Study of microstructural changes in cornea under different stresses using a custom-made 100 kHz swept-source optical coherence tomography imaging system.

Swept-source optical coherence tomography (SSOCT) is a non-destructive imaging technique that provides high-resolution visualisation of tissue microstructure. This study examined corneal morphology and central corneal thickness (CCT) variations under cold (0 °C), formalin (10%), and heat (80 °C) stress using a custom-built 100 kHz SSOCT system over 72 h. Quantitative evaluation of CCT and the optical attenuation coefficient (OAC) provided depth-resolved insight into microstructural alterations, eliminating the need for layer segmentation. In controls, stromal CCT increased from 839.8 ± 39.1 μm at 24 h to 2006.8 ± 47.6 μm at 72 h, indicative of post-immersion stromal oedema. Under cold stress, the epithelium-to-bowman’s layer measured 152.7 ± 16.9 μm and the stroma 772 ± 34.4 μm at 24 h, increasing to 1240 ± 37.4 μm at 72 h. Formalin fixation produced inverse trends, with epithelium-to-bowman’s CCT reducing from 101.2 ± 6.8 µm (24 h) to 50.1 ± 12.6 µm (72 h), and stromal CCT decreasing from 1090 ± 77.8 µm to 806.6 ± 61.8 µm, reflecting fixation-induced dehydration and collagen cross-linking. In contrast, heat stress resulted in an epithelium-to-bowman’s CCT of 163.7 ± 12.3 μm at 24 h and 227.1 ± 33.6 μm at 72 h, while stromal thickness declined from 908.2 ± 39.0 μm to 862.3 ± 50.1 μm. OAC mapping exhibited strong concordance with histopathological findings, validating it as a reliable, non-invasive indicator of tissue morphology. Statistical analysis (ANOVA, p < 0.001) confirmed significant morphological variations across all experimental conditions. Overall, SSOCT integrated with OAC analysis enables sensitive detection of corneal microstructural changes under stress, providing a quantitative and non-invasive framework for assessing tissue preservation, estimating postmortem interval, and advancing forensic and medicolegal investigations.