For cloning the PrESTs, a pool of RNA consisting of material from several human tissues is used in an RT-PCR approach. First, the amplified gene fragments are cloned and the sequence verified. Then, an E. coli recombinant protein expression system is used for expressing the clones, and the products are purified using nickel-containing matrices (IMAC).
The expressed PrEST Antigens are validated using ESI mass spectrometry. Purity is analyzed using the SDS page, and the PrEST Antigen amount is quantified using a Nanodrop system.