Bacterial adhesion tests were carried out using two different bacterial strains: reference Staphylococcus aureus (ATCC 25923) and clinical Staphylococcus aureus (MRSA 1030). Before testing, the bacteria inoculate was incubated for 18 h at 37 °C in 50 cm3 of BactoTM Tryptic Soy Broth (TSB) (Becton Dickinson). Then, the investigated samples were placed into 24-well plates. On their surfaces, 1 mL of bacterial suspension (~1∙108 CFU∙mL−1) was added, and the samples were incubated at 37 °C for 4 h with shaking at 60 rpm. Then, the samples were gently washed with PBS solution (8.0 g∙L−1 NaCl, 0.2 g∙L−1 KCl, 1.44 g∙L−1 Na2HPO4, 0.24 g∙L−1 KH2PO4). To remove the adhered bacteria, the samples were placed into new 24-well plates and treated intensively with 1 mL of 0.25% trypsin. Then, 100 μL of the bacteria and trypsin solution were collected and mixed with 0.9% NaCl at concentrations of 1:1000, 1:10 000, and 1:100 000. One hundred microliters of the prepared solutions were spread on agar plates. The agar plates were incubated at 37 °C for 24 h. After that, the bacterial colonies were counted.
Bactotm tryptic soy broth tsb
Manufactured by BD
Sourced in United States
BactoTM Tryptic Soy Broth (TSB) is a general-purpose growth medium used for the cultivation of a wide range of microorganisms, including bacteria and fungi. It provides essential nutrients to support the growth and proliferation of these microorganisms in a laboratory setting.
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4 protocols using bactotm tryptic soy broth tsb
1
Bacterial Adhesion Assay for Staphylococcus aureus
2
Characterizing Optimal Growth Conditions
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To determine the optimal temperature and pH for growth, the strain was incubated for 7 days in BactoTM Tryptic Soy Broth (TSB; Becton, Dickinson and Company, Sparks, MD, USA) at 5 °C, 10 °C, 20 °C, 25 °C, 28 °C, 37 °C, and 45 °C and at pH 3 to pH 13, respectively. Growth in various concentrations of NaCl was also examined after 7 days of incubation in TSB. Chemotaxonomic experiments were conducted on the basis of a previous report [7 (link)]. Physiological and biochemical characteristics were evaluated using API ZYM, API Coryne, and API 50CH Biochemical Test Kits (bioMérieux, Marcy I’Etoile, France) according to the manufacturer’s instructions. Assimilation of carbon sources at a final concentration of 1% (w/v) was tested using ISP 9 agar as the basal medium according to Pridham and Gottlieb [8 (link)].
3
Culturing and Identifying E. coli O157:H7
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Bacterial strains were stored at -80°C in BactoTM Tryptic Soy Broth (TSB; Becton Dickinson and Co., Sparks, MD, United States) containing 20% glycerol.
All the strains were cultured on chromogenic E. coli O157:H7 medium (Hopebio, Qingdao, China) for 24 h at 37°C. The bright red single colonies were used to identify presumptively the presence of E. coli O157:H7 (Figure1 ). And then cells were incubated in BactoTM TSB at 37°C overnight with shaking (110 r/min).
All the strains were cultured on chromogenic E. coli O157:H7 medium (Hopebio, Qingdao, China) for 24 h at 37°C. The bright red single colonies were used to identify presumptively the presence of E. coli O157:H7 (Figure
4
Bacterial Strains Adsorption of AgNps
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All the strains from the American Type Culture Collection (ATCC, United States) were preserved at -80°C in our laboratory until use, namely Escherichia coli O157:H7 (ATCC 43895), S. aureus (ATCC 27664) and Salmonella (ATCC 13076). Bacterial strains were stored at -80°C in BactoTM Tryptic Soy Broth (TSB; Becton, Dickinson and Co., Sparks, MD, United States) containing 20% glycerol, were inoculated in LB agar plates and incubated at 37°C overnight. Afterward, the pure culture was transferred to BactoTM TSB at 37°C overnight with shaking (110 rpm). The bacterial concentration was about 108 CFU/mL. One milliliter of cell suspension was centrifuged at 6,000 ×g for 5 min and the supernatant was discarded. The cell pellets were washed three times with double distilled water, centrifuged at 6,000 ×g for 5 min and finally resuspended with 10 mL of the silver colloidal nanoparticles or double distilled water. Next, the solution was mixed with a vortex to obtain a mixture that was as homogeneous as possible and then stood for 3–5 min. This procedure was designed to make AgNps and bacteria adsorb each other. Transmission electron microscopy (TEM) measurements were also performed to investigate the distribution of the AgNps coated on the Escherichia coli O157: H7.
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