Microscopic examinations of the histological sections of unwounded skin indicated the minimal changes in the collagen morphology in terms of collagen type, size and distribution and orientations. Vertically oriented red/yellow birefringent fibers were seen in un-wounded skin on day 10. Similar findings of occurrence of randomly oriented collagen fibers have also been reported from the earlier studies as a way to resist different mechanical/shearing forces. The microscopic images of the histological sections of un-illuminated control on days 5 and 10 post-wounding indicated the presence of thin weakly birefringent green and yellow fibers localized in papillary dermis of the regenerating tissue. On day 30 post-wounding, localization of yellow birefringent fibers were seen till mid dermis indicating the progression of healing. However, on day 45 and 60 post-wounding, yellow fibers were replaced by more bright red birefringent type I collagen. More interestingly, newly formed collagen fibers were vertically oriented in addition to parallel orientations. An exact explanation for the occurrence of vertically oriented collagen fibers in the regenerated skin is not clearly known. Single exposure of the optimum laser dose had significantly influenced the collagen content which was evident by the occurrence of both thick and thin fibers in papillary and mid dermal layers even on day 5 postwounding. In addition, brightly birefringent red fibers was prominently present in histological sections of the optimum laser treated animals on days 10, 30, 45 and 60 post-wounding clearly indicating the simulative effect of visible red light on collagen synthesis. On day 30 post-wounding, the type I collagen localized mainly in mid and deep dermal layers of the regenerated skin indicating the succession of healing under the influence of HeNe laser. Further, image analysis was performed for histological sections to quantify the total collagen deposited during the progression of healing. This was necessitated by the fact that histological assessments were qualitative and the prognostic potential of in vivo autofluorescence could only be tested with quantitative assessments through gold standard measurments. In the present study, since spectral discrimination of type I and III collagen were not performed, the results of image analysis of the histological sections were represented as total collagen, yellow and red fibers ) facilitating quantitative relationship between the two optical techniques. Wounded animals exposed to optimum dose of the red laser light displayed 5.70%, 5.90%, 1.73%, 1.43% and 1.09% fold increase in total collagen compared to un-illuminated control as reflected by the image analysis of Picro-Sirius stained histological sections. The microscopic images of the histological sections in RGB color space is firstly converted into HIS color scale through the “TissueQuant” software.