Team:USP-Brazil/Results:FreezeDry

Results

Lyophilization

Defining the best cryoprotectants

As the baker’s dry yeast may confirm, protocols for yeast cells are abundant. Using as reference the known methodologies for Saccharomyces cerevisiae and other yeast species[22], we tested combinations of two cryoprotectants for the lyophilization of P. pastoris: powdered milk and monosodic glutamate (see our Notebook). After realizing the lyophilization process with 1.5 mL tubes, the following results were obtained:

Table 2: CFU for each test in 1.5 mL tubes. 24h lyophilization process with 1 mL of cell suspension.

Although threalose was not used—what is also a good protectant for yeast species [22]—, the result showed itself interesting enough for a cheaper way to make lyophilized Pichia. Since lactose is not metabolized by this yeast [23], this might not affect P_AOX1 activation if the powdered milk does not have residual glucose quantities. Those interesting findings support the next subject of our project: the idealized device who could compartmentalize the Pichia powder in a simple, secure and user-friendly way. Check the next section where we try to put all the pieces together to built the product.