Denudation of human amniotic membrane by a novel process and its characterisations for biomedical applications.

This study was aimed to investigate the suitability of a modified method to get decellularised human amniotic membrane (DHAM). The obtained membrane was subjected to physico-chemical and biological evaluations to validate its potential for biomedical applications. The human amniotic membrane was processed with detergent and alkali followed by enzymatic treatments. Hematoxylin and eosin (H&E) and Masson's trichrome staining of membrane were in accordance with conjectures: the decellularised membrane stained for extracellular matrix is rich in collagen. Scanning electron micrograph also showed the denudation in the processed membrane with the cellular impressions on the basement membrane. Physical characteristics namely the differential scanning calorimetric, tensile, shrinkage behaviour and the Fourier transform infrared spectra of decellularised membrane showed its stability and intact structure similar to the unprocessed membrane. In the visible range of light, the membrane was found to be transparent from 90 to 98 %. Proliferation rate of fibroblasts, keratinocytes, myoblasts and hepatocytes were significantly upregulated compared to the control. The cell morphologies were normal and differentiation of myoblasts into myotubes were more pronounced in decellularised membrane. Proliferation of corneal limbal cells on decellularised membrane showed 92-100 % confluency on day 21 and the migrated cells displayed a spindle shape and changing later to a more cuboidal appearance.