I have decellularized the umbilical cord with detergent based methods, the problem is that I am unable to solubilize it with pepsin and HCl, kindly guide me with the exact protocol of concentrations and conditions to solubilize it completely?
Use Hcl 0.01 M and 10 mg/mL dECM in 1 mg/mL pepsin at 25C. There could be multiple reasons why you are having issues with the solubilization. 1. pH is not in the right range for pepsin activity. Pepsin activity is highest at 2. Once you add the HCl to the dECM please check the pH and adjust it to around 2. 2. Decellularizing agents also interferes with the pepsin digestion. To make sure that these agents are removed wash the decelluliarized tissue extensively. Use multiple washes. 3. Make sure that the umbilical cord is powdered very fine. Use a cryomill for this purpose. It would be very difficult to digest very large particles. 4. Make sure that the room in which the digestion happen is around 25C. Hopefully these suggestions should help Tayyaba Bari
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