Combined optical coherence tomography and light sheet fluorescence microscopy for embryonic imaging

Abstract

Optical coherence tomography (OCT) and light sheet fluorescence microscopy (LSFM) are well-established imaging techniques preferred in developmental biology, e.g., embryonic imaging. However, each technique has its own drawbacks, such as resolution and molecular specificity with OCT and field-of-view (FOV) and speed with LSFM. To overcome these limitations for small animal embryo imaging, we have developed a co-aligned multimodal imaging system combining OCT and LSFM. The OCT probe and LSFM excitation beams were combined and scanned with a galvanometer-mounted mirror through the same objective lens. The light sheet thickness was ~13 μm. The LSFM collection arm consisted of a 0.8 numerical aperture water immersion objective, tube lens, and CCD camera, resulting in a transverse resolution of ~2.1 μm. The OCT system was based on a 100 kHz swept-source laser with a central wavelength of 1050 nm and had a lateral resolution of ~15 µm and an axial resolution of ~7 μm. Images of fluorescent microbeads and a fluorescent-tagged mouse embryo at gestational day 9.5 showed the capabilities of the multimodal imaging system. Since the OCT system and LSFM system were co-aligned, image registration was straightforward and enabled high-throughput multimodal imaging without the need for complex registration techniques.