Purification of Streptolysin

The protocol is applicable to bacterial cultures of from 2 to 20 yields 0.2 to 0.6 mg toxin per liter of culture. 

Preparation of culture medium and production of culture supernatants containing SLO. Seeding cultures of Streptococcus pyogenes, Richards strain, are prepared in Brain-Heart-Infusion medium by overnight incubation at 37°C. For long term storage of the strain, the culture is supplemented to 50% (by volume) with sterile glycerol and kept at -70°C. 

Toxin production medium is prepared from Bacto Proteose Peptone No. 3 (Difco) 20 g, and Yeast Extract 20 g (Difco), and flasks of the appropriate volume (up to 51) are filled with distilled water to bottom of the neck. 

After autoclaving at 110°C for 30min, 25 ml of sterile 10% NaHCOs and 20 ml of sterile 50% glucose are added. 

The culture is started with 20 ml inoculum per liter. Streptococci are cultured at 37°C and after 4 h the addition of NaHCOs and glucose is repeated. The pH of the medium is monitored every 30 min and adjusted to 7.5 with 1 M NaOH, while stirring with a magnetic bar. 

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