Ultrahigh-resolution optical coherence tomography imaging of protein crystals using gel inclusion technique

Abstract

Protein crystals are required for X-ray crystallography to determine three-dimensional structures of proteins at atomic resolution. The conventional microscopy is currently used for observation and screening of protein crystals. However, the three-dimensional imaging, which is important for automated treatment of protein crystals, is generally difficult by light microscopy. In addition, the protein crystals in the media are frequently difficult to identify by conventional light microscopy owing to the appearance of salt crystals or amorphous materials. In this work, we successfully demonstrated micro-scale, non-invasive, three-dimensional cross-sectional imaging of protein crystals using ultrahigh resolution optical coherence tomography (UHR-OCT). A low noise, Gaussian like, high power supercontinuum at wavelength of 800 nm was used as the light source. The axial resolution of 2 um in sample and the sensitivity of 95 dB were achieved. Since the protein crystal has homogeneous nano-structure, the optical scattering is negligibly small. Therefore, we used gel-inclusion technique to enhance the intensity of scattered signals, and clear, sharp 3D cross-sectional images of protein crystals were successfully observed. As the gel concentration was increased, the OCT signal intensity was increased. Using this method, the protein crystals surrounded by substantial amount of precipitates could be visualized, which is difficult by conventional light microscopy. The discrimination of protein and salt crystals was also demonstrated by the OCT signal intensity. The wavelength dependence of OCT imaging for protein crystal was examined at wavelength of 800-1700 nm regions. It was confirmed that the finest images were observed using 800 nm wavelength system.