Imaging large, cleared samples requires microscope objectives that combine a large field of view (FOV) with a long working distance (WD) and a high numerical aperture (NA). Ideally, such objectives should be compatible with a wide range of immersion media, which is challenging to achieve with conventional lens-based objective designs.
Imaging large, cleared samples requires microscope objectives that combine a large field of view with a long working distance and a high numerical aperture . Ideally, such objectives should be compatible with a wide range of immersion media, which is challenging to achieve with conventional lens-based objective designs. Here we introduce the multi-immersion ‘Schmidt objective’ consisting of a spherical mirror and an aspherical correction plate as a solution to this problem.
To address this challenge, we introduce an approach for designing a multi-immersion objective based on a mirror instead of lenses. The underlying idea is that the reflection of a light ray by a mirror is independent of the refractive index of the medium the mirror is in contact with. Consequently, if we submerge a curved mirror inside a liquid-filled chamber and use it to focus light, the ray paths and location of the focus do not change when changing the bulk index of the immersion medium.
Fabian F. Voigt, Anna Maria Reuss, Martina Schaettin, Adriana L. Hotz, Stephan C. F. Neuhauss, Esther T. Stoeckli, Adriano Aguzzi & Fritjof HelmchenSven Hildebrand & Alard RoebroeckMartina Schaettin, Adriana L. Hotz, Stephan C. F. Neuhauss & Esther T. StoeckliF.F.V. and F.H. designed the project. F.F.V designed the objective, built the microscope, imaged samples and analyzed data. A.M.R., T.N., S.H., M.S., A.L.H., A.B., S.C.F.N., A.R., E.T.S., S.S.L. and A.A. provided samples. L.W.
Extended Data Fig. 5 Correlative imaging of a 4-day old Chicken embryo with the mesoSPIM light-sheet microscope and the Schmidt objective.) Maximum projections of stacks taken at higher zoom level showing individual axons. The white cells with strong autofluorescence are red blood cells which have not lost their nucleus yet at this developmental stage. The images are examples from one of two imaged chicken embryos.
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