To ensure quality control and assurance in tissue engineering, noninvasive, real-time and aseptic evaluation of cell-based devices is required before product release. In this study, Raman spectroscopy was applied to monitor the cellular activities of an oral mucosa equivalent (EVPOME) produced ex vivo from cultured autogenous oral keratinocytes and acellular dermis—AlloDerm. Raman spectra showed a positive correlation of the peak area ratio of amide I (1655 cm−1)/phenylalanine (1004 cm−1) with a negative linear regression (R2 > 0.95) according to the number of cultured days, especially on the 14thand 21st day. This work demonstrates the successful application of Raman spectroscopy for quantitatively monitoring and evaluating the maturity of EVPOME. Copyright © 2010 John Wiley & Sons, Ltd.