyses reveal comprehensive responses of insect hemocytes to mycopathogen Beauveria bassiana, and fungal virulence-related cell wall protein assists pathogen to evade host cellular defense
    Article Snippet: .. Lectins included concanavalin A (ConA), Galanthus nivalis lectin (GNL), peanut agglutinin (PNA), and wheat germ agglutinin (WGA), and were purchased from Vector Laboratories, Inc. (Burlingame, California, USA). ..

    Incubation:

    Article Title: The Metabolic Chemical Reporter 6‑Azido-6-deoxy-glucose Further Reveals the Substrate Promiscuity of O‑GlcNAc Transferase and Catalyzes the Discovery of Intracellular Protein Modification by O‑Glucose
    Article Snippet: .. The blots washed three times in TBST for 5 min and incubated with biotin-conjugated Concanavalin A (Vector Lab), diluted 1:1000 in TBST, for 1 h. The blot was then washed 3× with TBST for 10 min. Then the blot was then incubated with Strep-HRP at 1:1000 in blocking buffer for 1 h. After being washed 3× with TBST for 10 min, the blot was developed using ECL reagents. .. BL21(DE3) chemically competent E. coli (Novagen) were transformed with a pET24b plasmid encoding 6His-tagged ncOGT (nucleocytoplasmic OGT) by heat shock and plated on selective LB agar plates containing 50 μ g mL−1 kanamycin (LB-kan).

    Article Title: Glycan Profiling Shows Unvaried N-Glycomes in MSC Clones with Distinct Differentiation Potentials
    Article Snippet: .. Following 15 min incubation on ice, 120 μL of 10 μg/mL FITC-ConA (Vector Labs) was added to each sample. .. After two 15 min incubations interspersed with flicking of the tubes, 1 mL of washing buffer was added and the samples were centrifuged for 5 min at 450 g. The cell pellet was resuspended in 100 μL of washing buffer containing 1 μg/mL DAPI and incubated on ice in the dark for 5 min, followed by the addition of 1 mL washing buffer, and centrifugation for 5 min at 450 g. The pellet was resuspended in 400 μL PBS for flow cytometry.

    Blocking Assay:

    Article Title: The Metabolic Chemical Reporter 6‑Azido-6-deoxy-glucose Further Reveals the Substrate Promiscuity of O‑GlcNAc Transferase and Catalyzes the Discovery of Intracellular Protein Modification by O‑Glucose
    Article Snippet: .. The blots washed three times in TBST for 5 min and incubated with biotin-conjugated Concanavalin A (Vector Lab), diluted 1:1000 in TBST, for 1 h. The blot was then washed 3× with TBST for 10 min. Then the blot was then incubated with Strep-HRP at 1:1000 in blocking buffer for 1 h. After being washed 3× with TBST for 10 min, the blot was developed using ECL reagents. .. BL21(DE3) chemically competent E. coli (Novagen) were transformed with a pET24b plasmid encoding 6His-tagged ncOGT (nucleocytoplasmic OGT) by heat shock and plated on selective LB agar plates containing 50 μ g mL−1 kanamycin (LB-kan).

    Staining:

    Article Title: Ischaemia-induced retinal neovascularisation and diabetic retinopathy in mice with conditional knockout of hypoxia-inducible factor-1 in retinal Müller cells
    Article Snippet: Briefly, mice were perfused through the left ventricle to remove circulating leucocytes in blood vessels. .. The adherent leucocytes in the vasculature were stained by perfusion with FITC-conjugated concanavalin-A (40 μg/ml; Vector Laboratories) and counted. .. The quantitative data were analysed and compared with those from wild-type mice using unpaired Student’s t test (two-tailed test).

    Labeling:

    Article Title: Arginase as a Mediator of Diabetic Retinopathy
    Article Snippet: The images were analyzed for reaction intensity by using the MetaMorph Image System (Molecular Devices). .. Leukocyte adhesion Retinal leukostasis was assayed by labeling the adherent leukocytes using Concanavalin A (Vector Laboratories). .. TUNEL assay Endothelial cell death was studied using TUNEL (Terminal deoxynucleotidyl transferase dUTP nick end labeling) assay using Fluorescein in situ cell death detection kit (Millipore) according to the manufacturer’s protocol.

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    Vector Laboratories biotin conjugated concanavalin a
    A significant fraction of 6AzGlc-dependent labeling is O-linked. (A) Known O -GlcNAcylated proteins are labeled by 6AzGlc. H1299 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO for 16 h, followed by CuAAC with a cleavable alkyne-biotin tag. After enrichment on streptavidin beads, the labeled proteins were eluted and visualized by Western blotting. The nonglycosylated protein β -actin is a negative control. (B) A notable fraction of 6AzGlc-dependent signal is sensitive to β -elimination. NIH3T3 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO vehicle for 16 h, followed by CuAAC with alkyne-biotin, separation by SDS-PAGE and transfer to a PVDF membrane. The indicated membranes were then treated for 24 h with either H 2 O or 55 mM NaOH before analysis by streptavidin or Western blotting. (C) 6AzGlc is not incorporated into N-linked glycans. NIH3T3 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO vehicle for 16 h. The corresponding cell lysates were then incubated with either PNGase-F or H 2 O vehicle as indicated before CuAAC with alkyne TAMRA and analysis by in-gel fluorescence. A fraction of the treated lysate was separated before CuAAC and analyzed by Lectin blotting with <t>Concanavalin</t> A (ConA).
    Biotin Conjugated Concanavalin A, 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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    A significant fraction of 6AzGlc-dependent labeling is O-linked. (A) Known O -GlcNAcylated proteins are labeled by 6AzGlc. H1299 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO for 16 h, followed by CuAAC with a cleavable alkyne-biotin tag. After enrichment on streptavidin beads, the labeled proteins were eluted and visualized by Western blotting. The nonglycosylated protein β -actin is a negative control. (B) A notable fraction of 6AzGlc-dependent signal is sensitive to β -elimination. NIH3T3 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO vehicle for 16 h, followed by CuAAC with alkyne-biotin, separation by SDS-PAGE and transfer to a PVDF membrane. The indicated membranes were then treated for 24 h with either H 2 O or 55 mM NaOH before analysis by streptavidin or Western blotting. (C) 6AzGlc is not incorporated into N-linked glycans. NIH3T3 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO vehicle for 16 h. The corresponding cell lysates were then incubated with either PNGase-F or H 2 O vehicle as indicated before CuAAC with alkyne TAMRA and analysis by in-gel fluorescence. A fraction of the treated lysate was separated before CuAAC and analyzed by Lectin blotting with Concanavalin A (ConA).

    Journal: Journal of the American Chemical Society

    Article Title: The Metabolic Chemical Reporter 6‑Azido-6-deoxy-glucose Further Reveals the Substrate Promiscuity of O‑GlcNAc Transferase and Catalyzes the Discovery of Intracellular Protein Modification by O‑Glucose

    doi: 10.1021/jacs.7b13488

    Figure Lengend Snippet: A significant fraction of 6AzGlc-dependent labeling is O-linked. (A) Known O -GlcNAcylated proteins are labeled by 6AzGlc. H1299 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO for 16 h, followed by CuAAC with a cleavable alkyne-biotin tag. After enrichment on streptavidin beads, the labeled proteins were eluted and visualized by Western blotting. The nonglycosylated protein β -actin is a negative control. (B) A notable fraction of 6AzGlc-dependent signal is sensitive to β -elimination. NIH3T3 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO vehicle for 16 h, followed by CuAAC with alkyne-biotin, separation by SDS-PAGE and transfer to a PVDF membrane. The indicated membranes were then treated for 24 h with either H 2 O or 55 mM NaOH before analysis by streptavidin or Western blotting. (C) 6AzGlc is not incorporated into N-linked glycans. NIH3T3 cells were treated with either Ac 4 6AzGlc (200 μ M) or DMSO vehicle for 16 h. The corresponding cell lysates were then incubated with either PNGase-F or H 2 O vehicle as indicated before CuAAC with alkyne TAMRA and analysis by in-gel fluorescence. A fraction of the treated lysate was separated before CuAAC and analyzed by Lectin blotting with Concanavalin A (ConA).

    Article Snippet: The blots washed three times in TBST for 5 min and incubated with biotin-conjugated Concanavalin A (Vector Lab), diluted 1:1000 in TBST, for 1 h. The blot was then washed 3× with TBST for 10 min. Then the blot was then incubated with Strep-HRP at 1:1000 in blocking buffer for 1 h. After being washed 3× with TBST for 10 min, the blot was developed using ECL reagents.

    Techniques: Labeling, Western Blot, Negative Control, SDS Page, Incubation, Fluorescence

    N -glycan profile of MSCs does not correlate with differentiation capacity. (A) Osteogenic capacities of Y101 and Y202 hTERT-MSC lines. For details see Figure 2A . (B) N -glycan profiles of the Y101 and Y202 hTERT-MSC lines following 24 h culture in basal medium. Representative MALDI-TOF spectra as in Figure 1 , but glycan assignments are omitted for clarity. (C) Heat map display comparing the N -glycan profiles of Y101, Y202 hTERT-MSCs, and osteoblasts derived from the Y101 hTERT-MSCs. Averaged (Y101, Y202 n = 5, osteoblasts n = 3) normalized peak intensities were compared. Each row in the heat map display represents a single glycan structure. Glycans are grouped by type: oligomannose (O), hybrid (H), or complex (C), and sorted by abundance in the Y101 profile within each type. The cells' colors denote glycan abundance as indicated in the legend. (D) Y101 or Y202 cells were grown for 0 or 8 days in basal or osteogenic (osteo) medium, then single cell suspensions stained with ConA-FITC and DAPI and analyzed by flow cytometry. The median intensity of the FITC fluorescence of live cells is shown with SEM from three independent replicates. (E) Y101 cells were cultured in basal medium containing 10 μg/mL swainsonine for 48 h before addition of osteogenic medium containing swainonsine at the same concentration. Mineral deposition and ALP activity were tested by ALP/von Kossa staining 0, 7, 14, and 21 days after addition of osteogenic medium. *** P

    Journal: Frontiers in Cell and Developmental Biology

    Article Title: Glycan Profiling Shows Unvaried N-Glycomes in MSC Clones with Distinct Differentiation Potentials

    doi: 10.3389/fcell.2016.00052

    Figure Lengend Snippet: N -glycan profile of MSCs does not correlate with differentiation capacity. (A) Osteogenic capacities of Y101 and Y202 hTERT-MSC lines. For details see Figure 2A . (B) N -glycan profiles of the Y101 and Y202 hTERT-MSC lines following 24 h culture in basal medium. Representative MALDI-TOF spectra as in Figure 1 , but glycan assignments are omitted for clarity. (C) Heat map display comparing the N -glycan profiles of Y101, Y202 hTERT-MSCs, and osteoblasts derived from the Y101 hTERT-MSCs. Averaged (Y101, Y202 n = 5, osteoblasts n = 3) normalized peak intensities were compared. Each row in the heat map display represents a single glycan structure. Glycans are grouped by type: oligomannose (O), hybrid (H), or complex (C), and sorted by abundance in the Y101 profile within each type. The cells' colors denote glycan abundance as indicated in the legend. (D) Y101 or Y202 cells were grown for 0 or 8 days in basal or osteogenic (osteo) medium, then single cell suspensions stained with ConA-FITC and DAPI and analyzed by flow cytometry. The median intensity of the FITC fluorescence of live cells is shown with SEM from three independent replicates. (E) Y101 cells were cultured in basal medium containing 10 μg/mL swainsonine for 48 h before addition of osteogenic medium containing swainonsine at the same concentration. Mineral deposition and ALP activity were tested by ALP/von Kossa staining 0, 7, 14, and 21 days after addition of osteogenic medium. *** P

    Article Snippet: Following 15 min incubation on ice, 120 μL of 10 μg/mL FITC-ConA (Vector Labs) was added to each sample.

    Techniques: Derivative Assay, Staining, Flow Cytometry, Cytometry, Fluorescence, Cell Culture, Concentration Assay, ALP Assay, Activity Assay

    Prevention of diabetes-induced increase in leukocyte adhesion by arginase deletion . Wild type controls, wild type diabetic or arginase AI +/− AII −/− (KO) diabetic mice were perfused through left ventricle with Concanavalin A to label leukocytes attached to the vascular endothelium. The number of attached leukocytes was significantly increased in the wildtype diabetic mice as compared to non-diabetic controls and arginase KO significantly blunted this effect (* p

    Journal: Frontiers in Immunology

    Article Title: Arginase as a Mediator of Diabetic Retinopathy

    doi: 10.3389/fimmu.2013.00173

    Figure Lengend Snippet: Prevention of diabetes-induced increase in leukocyte adhesion by arginase deletion . Wild type controls, wild type diabetic or arginase AI +/− AII −/− (KO) diabetic mice were perfused through left ventricle with Concanavalin A to label leukocytes attached to the vascular endothelium. The number of attached leukocytes was significantly increased in the wildtype diabetic mice as compared to non-diabetic controls and arginase KO significantly blunted this effect (* p

    Article Snippet: Leukocyte adhesion Retinal leukostasis was assayed by labeling the adherent leukocytes using Concanavalin A (Vector Laboratories).

    Techniques: Mouse Assay

    Morniga-G activates healthy human lymphocytes and induces cell death in leukemia cells. PBMCs from healthy donors were cultured for 3 days: ( A ) in the presence of increasing concentrations of Morniga-M (MorM), Morniga-G (MorG), and Con A and proliferative index (from the [ 3 H]-thymidine incorporation) was calculated; and ( B ) in the presence of lectins at concentrations triggering maximal proliferation. CD25 expression was evaluated using flow cytometry, in CD3+ T lymphocytes, in CD19+ CD3- B lymphocytes, and in CD56+ CD3- NK lymphocytes. Values are means ± SD of three experiments performed with three to four different healthy donors. ( C ) Resting PBMCs and Jurkat A3 leukemic cells were incubated with anti-Tn mouse monoclonal antibody + PE-conjugated anti-mouse antibody (αTn + antiα-PE) or FITC-conjugated Morniga G (MorG-FITC) and analyzed using cytofluorimetry. Healthy peripheral lymphocytes (PBLs) were analyzed in a gate corresponding to lymphocytes as defined by the size and granularity parameters. Autofluorescence: purple histograms, dashed green histogram: fluorescent positive cells. ( D ) PBMCs of healthy donors and Jurkat A3 leukemic cells were cultured for 24 h with different concentrations of MorG, then cell viability was evaluated in 3-(4,5-dimethylthiazol)-2-5-diphenyl terazolium bromide MTT reduction assays (mean values ± SD of four independent experiments).

    Journal: International Journal of Molecular Sciences

    Article Title: Morniga-G, a T/Tn-Specific Lectin, Induces Leukemic Cell Death via Caspase and DR5 Receptor-Dependent Pathways

    doi: 10.3390/ijms20010230

    Figure Lengend Snippet: Morniga-G activates healthy human lymphocytes and induces cell death in leukemia cells. PBMCs from healthy donors were cultured for 3 days: ( A ) in the presence of increasing concentrations of Morniga-M (MorM), Morniga-G (MorG), and Con A and proliferative index (from the [ 3 H]-thymidine incorporation) was calculated; and ( B ) in the presence of lectins at concentrations triggering maximal proliferation. CD25 expression was evaluated using flow cytometry, in CD3+ T lymphocytes, in CD19+ CD3- B lymphocytes, and in CD56+ CD3- NK lymphocytes. Values are means ± SD of three experiments performed with three to four different healthy donors. ( C ) Resting PBMCs and Jurkat A3 leukemic cells were incubated with anti-Tn mouse monoclonal antibody + PE-conjugated anti-mouse antibody (αTn + antiα-PE) or FITC-conjugated Morniga G (MorG-FITC) and analyzed using cytofluorimetry. Healthy peripheral lymphocytes (PBLs) were analyzed in a gate corresponding to lymphocytes as defined by the size and granularity parameters. Autofluorescence: purple histograms, dashed green histogram: fluorescent positive cells. ( D ) PBMCs of healthy donors and Jurkat A3 leukemic cells were cultured for 24 h with different concentrations of MorG, then cell viability was evaluated in 3-(4,5-dimethylthiazol)-2-5-diphenyl terazolium bromide MTT reduction assays (mean values ± SD of four independent experiments).

    Article Snippet: Concanavalin A (Con A) was purchased from Vector Laboratories (Burlingame, CA, USA).

    Techniques: Cell Culture, Expressing, Flow Cytometry, Cytometry, Incubation, MTT Assay