- -Start by thawing the serum and the control serum bottle at 2-8°C.
- -Stir the bottled serum to avoid a precipitant from forming during the thawing, then place it in the water bath adjusted to 80°C.
- -Once the control bottle temperature reaches 50°C, adjust the bath thermostat to 70°C; when the bottle temperature reaches 55°C, change the thermostat to 60°C.
- -When the serum reaches the temperature of 56°C, use a timer (30 minutes is the registered time) and change the thermostat of the water bath to 56°C. During those 30 minutes, check the temperature regularly and maintain it stable by adjusting the thermostat, by adding some cold water, and by lowering the lid or not. Stir the serum every 5 to 10 minutes.
- -Keep the serum at room temperature for 30 minutes, and then freeze the serum at -20°C.
The stirring is crucial for good serum quality. If the serum is not correctly mixed, a precipitant of salts, proteins and lipids will appear, and allow crystalline and flocculation deposits to form. These deposits are not toxic for cell culture but affect the serums appearance and consistency.
Heating the serum for long periods can destroy the growth promoters, which can also enable deposits to form.