Sambucus nigra

Manufactured by Vector Laboratories

Sourced in United States

Sambucus nigra is a naturally occurring plant material sourced from the elderberry shrub. It is provided as a powder for use in various research and laboratory applications.

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6 protocols using sambucus nigra

Biotinylated Lectin Binding Assay

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The biotinylated lectins were obtained from commercial sources. Concavalia ensiformis (ConA)^{17 (link)} was obtained from Dako (Carpinteria, CA, USA). The lectins obtained from Vector Laboratories (Burlingame, CA, USA) were as follows: Maackia amurensis (MAL-1),^{18 (link)} Sambucus nigra (SNL),^{19 (link)} Solanum tuberosum (STA),^{20 (link)} wheat germ agglutinin (WGA),^{21 (link)}

Servais A.B., Kienzle A., Ysasi A.B., Valenzuela C.D., Wagner W.L., Tsuda A., Ackermann M, & Mentzer S.J. (2018). Structural heteropolysaccharides as air-tight sealants of the human pleura. Journal of biomedical materials research. Part B, Applied biomaterials, 107(3), 799-806.

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Platelet Desialylation by Neuraminidase

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Human platelets were treated with α2–3,6,8 neuraminidase from Clostridiumperfringens (New England Biolabs) to remove terminal sialic acid (18 (link)). Isolated platelets in Walsh’s buffer (∼1.5 × 10⁹ platelets per milliliter) were incubated with 2.5 mU of α2–3,6,8 neuraminidase for 15 min at 37 °C. Desialylation was confirmed by lectin binding [biotinylated version of sambucus nigra (Vector Laboratories Ltd.)] via flow cytometry (SI Appendix).

Juan-Colás J., Hitchcock I.S., Coles M., Johnson S, & Krauss T.F. (2018). Quantifying single-cell secretion in real time using resonant hyperspectral imaging. Proceedings of the National Academy of Sciences of the United States of America, 115(52), 13204-13209.

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Platelet Glycoprotein and Lectin Analysis

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Platelet membrane glycoproteins (GPs) were detected using fluorescein-conjugated antibodies and flow cytometry. Citrated whole blood was centrifuged at 150 g for 15 minutes without brake at room temperature. Subsequently, platelets rich plasma was stained with the fluorescein isothiocyanate-conjugated (FITC) anti-human CD41a (GPIIb), phycoerythrin-conjugated (PE) anti-human CD42b (GPIbα, both from BioLegend) or allophycocyanin-conjugated anti-human CD36 (BD Biosciences) for 20 minutes. To analyze lectin expression, the fluorescein isothiocyanate-labelled sialic acid-binding lectin, Sambucus nigra (SNA) lectin, and the PE-streptavidin labeled sialic acid-binding lectin, Maackia amurensis lectin II (MAL-II) (both obtained from Vector Laboratories, California), for 30 minutes at room temperature. The BD FACSAria II (Becton Dickinson, Franklin Lakes, NJ) was used for analysis. To correct for the increased fluorescence intensity in giant platelets, the mean fluorescence intensities (MFIs) of lectins were divided by MFIs of the CD41a for comparison with those of normal platelets.

Mekchay P., Ittiwut C., Ittiwut R., Akkawat B., Le Grand S.M., Leela-adisorn N., Muanpetch S., Khovidhunkit W., Sosothikul D., Shotelersuk V., Suphapeetiporn K, & Rojnuckarin P. (2020). Whole exome sequencing for diagnosis of hereditary thrombocytopenia. Medicine, 99(47), e23275.

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Multicolor Flow Cytometry Immunophenotyping

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The following monoclonal antibodies (derived either from BD biosciences or eBioscience) were used: FITC-labeled anti-CD3 (145-2C11), -CD4 (RM4-5), -NK1.1 (PK136 ebio), -MHC-I (H-2Kb; AF688.5BD),-CD8b (eBIOH35); PE-labeled anti-CD8b (eBioH35-17.2), -MHC-II (M5/114.15.2) and −NK1.1 (PK136); and APC-labeled anti-FoxP3 (FJK-169). Biotin-labeled plant-derived lectin SambucusNigra (SNA) was obtained from Vector Labs, CA, USA, and binding was detected with PE-labeled Streptavidin (Jackson Immunoresearch, UK).

Perdicchio M., Cornelissen L.A., Streng-Ouwehand I., Engels S., Verstege M.I., Boon L., Geerts D., van Kooyk Y, & Unger W.W. (2016). Tumor sialylation impedes T cell mediated anti-tumor responses while promoting tumor associated-regulatory T cells. Oncotarget, 7(8), 8771-8782.

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Glycan Profiling of Confluent Cells

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Confluent cells were detached from cell culture dishes by gentle resuspension in PBS and were transferred to V-bottom 96-well plates for staining (1–2 × 10⁶ cells/well). Prior to this, control cells were treated with 4 µg/ml kifunensine (Bio-Techne, Minneapolis, MN, USA) overnight. Cells were washed with staining buffer (PBS supplemented with 2% (v/v) fetal calf serum, 2 mM EDTA) and were then stained successively with a near-IR fluorescent reactive dye (1:500, Thermo Fisher Scientific), biotin-conjugated lectins and streptavidin-PE (1:1000, Thermo Fisher Scientific) diluted in FACS buffer. The following biotinylated lectins were used: Sambucus nigra (5 µg/ml), Griffonia simplicifolia (0.5 µg/ml), Pisum sativum agglutinin (1 µg/ml), Lens culinaris agglutinin (0.5 µg/ml), Phaseolus vulgaris erythroagglutinin (1 µg/ml), Phaseolus vulgaris leucoagglutinin (1 µg/ml), and succinylated wheat germ agglutinin (2 µg/ml) (all from Vector laboratories, San Francisco, CA, USA) as well as Concanavalin A Type IV (1 µg/ml) and Tritium vulgaris (0.5 µg/ml) (from Sigma–Aldrich). Stained cells were fixed with 2% formaldehyde (FA) in PBS (Polysciences) and were analysed using a FACSCanto II and the FlowJo 10.6.2 software package (Becton Dickinson).

Hobohm L., Koudelka T., Bahr F.H., Truberg J., Kapell S., Schacht S.S., Meisinger D., Mengel M., Jochimsen A., Hofmann A., Heintz L., Tholey A, & Voss M. (2022). N-terminome analyses underscore the prevalence of SPPL3-mediated intramembrane proteolysis among Golgi-resident enzymes and its role in Golgi enzyme secretion. Cellular and Molecular Life Sciences, 79(3), 185.

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Lectin ELISA for Glycan Detection

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Lectin ELISAs were performed as described previously (3 (link)). In brief, samples were coated on plates and detected with biotinylated lectin [wheat germ agglutinin (WGA, Vector Laboratories), Ricinus communis agglutinin (RCA, Vector Laboratories), or Sambucus nigra agglutinin (SNA, Vector Laboratories)], streptavidin labeled with horseradish peroxidase, and tetramethylbenzidine. The reaction was stopped with 2 M H₂SO₄, and the absorbance was measured at 450 and 540 nm using a BioTek microtiter plate reader. We previously confirmed that SNA does not bind Fc glycans (3 (link)).

van de Bovenkamp F.S., Derksen N.I., van Breemen M.J., de Taeye S.W., Ooijevaar-de Heer P., Sanders R.W, & Rispens T. (2018). Variable Domain N-Linked Glycans Acquired During Antigen-Specific Immune Responses Can Contribute to Immunoglobulin G Antibody Stability. Frontiers in Immunology, 9, 740.

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