To determine the antibacterial activity of methanol extract and different fractions of S. caseolaris fruit, bacterial strains such as E. coli (ATCC 25922), S. aureus (ATCC 29247), and B. subtilis (ATCC 6633) were used in this experiment. The following chemicals were used in this study including methanol (XiLong, China), hexane (C6H14; Vietnam), ethyl acetate (C4H8O2, Vietnam), n-Butanol (C4H8O2, China), Amoxicillin (Vietnam), Gallic acid (C7H6O5; China), Folin-Ciocalteu reagent (Germany), Sodium carbonate (Na2CO3; Germany), Quercetin (C15H10O7; Sigma-Aldrich, Singapore), Aluminium chloride (AlCl3; China), Tryptic Soy Agar medium (Sigma-Aldrich, Germany).
Tryptic soy agar medium
Manufactured by Merck Group
Sourced in United States, Germany
Tryptic soy agar medium is a nutrient-rich culture medium used for the growth and isolation of a wide range of microorganisms, including bacteria and fungi. It provides the necessary nutrients and growth factors to support the cultivation of a diverse microbial population.
Automatically generated - may contain errors
Lab products found in correlation
Aluminium chloride, by Merck Group (1 mentions)
Amoxicillin, by Merck Group (1 mentions)
N butanol, by Merck Group (1 mentions)
Methanol, by Merck Group (1 mentions)
Gallic acid, by Merck Group (1 mentions)
Quercetin, by Merck Group (1 mentions)
Folin ciocalteu reagent, by Merck Group (1 mentions)
Sodium carbonate, by Merck Group (1 mentions)
Blood agar base, by Merck Group (1 mentions)
7890b system, by Agilent Technologies (1 mentions)
Omegawax 250, by Merck Group (1 mentions)
Milli q iq 7005 water purification system, by Merck Group (1 mentions)
Penicillin, by Merck Group (1 mentions)
Sodium hydroxide (naoh), by Avantor (1 mentions)
Chloroform, by Avantor (1 mentions)
9 protocols using tryptic soy agar medium
1
Antibacterial Activity of S. caseolaris Fruit
2
Oral Supplementation of Akkermansia muciniphila
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A. muciniphila culture and oral supplementation were performed as previously described22 (link) with a few modifications. A. muciniphila (ATCC BAA-835) was grown in fresh pre-reduced Brain Heart Infusion (BHI) broth (Sigma-Aldrich, BD 237,500, USA) under anaerobic conditions (75% N2, 20% CO2, and 5% H2) at 37°C and incubated for 3–4 days. Afterward, cultures were centrifuged and condensed in anaerobic PBS containing 20% (vol/vol) glycerol to a concentration of ~ 1 × 1010 c.f.u/mL under strictly anaerobic conditions and stored at −80°C until use. The bacteria were incubated in a pre-reduced Tryptic Soy Agar medium (Sigma-Aldrich, BD 236,950, USA) for 4–5 days to determine the A. muciniphila counts (c.f.u./mL). A. muciniphila glycerol stocks were diluted with anaerobic PBS to a final concentration of 2 × 108 viable c.f.u. per 0.2 mL. Mice were treated by oral gavage with 200 μL of either A. muciniphila suspension or anaerobic PBS three days a week (on days 1st, 3rd and 5th) for four weeks, starting 48 h after the last gavage of antibiotics.
3
Phagocytic Activity of Aeromonas hydrophila
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Aeromonas hydrophila MTCC-1739 was cultured in tryptic soy agar medium (Sigma-Aldrich) for 24 h at 37°C. The cell number was adjusted to 5 × 106 CFU/mL. The phagocytic activity of the blood leukocyte was determined according to the method of Cai et al. [33 (link)]. The phagocytic activity (%) was calculated as using the formula:
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Aeromonas hydrophila MTCC-1739 was cultured in tryptic soy agar medium (Sigma-Aldrich) for 24 h at 37°C. The cell number was adjusted to 5 × 106 CFU/mL. The phagocytic activity of the blood leukocyte was determined according to the method of Cai et al. [33 (link)]. The phagocytic activity (%) was calculated as using the formula:
4
Quantifying Bacterial Load in Blood
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Blood samples were collected by cardiac puncture, 100μl blood was plated on tryptic soy agar medium (Sigma, USA) at 37°C. The bacterial load was quantified by counting CFU on each plate after overnight incubation.
5
Preparation and Sterilization of Cell Culture Media
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Pre-formulated, dehydrated tryptic soy agar medium and the following chemicals were purchased from Sigma Aldrich (St. Louis, MO, USA) and prepared according to the manufacturer's instructions: PBS tablets (pH ± 0.2, 0.1 mol/l), non-essential amino acid solution, trypsin-EDTA solution and Dulbecco's modified Eagle medium with GlutaMAX-1 (DMEM). PBS was prepared according to the manufacturer's instructions and filter-sterilized with a 0.2 µm syringe filter. Faetal bovine serum (FBS) was obtained from American Type Culture Collection (ATCC, Manassas, VA, USA).
6
Isolation of Hemolymph Bacteria from Leafhoppers
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CO2-anesthetized leafhoppers were surface-sterilized by submerging them first in 95% ethanol for 1 to 2 min, then in 1.2 to 1.5% sodium hypochlorite solutions for 2 min and, eventually, rinsing them 2 or 3 times in sterile water [40 (link)]. Under a dissecting microscope, the hemolymph was aspirated with a fine, flame-drawn needle inserted into the insect body, between the thorax and the abdomen. The fluid was transferred to a tube containing 1X PBS. The hemolymph of five individuals was combined, then split in two and plated onto chocolate (Blood Agar Base—Sigma Aldrich—added with 7% horse defibrinated blood) and purple agar (Bromocresol Purple Broth—Sigma Aldrich—added with 1.5% agar) solid medium. The hemolymph of a total of 40 individuals was obtained. Plates were kept at 26 °C in the dark for up to 14 days. Colonies were numbered (using C when isolated from chocolate agar and P when isolated from purple agar plates) and transferred onto new chocolate and purple plates. For subcultures and maintenance, a Tryptic Soy Agar (TSA) medium (Sigma Aldrich) was eventually used.
7
Profiling Volatile Organic Compounds of Lactobacilli
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The profile of volatile organic compounds (VOCs) produced by L. paracasei 85 and L. buchneri 93 were determined using an Agilent 7890bsystem (Santa Clara, CA, USA) coupled with Agilent 5977 Inert XL MSD apparatus. The system was equipped with a fused-silica capillary column coated with a polar stationary phase (OMEGAWAX™250, 30 m × 0.25 mm × 0.25 μm, Supelco, Bellefonte, PA, USA). The bacterial culture with or without ZEN treatment was inoculated with Tryptic Soy Agar (TSA) medium (Sigma-Aldrich, Steinheim, Germany). The vials were placed at an angle into the incubator. The VOCs were extracted with 50 μm polydimethylsiloxane (PDMS)/divinylbenzene (DVB) fiber (Supelco, Bellefonte, PA, USA) at 37 °C for 50 min. The absorbed molecules were desorbed into the injection port for 5 min at 250 °C. VOCs were separated with the following program: held 40 °C for 5 min, heated at 5 °C/min to 185 °C, held for 1 min, heated at 5 °C/min to 200 °C, held for 10 min, and then heated at 10 °C/min to 240 °C. Helium was used as the carrier gas. The MS interface and the ion source were maintained at 250 and 230 °C, respectively. Acquisition was performed in electron impact mode (70 eV) with the mass range of 30–300 m/z. Compounds were identified by matching mass spectra with the NIST14.1 library [2 (link)]. The analyses for each sample were carried out in triplicate.
8
Chitosan Characterization and Application
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Low molecular weight CS (LMW) with 50–190 kDa molecular weight and 75–85% deacetylation degree, medium molecular weight CS (MMW) with 190–310 kDa molecular weight and 75–85% deacetylation degree, and high molecular weight CS (HMW) with 310–390 kDa molecular weight and above 75% deacetylation degree were purchased from Merck (Darmstadt, Germany). 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), streptomycin, and penicillin were purchased from Sigma-Aldrich, St. Louis, MO, USA. IPA was purchased from POCH (Gliwice, Poland). MilliQ water was used to prepare all the aqueous solutions (Milli-Q® IQ 7005 Water Purification System, Millipore, Boston, MA, USA). All other reagents were of analytical grade or higher. Chloroform, hydroxyacetic acid, and sodium hydroxide were purchased from Avantor Performance Materials Poland S. A. (Gliwice, Poland). The carbon dioxide used to saturate the CS precipitate was obtained from Linde Gaz Polska Sp. z o. o. (Gdańsk, Poland). Tryptic soy agar (TSA) medium was purchased from Merck (Darmstadt, Germany).
9
Isolation of Endophytes from Cherry Roots and Seeds
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The young, healthy feeding roots and ripened cherries were washed thoroughly under tap water followed by 3 rinses with Sterile Saline Solution (3S) containing 0.85% (w/v) of sodium chloride under a laminar flow hood. The excess of moisture was removed on sterile filter paper. The roots without external damages were excised in a small fragment of 2-3 cm, and the seeds were retrieved by removing the exocarp (skin), the mesocarp (pulp), and the endocarp (parchment). At this step, the plant materials were surface-sterilized with sodium hypochlorite (2.5% available chlorine) for 2 min., followed by 5 rinses with 3S to remove the surface sterilizing agent. The surface-sterilized roots were crushed with sterile mortar-pestle and resuspended in 5 mL of 3S. Seeds were cut in 2-4 fragments on a sterile filter paper. The sterilized roots and seeds were then plated on Petri dishes containing Tryptic Soy Agar (TSA) medium (Merck KGaA, Darmstadt, Germany). To assess surface sterilization efficiency, the 3S from the last rinse was inoculated by flooding on control TSA containing Petri dishes in triplicate. All the Petri dishes were incubated at 28 • C for up to 14 days. Isolates were purified, stored in glycerol 25% at -80 • C, and considered as endophytes if no growth was observed on the sterilization control plates.
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