BRAIN HEART INFUSION AGAR

Protocol tips

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	-The medium is tested for compatibility using 7% v/v oxalated horse blood, defibrinated horse blood or defibrinated sheep blood. There should be no evidence of lysis or darkening after incubation at 37°C, 25°C or 4°C for 72 hours.

Protocol tips
-The medium is tested for compatibility using 7% v/v oxalated horse blood, defibrinated horse blood or defibrinated sheep blood. There should be no evidence of lysis or darkening after incubation at 37°C, 25°C or 4°C for 72 hours.

Publication protocol

Prior to testing, one vial of each strain was removed from the freezer and subcultured twice on Brain Heart Infusion (BHI) blood agar plates (Oxoid) and incubated for 24 h at 37°C in a microaerophilic atmosphere, in order to ensure its purity and vitality. The inoculation procedure was carried out according to the recipient the media were poured in. The inoculation procedure for pneumococci kept into plates or glass tubes was as follows: a portion of the blood culture was taken with a 0.001 mL calibrated loop and streaked out onto the surface of the plates or glass tubes with supplemented media. A 4-quadrant streaking method was adopted to inoculate the plates. The tubes, partially closed, and the plates were incubated for 24 h at 37°C in a microaerophilic atmosphere. In order to inoculate pneumococci into microtubes, a portion of the overnight blood culture was taken with a cotton swab and rolled twice over the surface of the agar. Microtubes were completely closed and incubated for 24 h at 37°C in a microaerophilic atmosphere. After incubation, glass tubes and microtubes were sealed with parafilm, while plates were placed inside hermetically sealed plastic bags. Finally, all subcultures were maintained at room temperature until further evaluation.

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