how do you prepare substrate (chicken keratin powder)?

Chicken feathers were defatted with chloroform–methanol (1:1) in continuous agitation for 24 h, then washed three times with distilled water, and dried in the air. For keratin extraction, the

chicken feathers (50 g) were immersed in 1000 mL of 0.5 M sodium sulfide for 6 h with continuous agitation at ambient conditions. Soluble keratin was first centrifuged for 10 min at

10,000× g and then precipitated from the supernatant using 70% ammonium sulfate. The

precipitate was washed several times with distilled water and then dried at 40 ◦C