The pioneering work on the influence of age and sex on collagen autofluorescence was reported earlier by Shuster and co-workers involving human subjects which concluded that skin collagen decreases with age. On the contrary, in a study by Kollias and co-workers, a positive correlation between age and collagen deposition by measuring endogenous skin fluorescence using albino hairless mouse models was observed. In the present study although, there were variations in the collagen intensities among un-illuminated control and laser treatment groups with age, it was found to be nonsignificant, may be attributed to the selected age range, which would have been improved if the age range selected was between 5-55 weeks. Similarly, autofluorescence results indicated higher collagen deposition in male mice compared to females in all the tested time points, however the difference in collgen deposition was not found to be statistically significant. Present report is the first of its kind, wherein autofluorescence technique has been utilized to monitor collagen non-invasively as well as to reveal the differential collagen deposition with gender during acute wound healing. Previous studies from our laboratory contributed significantly in demonstrating the augmented effect of red light laser irradiation on collagen synthesis in pre-clinical models during tissue regeneration. Thus, the major focus of the current work was to elucidate the prognostic potential of in vivo autofluorescence in the factual measurement of collagen rather than proving the beneficial effect of LLLT in collagen deposition. In order to achieve the stated objective, normalized spectral intensity and area under the curves for collagen peaks were computed and compared. In our previous report, we have utilized normalized NADH/collagen ratio to assess the collagen deposition in the spectra. However, in the present study we believed that the inclusion of collagen intensity and an additional parameter such as AUC could help in acquiring more accurate information about the collagen. Although, collagen intensities and AUC were extracted from the same spectra, we believe that the intensity could provide information at a single point i.e., at a wavelength of 405 nm whereas, AUC could provide supplementary information to the spectral intensity values. Thus, AUC measurments in the present study were performed only to support/confirm the findings of collagen intensity at 405 nm. Single exposure of the optimum laser treated animals displayed 1.171, 1.165, 1.125, 1.020 and 1.012 fold increase in collagen intensity and 1.165, 1.151, 1.127, 1.019 and 1.016 fold increase in AUC values compared to un-illuminated controls on days 5, 10, 30, 45 and 60 post-wounding respectively. Two different spectral parameters i.e. spectral intensity and AUC displayed similar and comparable value for collagen deposition substantiating.