Staphylococcus aureus TISTR 517, obtained from the Thailand Institute of Scientific and Technological Research, Pathum Thani Province, Thailand, was cultured in Luria Bertani (LB) agar medium (Himedia, India) at 37 °C in a static incubator for 24 h. A single colony was inoculated into 25 × 150 mm of the screw cap test tube containing 10 ml of LB broth medium, incubated at 37 °C, 200 rpm in a shaking incubator for 24 h. The cells were stored in 15% glycerol at −80 °C until use.
Lb agar medium
Manufactured by HiMedia
Sourced in India
LB agar medium is a common culture medium used for the growth and maintenance of various bacterial strains. It provides the necessary nutrients and solidifying agent for bacterial colonies to develop and thrive in a controlled laboratory environment.
Automatically generated - may contain errors
3 protocols using lb agar medium
1
Culturing and Preserving Staphylococcus aureus
2
Preparing Bacterial and Fungal Cultures
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The standard strains of Pseudomonas aeruginosa (ATCC 9027), Staphylococcus aureus (ATCC 6538), Bacillus subtilis (9372) and Candida albicans (ATCC 10231) used in this project were obtained in lyophilized form (purchased from the Pasture Institute of Iran). These strains were activated by culturing in Luria Bertani (LB) agar medium (Hi Media, India) and incubated at 37°C for 24 hours. Next, 4 ml of LB medium was used to obtain individual colonies. Then, it was incubated over night at 37°C while being shaken at a speed of 200 rpm. The cells were separated using centrifugation at 4°C and at a rate of 3000 rpm for 15 minutes.
Consequently, they were cleaned by washing twice before re-suspending in Ringer’s solution to achieve bacterial concentrations ranging between 10
7–108 colony forming units (CFU)/mL.
Consequently, they were cleaned by washing twice before re-suspending in Ringer’s solution to achieve bacterial concentrations ranging between 10
7–108 colony forming units (CFU)/mL.
3
Microbial CCA Production Protocol
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Escherichia coli mutant as the CCA producer was provided by The National Research and Innovation Agency (BRIN), previously named was Biotech Center, Agency for the Assessment and Application of Technology -Indonesia. The culture was maintained in Miller Luria Bertani (LB) agar medium (Himedia, Mumbai, India) pH 7.0, containing 100 µgmL -1 ampicillin. The chemicals used were ammonium sulfate p.a (Merck, Germany), chitosan powder with a degree of deacetylation (DD) > 75% obtained from the local market, CPC-Na (Biorbyt, Netherlands), and 7-ACA (Tokyo Chemical Industry, Japan). Other chemicals such as 50% glutaraldehyde, absolute methanol, p-dimethylaminobenzaldehyde (p-DAB), NaOH granules, and acetic acid glacial were obtained from Sigma-Aldrich (Singapore).
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