Optical clarity and mechanical strength are properties of cornea that are dictated by the microscopic superstructure of collagen lamellae. In this study, we used second harmonic generation microscopy to characterize and study chicken cornea model in thee -dimensions. Through the use of a combination of second harmonic generation (SHG) microcopy and Fouriertransform image analysis, adult chicken corneas were imaged. We found that adjacent lamellae of anterior chicken cornea stroma rotate in a counterclockwise fashion with a rotational pitch of around 0.7 degree/μm. However, the rotation stops upon reaching posterior stroma. Our approach can be extended into understanding structural architecture of human cornea and contribute to improved engineering products for corneal transplantation.