Il10-/- mice were weaned 21 days after birth and supplemented with a 5 mM solution of either 2-fucosyllactose (2FL), 3-fucosyllactose (3FL), 3-sialyllactose (3SL), 6-sialyllactose (6SL) (Glycom A/S, Hørsholm, Denmark) or D-lactose (Sigma-Aldrich, Buchs, Switzerland) in sterile filtrated water ad libitum for 28 days. For depletion of intestinal microbiota, 4–6 weeks old mice were supplied with 0.5 g/L vancomycin (AppliChem), 1 g/L ampicillin (VWR), 1 g/L neomycin (Fisher Bioreagents) and 0.2% aspartame (Sigma) in drinking water for 6 days. Once per day, mice received intragastric gavage of 250 μL of a 1 g/L metronidazole (Sigma) solution. After antibiotic treatment, mice were treated with 10% PEG 3000 solution (Sigma) in drinking water to flush the remaining bacteria and antibiotic. Neo-colonization with specific bacterial strains was done by intragastric gavage of 250 μL containing 109 CFU of the respective strain in PBS. Fecal pellets were collected to monitor changes of the microbiota.
Vancomycin
Manufactured by AppliChem
Sourced in Germany
Vancomycin is a laboratory product used for antimicrobial susceptibility testing. It is a glycopeptide antibiotic effective against Gram-positive bacteria, including methicillin-resistant Staphylococcus aureus (MRSA).
Automatically generated - may contain errors
Lab products found in correlation
Metronidazole, by Merck Group (2 mentions)
Deferoxamine, by Merck Group (2 mentions)
Piperacillin, by Merck Group (2 mentions)
2 2 bipyridyl, by Merck Group (2 mentions)
Novobiocin, by Merck Group (2 mentions)
Clavulanic acid, by Merck Group (2 mentions)
Mecillinam, by Merck Group (2 mentions)
Teicoplanin, by Merck Group (2 mentions)
Neomycin, by Thermo Fisher Scientific (1 mentions)
Ampicillin, by Avantor (1 mentions)
Aspartame, by Merck Group (1 mentions)
D lactose, by Merck Group (1 mentions)
Alexa fluor 488 donkey anti rabbit igg, by Thermo Fisher Scientific (1 mentions)
Penicillin, by Merck Group (1 mentions)
Dimethyl sulfoxide (dmso), by AppliChem (1 mentions)
Phosphomycin, by Merck Group (1 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (1 mentions)
Chloramphenicol, by Merck Group (1 mentions)
Nystatin, by AppliChem (1 mentions)
Ampicillin, by Merck Group (1 mentions)
7 protocols using vancomycin
1
Modulation of Intestinal Microbiota in Mice
2
Production and Characterization of Anti-Waddlia Antibodies
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Polyclonal rabbit antibodies against W. chondrophila were produced in our laboratory as described previously [44 (link)]. Secondary antibody, Alexa Fluor® 488 donkey anti-rabbit IgG, was purchased from Thermo Fisher Scientific (Waltham, MA, USA). Clavulanic acid, deferoxamine, mecillinam, novobiocin, penicillin, phosphomycin, piperacillin, teicoplanin, and 2,2′-bipyridyl were purchased from Sigma-Aldrich (St. Louis, MO, USA). Vancomycin was obtained from AppliChem (Darmstadt, Germany). MP265 was purchased from American Custom Chemicals Corporation (San Diego, CA, USA). All drugs were diluted in deionized water with the exception of MP265 and 2,2′-bipyridyl, which were diluted in dimethyl sulfoxide (DMSO, AppliChem) and in ethanol, respectively, in order to obtain the specific concentrations described in Table 1 . Finally, the solutions were filtered through a 0.22 μm pore filter and stored at −20 °C.
3
Culturing Human Cancer Cell Lines and Helicobacter pylori
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Human gastric carcinoma AGS and NCI-N87 cells and human cervical carcinoma HeLa cells (ATCC) were cultured in RPMI 1640 medium (Gibco®/Life Technologies) supplemented with 10% heat-in-activated foetal calf serum (FCS) (Gibco®/Life Technologies) in a humidified incubator at 37 °C with 5% CO2 and passaged once every 2–3 days.
H. pylori strain P1wt (wildtype) and isogenic mutants P1cagA (CagA-deficient) and P1virB7 (T4SS-deficient) as well as strain P12 were grown on agar plates containing 10% horse serum, 5 µg/ml trimethoprim, 1 µg/ml nystatin, and 10 µg/ml vancomycin (AppliChem) under microaerophilic conditions at 37 °C for three days. For the P1cagA and P1virB7 strains, the agar plates were supplemented with chloramphenicol (Sigma). Bacteria were replated and cultured for another two days before use.
H. pylori strain P1wt (wildtype) and isogenic mutants P1cagA (CagA-deficient) and P1virB7 (T4SS-deficient) as well as strain P12 were grown on agar plates containing 10% horse serum, 5 µg/ml trimethoprim, 1 µg/ml nystatin, and 10 µg/ml vancomycin (AppliChem) under microaerophilic conditions at 37 °C for three days. For the P1cagA and P1virB7 strains, the agar plates were supplemented with chloramphenicol (Sigma). Bacteria were replated and cultured for another two days before use.
4
Glioma Syngeneic Mouse Model with ABX
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Glioma syngeneic mouse model (GM) are obtained injecting glioma cells (GL261; CT2-a and GL261-rfp) as previously described [10 (link)]. Mice were treated with not-absorbable ABX (0.5 g/l vancomycin, AppliChem; [0.5 g/l], gentamicin, Aurogene) and sucralose (0.5%) to improve palatability or with sucralose alone (control solution) in autoclaved water. ABX and control solutions were changed every 2 days, and continuously administrated for 2 weeks before and 3 weeks after tumor injection.
5
Depleting Gut Commensal Microbiota
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For the depletion of gut commensal microbiota, animals were given ampicillin (1 g l−1; Sigma), vancomycin (500 mg l−1; Applichem), neomycin sulphate (N; 1 g l−1; Applichem), and metronidazole (1 g l−1; Sigma) in their drinking water for four weeks as described previously [52 (link)].
6
Inducing Aberrant Body Formation in Waddlia Infection
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HEp-2 and Vero cells were seeded at 4 × 106 cells per 25-cm2 flask (for RNA extraction) or at 2.5 × 105 cells per well in 24-well plates (for immunofluorescence) the day before infection. Waddlia suspension was prepared by filtering a 5-day-old co-culture through a 5-μM pore filter (Millipore, Carrigtwohill, Ireland) to eliminate amoebal cysts and trophozoites. Mammalian cells were infected with the bacterial filtrate diluted 100-fold (in case of subsequent treatment with drugs that induce aberrant body formation) or 1,000-fold (for kinetics of untreated infections). Infected cells were centrifuged at 1,790 × g for 10 min at room temperature and incubated at 37°C with 5% CO2 for 15 min. The medium containing non-internalized bacteria was then removed and replaced with fresh DMEM supplemented with 10% FBS.
In order to induce aberrant body formation, drugs were added at 2 hpi (teicoplanin, 250 µg/mL, and Vancomycin, 500 µg/mL) or 8 hpi (BPDL, 75 µM; deferoxamine, 260 µg/mL; novobiocin, 250 µg/mL; penicillin G, 1,000 µg/mL; piperacillin, 500 µg/mL; mecillinam, 200 µg/mL; clavulanate, 900 µg/mL; and fosfomycin, 500 µg/mL). Clavulanic acid, deferoxamine, mecillinam, novobiocin, penicillin G, fosfomycin, piperacillin, teicoplanin, and 2,2′-bipyridyl were purchased from Sigma-Aldrich (Buchs, Switzerland). Vancomycin was purchased from AppliChem.
In order to induce aberrant body formation, drugs were added at 2 hpi (teicoplanin, 250 µg/mL, and Vancomycin, 500 µg/mL) or 8 hpi (BPDL, 75 µM; deferoxamine, 260 µg/mL; novobiocin, 250 µg/mL; penicillin G, 1,000 µg/mL; piperacillin, 500 µg/mL; mecillinam, 200 µg/mL; clavulanate, 900 µg/mL; and fosfomycin, 500 µg/mL). Clavulanic acid, deferoxamine, mecillinam, novobiocin, penicillin G, fosfomycin, piperacillin, teicoplanin, and 2,2′-bipyridyl were purchased from Sigma-Aldrich (Buchs, Switzerland). Vancomycin was purchased from AppliChem.
7
Synthesis and Characterization of AMP Palm-KGK-NH2
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The enzyme DNase I and the antibiotic vancomycin were purchased by AppliChem (Germany) and the AMP Palm-KGK-NH 2 (Palm) was synthesized manually by solid-phase synthesis method on polystyrene AM-RAM resin, using Fmoc/tButyl strategy as previously described [24] . Coupling was performed with HOBt/DIPCDI method, the Fmoc protecting group were removed with 20% piperidine. Crude peptides were cleaved from resin using a mixture of trifluoroacetic acid (TFA), triisopropylsilane (TIS) and water as scavengers. The final products were purified by reverse-phase high performance liquid chromatography (RP-HPLC) in a mixture of acetonitrile -water with 0.1% TFA as an eluent. Molecular weights of peptides were determined by matrix -assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF).
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