Lactose (1 ) was from Lachema (Brno, Czech Republic). 2′-Fucosyllactose (2 ) was purchased from Biosynth (San Diego, CA, USA). Lacto-N-tetraose (3 ), lacto-N-neotetraose (4 ), blood-group antigen A type 6 (5 ), and blood-group antigen B type 6 (6 ) came from Elicityl (Crolles, France; in the catalog, they are rather uncommonly denoted as type 5). Protein-assay-dye-reagent concentrate and bovine plasma γ-globulin (IgG) for protein calibration were from BioRad (Watford, Hertfordshire, UK). p-Nitrophenyl β-d -galactopyranoside (pNP-Gal), 3,4-diethoxy-3-cyclobutene-1,2-dione (squaric acid diethyl ester), and Dowex 66 base-free were purchased from Sigma-Aldrich (Prague, Czech Republic). Buffers for ELISA assays were prepared from salts bought from Roth (Karlsruhe, Germany). If not stated otherwise, chemicals, solvents, general buffers, and cultivation media were purchased from VWR (Stříbrná Skalice, Czech Republic).
2 fucosyllactose
Manufactured by Biosynth
Sourced in United Kingdom, United States
2'-Fucosyllactose is a human milk oligosaccharide (HMO) that serves as a structural component of human breast milk. It is a complex carbohydrate molecule composed of fucose, glucose, and galactose. 2'-Fucosyllactose is involved in various biological functions, including prebiotic and immunomodulatory properties.
Automatically generated - may contain errors
Lab products found in correlation
Lactose, by Biosynth (2 mentions)
Protein assay dye reagent concentrate, by Bio-Rad (1 mentions)
Dextran sulfate sodium (dss), by MP Biomedicals (1 mentions)
X gal, by Thermo Fisher Scientific (1 mentions)
Ampicillin, by Merck Group (1 mentions)
Lactose, by Avantor (1 mentions)
Lacto n tetraose, by Biosynth (1 mentions)
Lacto n hexaose, by Biosynth (1 mentions)
Lacto n neotetraose, by Biosynth (1 mentions)
3 sialyllactose, by Biosynth (1 mentions)
6 sialyllactose, by Biosynth (1 mentions)
N acetylneuraminic acid, by Biosynth (1 mentions)
Ckk 8, by Merck Group (1 mentions)
Sodium hydroxide naoh pellets, by Merck Group (1 mentions)
Ammonium hydroxide solution nh4oh, by Merck Group (1 mentions)
Hplc grade acetonitrile acn, by Honeywell (1 mentions)
Iodomethane, by Merck Group (1 mentions)
Amylopectin, by Biosynth (1 mentions)
Ammonium acetate nh4ac, by Merck Group (1 mentions)
Sophorose, by Megazyme (1 mentions)
7 protocols using 2 fucosyllactose
1
Carbohydrate-based Reagents and Buffers
2
Characterization of Il22ra1 and RegIIIγ Knockout Mice
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Il22ra1tm1a/tm1a, RegIIIγtm1a/tm1a, and WT C57BL/6N mice were maintained and phenotyped by the Sanger Mouse Genetics Programme (White et al., 2013 (link)). Endogenous Il22ra1 expression was detected by overnight incubation with 0.1% (w/v) X-gal (Invitrogen). All animals were kept under specific pathogen-free conditions, and colony sentinels tested negative for Helicobacter spp. Mice were infected orally with 109 cfu of Kanamycin (Kan)- and nalidixic acid (Nal)-resistant luminescent Citrobacter rodentium ICC180 (Wiles et al., 2006 (link)) or intravenously (i.v.) with 105 cfu Salmonella enterica serovar Typhimurium M525 TETc (Clare et al., 2003 (link)). Where indicated, C. rodentium-infected mice were administered intraperitoneal (i.p.) ampicillin (50 mg/kg; Sigma-Aldrich), oral 2′-fucosyllactose, H disaccharide, lactose (2 mg/day; Carbosynth), or PBS controls. DSS colitis was induced by administering 2% (w/v) DSS (MP Biomedicals) in drinking water for 7 days, followed by normal autoclaved water. The care and use of mice were in accordance with the UK Home Office regulations (UK Animals Scientific Procedures Act 1986) and were approved by the Sanger Institute’s Animal Welfare and Ethical Review Body.
3
Oligosaccharide Standards for Research
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The oligosaccharide standards; 2′-Fucosyllactose (2′FL), Lacto-N-tetraose (LNT), 3′-Sialyllactose (3′SL), 6′-Sialyllactose (6′SL), Disialyllactose (DSL), Lacto-N-hexaose (LNH), N-Acetylneuraminic acid (Sialic Acid), LS-tetrasaccharide c (LSTc), Lacto-N-neotetraose (LNnT) and Lacto-N-neohexaose (LNnH) were purchased from Carbosynth Ltd. (Berkshire, UK) and lactose was obtained from VWR (Dublin, Ireland).
4
Atropine and Supplements Effects on HCE-F
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Initially, 5 × 103 HCE-F/well were seeded in 96-well plates and left to adhere, overnight, in complete culture medium. In a first set of experiments the following treatments were carried out in SFM: atropine (1, 1.7, and 3.4 mM), for 24, 48, and 72 h. In a second set of experiments, HCE-F cells were treated with 0.15% HA (HMW: 1.8 MDa; Fidia pharmaceutical SpA, Abano Terme, Italy), 0.5% colostrum (NZpurehealth, Auckland, New Zeland), 0.5% 2-Fucosyl-Lactose (Carbosynth, Berkshire, UK, cat. No. OF06739) and two mixes (Mix A: 0.15% HA + 0.5% colostrum; Mix B: 0.15% HA + 0.5% 2FL) in the presence of atropine 1 mM; the medium with the supplements was changed every day for 4 days. Cytotoxicity was measured by the MTT method with the cell counting kit-8 assay (CKK8, Sigma-Aldrich, St. Louis, Missouri, USA, Cat. No. 96992) according to the manufacturer’s protocol.
5
Carbohydrate Standards Preparation Protocol
6
Enzymatic Assay for Fucosyltransferase Activity
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Yeast extract and casein peptone were purchased from B. D. Bioxon (Mexico City, Mexico). pNP-Fuc, pNP, D-lactose, D-glucose, D-galactose, L-fucose, and Man Rogosa and Sharpe (MRS) agar were purchased from Sigma-Aldrich (St. Louis, MO, USA), 2′-fucosyllactose was purchased from Carbosynth (Berkshire, UK). Sodium phosphate and sodium hydroxide were purchased from J. T. Baker (Mexico City, Mexico). Milli-Q® (Billerica, MA, USA) water was used throughout the experiments.
7
Hydrolytic Activity of LacA β-Galactosidase
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Lactose monohydrate and acetonitrile (HPLC grade) were supplied by VWR (Barcelona, Spain). MOPS (3-(N-morpholino) propanesulfonic acid), xylitol, phenyl-βglucoside, maltulose, maltose, cellobiose, mannitol, sorbitol, sucrose, and pNP-β-Dgalactopyranoside were purchased from Sigma Aldrich (Stenheim, Germany).
Erythritol, isomaltulose, gentiobiose, isomaltose, lactulose and 2'-fucosyl-lactose, were purchased from Carbosynth (Berkshire, UK). KH2PO4 and K2HPO4 were supplied by Merck (Steinheim, Germany). All other chemicals were of analytical grade. The hydrolytic activity of LacA β-galactosidase (1.16 U mL -1 ) was also assayed by incubation at 30⁰C of different carbohydrates such as lactose, lactulose, cellobiose, gentiobiose, maltose, isomaltulose and sucrose at 200 g L -1 with 300 μL of enzyme in 50 mM of MOPS buffer supplemented with 20 mM of NaCl (pH=7) to a final volume of 1 mL. In the case of 2-fucosyl-lactose, isomaltose, nigerose and maltulose the concentration was 0.5 g L -1 . Reactions were stopped at 24 hours by heating at 100ºC for 5 min and samples were stored at -20⁰C for the subsequent analysis. Reactions were carried out in triplicate (n = 3). All these hydrolytic reactions were monitored by GC-FID.
Erythritol, isomaltulose, gentiobiose, isomaltose, lactulose and 2'-fucosyl-lactose, were purchased from Carbosynth (Berkshire, UK). KH2PO4 and K2HPO4 were supplied by Merck (Steinheim, Germany). All other chemicals were of analytical grade. The hydrolytic activity of LacA β-galactosidase (1.16 U mL -1 ) was also assayed by incubation at 30⁰C of different carbohydrates such as lactose, lactulose, cellobiose, gentiobiose, maltose, isomaltulose and sucrose at 200 g L -1 with 300 μL of enzyme in 50 mM of MOPS buffer supplemented with 20 mM of NaCl (pH=7) to a final volume of 1 mL. In the case of 2-fucosyl-lactose, isomaltose, nigerose and maltulose the concentration was 0.5 g L -1 . Reactions were stopped at 24 hours by heating at 100ºC for 5 min and samples were stored at -20⁰C for the subsequent analysis. Reactions were carried out in triplicate (n = 3). All these hydrolytic reactions were monitored by GC-FID.
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