We observed thin film covering the protein drop within 6 days at 22 C incubator in the microbatch plate, using 1 microliter of protein and 1 uL of crystallizing agent per drop, covering with oil. As I‘ve known this film may be due to oxidation of the denatured protein. Next we are planning to include reducing agent like DTT in the protein complex, though the enzyme does not need a reducing agent for activity.

However,I wonder if there are other factors that can cause this phenomenon and how to prevent formation of the film.