Oblique Plane Microscopy With Tilted Objective for High-NA Imaging
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Solution Overview
Problem
Conventional oblique plane microscopy systems suffer from low numerical aperture (NA) values, making them less effective for high-resolution imaging, and are not compatible with standard sample preparation techniques like glass slides and multi-well plates.
Innovation Solution
An oblique plane microscopy system is configured with a first objective lens having a numerical aperture NA1≥n0 and a refractive index n1≈n0, a second objective lens with NA2≥0.95, and a third objective lens tilted relative to the second, allowing for high NA imaging compatible with standard sample preparations.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional oblique plane microscopy systems are used, then the system is compatible with standard sample preparation techniques, but the numerical aperture is too low for high-resolution imaging
Solution Approach 1:
The system divides the imaging function into three separate objective lenses, each with specific functions: the first objective (O1) provides illumination and collection with high NA, the second objective (O2) re-images the oblique plane, and the third objective (O3) collects light from O2. This segmentation allows each lens to be optimized independently, achieving high NA while maintaining compatibility with standard samples.
Solution Approach 2:
The patent introduces a tilted third objective lens at an angle relative to the optical axis, creating a three-dimensional arrangement of objectives. This angular dimension allows the system to capture light from oblique planes while maintaining high numerical aperture, resolving the contradiction between resolution and sample compatibility.
2Object-affected harmful factors
If light-sheet microscopy is used, then photobleaching and phototoxicity are reduced, but the implementation is awkward and not compatible with standard sample preparation
Solution Approach 1:
The first objective lens O1 serves multiple functions simultaneously: it acts as the illumination lens, the collection lens, and maintains compatibility with standard sample preparation techniques. This multi-functionality simplifies the system while retaining the benefits of light-sheet microscopy for living samples.
Solution Approach 2:
Instead of using complex orthogonal lens arrangements, the patent inverts the approach by using a single objective for illumination and collection, then using additional objectives to re-image and collect light from oblique planes. This inversion simplifies the implementation while maintaining compatibility with standard samples.
3Measurement precision
If the numerical aperture of the first objective is increased to approach the theoretical limit, then imaging resolution is improved, but the working distance is reduced
Solution Approach 1:
The second objective O2 acts as an intermediary that re-images the oblique plane formed by O1 onto a new plane where the third objective O3 can collect light. This intermediary imaging step allows the system to achieve high NA with extended working distance by separating the illumination/collection function from the imaging function.
Solution Approach 2:
By tilting the third objective at an angle and using three-dimensional light cone geometry, the system achieves high numerical aperture while maintaining adequate working distance. The angular arrangement in three dimensions allows light collection from oblique planes without requiring the objective to be extremely close to the sample.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The system achieves a numerical aperture approaching the theoretical limit of 1.33, providing high-resolution, low photobleaching, and phototoxicity imaging compatible with standard sample preparations.
Implementation Method 1
a first objective lens having at least one of a first numerical aperture NA1≥n0 and a first immersion medium with a first refractive index n1 that is approximately equal to n0
Implementation Method 2
a second objective lens with NA2≥0.95... the second objective lens being configured and arranged to collect substantially all the light from the first objective
Implementation Method 3
a third objective lens tilted relative to the second... the third objective being configured and arranged to collect substantially all the light from the second objective
Implementation Method 4
a combination of the first and second microscopes being configured to produce an intermediate image of the sample with a magnification MRR, wherein the magnification MRR is approximately equal to a ratio (n1/n2) of the refractive indices
Data Source
AI summary
Systems and methods for oblique plane microscopy are disclosed.