Cylindrical grid light lens with simplified grid lighting for light lens microphone

Description

RELATED APPLICATION DATA

[0001] This application claims the benefit of and priority under 35 U.S.C. §119(e) to U.S. Patent Application No. 62 / 961,058, filed January 14, 2020, entitled "System for a Lightsheet Microscope Lattice Illuminator Using a Cylindrical Lattice Lightsheet".BACKGROUND

[0002] The present disclosure is generally directed to lightsheets in optical microscopes and, more specifically, to illumination devices for optical microscopes.

[0003] It has been shown that an optical lattice used for lightsheet microscopy gives better resolution, optical sectioning, and light dosage performance than other lightsheet techniques. This combined with the natural large reduction in light dosage native to lightsheets has led to commercial instruments that have been able to capture images that were previously unable to be imaged. Certainly in the domain of single cell imaging, these instruments can be considered to be the best current microscopes.

[0004] Unfortunately, the current means for generating optical lattices are expensive and have very low light efficiency (typically <10% of the input light can be used at the sample). These means usually are complicated, contain many moving and electronic parts, and are difficult to align and maintain. Currently there are researchers that are trying to generate optical lattices with femtosecond pulsed IR lasers to achieve a multi-photon effect. These efforts are made extremely difficult due to the low efficiency. US 2018 / 0011303 A1 relates to methods and systems for generating non-diffracting light sheets for multicolor fluorescence microscopy. US 10,310,246 B2 relates to a converter, illuminator, and light sheet fluorescence microscope, wherein improved image quality by structured illumination or pivoting illumination and faster image acquisition are both achieved.

[0005] A further lattice light-sheet microscope is known from Ellefsen, Kyle L., et al, "Dynamic Ca2+ imaging with a simplified lattice light-sheet microscope: A sideways view of subcellular Ca2+ puffs", Cell Calcium, Elsevier, Amsterdam, NL, vol. 71, 01.12.2017, pages 34-44, ISSN: 0143-4160, DOI: 10.1038 / S41592-019-0327-9.SUMMARY

[0006] The present disclosure generally relates to lightsheets in optical microscopes. More specifically, an exemplary embodiment of the present disclosure relates to a simplified illumination device for optical microscopes. Even more specifically, an exemplary embodiment of the present disclosure relates to an optical system for generating an optical lattice as illumination for an optical microscope.

[0007] The invention is defined by claim 1.

[0008] A lattice can be generated piecewise by breaking down the optical lattice into constituent parts at the pupil plane. The three parts of the optical lattice in the pupil plane can be generated separately and integrated in time to give the same result as a lattice generated by a Spatial Light Modulator (SLM) at the image plane. Additionally, since a lattice is most often used dithered (or smeared in on axis), and since a dithered lattice has no information in the axis of dither, there is no need for interference between the beams in that direction. Accordingly, the extension is that a beam that has the same structure at the pupil plane as one constructed by an SLM in the image plane will give the same results. Therefore one must only generate the pupil plane pattern and one gets the optical lattice at the image plane. To get the same effect as a dithered lattice one can additionally remove the interference between the beams at the pupil plane in the dither direction.

[0009] If one considers the anatomy of the lattice beam at the pupil plane, one finds that a lattice beam can be generated by three parallel lines clipped by an annulus. This forms a total of four light beams. The two side beams create an envelope that reduces the sidelobes of the lattice. If the light beams interfere with each other, they form a regular structure along the dither axis. The two center beams interfere with each other to generate a single dimensional Bessel which provides the sharp peak in the center of the lattice. All of the beams together generate the complete lattice.

[0010] The angular content of the beams at the pupil plane are of particular importance to generating a lattice. A large angular content in the dither axis generates a long lattice which is used to form a wide sheet. In the other axis (i.e., the axial direction in the imaging path), perpendicular to the dither axis (normal to the sheet), there must be no angular content (the beam is collimated) so that one only gets one central lobe in the sheet. Therefore the simplest implementation of a lattice would be to place three cylindrical lenses in a collimated laser beam and focus these lines onto the pupil plane. An annulus is placed at the pupil plane to aperture the three lines and generate a lattice. This setup is very simple and generates a complete lattice. The width of the lattice is dependent on the focal length of the cylindrical lenses. This method of generating a lattice is called Cylindrical Lattice Lightsheet (CLLS).

[0011] CLLS is much simpler to make than other methods, but the implementation above has poor throughput since most of the light is blocked by the annulus. CLLS is further improved by adding a pair of axicon lenses to move the light from the input beam of CLLS to the ring of the annulus. This improves the light throughput by many factors and the total efficiency of the system can be >90%, which is much better than previous methods.

[0012] The lattice generated by this means is the same as previous methods, but what is actually desired is a dithered lattice which removes the content of the lattice in the dither axis. A typical lattice will do this by using a scanner to smear the lattice in that direction. CLLS can use a scanner in this method, but better techniques may be used.

[0013] To generate a dithered lattice, one must remove the interference between the three constituent beams. One could time modulate the beams and integrate, but this would lead to complexity and loss of efficiency. One can remove the interference from one of the three beams by adding a waveplate to one of the beams. There are now only two beams interfering and we get partial dither. The third beam can be altered with a time-varying waveplate using a liquid crystal (LC) device or by moving shear waveplates. These devices are inexpensive and have good throughput. The resulting beam is now identical to a standard dithered lattice without the need for a scanner. The beam can be used as a lightsheet directly.

[0014] Considering further the effect of the beam at the back aperture on a scanned beam, the lateral position of any part of the beam has little effect on the resultant sheet. Therefore there is no need for the two side beams to be on the outside of an annulus, but they can be anywhere horizontally. One can then combine the two side beams and move them to the middle of the back aperture. One must still break the symmetry between this now middle beam and the top and bottom beams, so a waveplate is added to either the middle beam or the top and bottom beams. This means that the complete scanned lattice sheet can be created with only a single cylindrical lens.

[0015] The much higher efficiency and simpler design of CLLS leads to easier implementation of multi-photon lattice systems, and lattice microscopes in general. The reduction in cost alone is at least 2x for the complete system, coming from the expensive SLM, galvos and high power lasers.

[0016] The exemplary apparatus can comprise: An optical device for generating three parallel lines that are focused in the dither axis and collimated in the other axis. This device could exemplarily be a set of three cylindrical lenses. A mask for blocking unwanted light from the three lines. The exemplary apparatus further comprises a pair of axicons before the cylindrical lenses as means for increasing the light through the annulus.

[0017] This exemplary apparatus could additionally comprise: Means for removing the interference between the three generated lines. This means could be a fixed waveplate on one line and a time-varying waveplate on another line.

[0018] This apparatus when combined would constitute a simplified optical lattice illumination system for a lightsheet.

[0019] This device has the advantage of much less cost, easier alignment and maintenance, and much higher light efficiency.

[0020] Aspects of the disclosure are thus directed towards lightsheets in optical microscopes.

[0021] Still further aspects of the disclosure are directed toward a simplified illumination device for optical microscopes.

[0022] Even further aspects of the disclosure are directed toward an optical system for generating an optical lattice in a lightsheet microscope.

[0023] These and other features and advantages of this disclosure are described and, or are apparent from, the following detailed description of the exemplary embodiment.BRIEF DESCRIPTION OF THE DRAWINGS

[0024] The exemplary embodiments of the present disclosure will be described in detail, with reference to the following figures wherein: Figure 1 illustrates the pattern of illumination at the pupil plane to generate a lattice. Figure 2 illustrates an exemplary embodiment of the present disclosure with the minimal optical components needed to generate a lattice lightsheet. Figure 3 is an image of data generated by an exemplary embodiment of the present disclosure. Figure 4 illustrates where additional optics can be added to modify the beam in an exemplary embodiment of the present disclosure. Figure 5 illustrates an exemplary embodiment of the present disclosure showing additional optics to smear the lightsheet. Figure 6 is an image of the data generated by an exemplary embodiment of the present disclosure. Figure 7 illustrates the pattern of illumination where the side beams are combined to the middle of the pupil plane. Figure 8 illustrates an exemplary embodiment of the present disclosure with combined side beams to generate a lattice lightsheet. Figure 9 illustrates the pattern of illumination at the pupil plane to generate a hex lattice. DETAILED DESCRIPTION OF THE DRAWINGS

[0025] The exemplary embodiments of the present disclosure will be described in relation to microscopes, imaging systems, and associated components. However, it should be appreciated that, in general, known components will not be described in detail. For purposes of explanation, numerous details are set forth in order to provide a thorough understanding of the present disclosure. It should be appreciated however that the present disclosure may be practiced in a variety of ways beyond the specific details set forth herein.

[0026] Figure 1 illustrates the pattern of illumination required to generate an optical lattice as seen at a pupil plane 10. The pattern comprises three lines 11, 12, and 13. The three lines 11, 12, and 13 may indicate locations on an annulus through which light passes. In some embodiments, the three lines may be vertical (e.g., spanning from one side of the annulus to the other) and / or parallel. As viewed in Figure 1, the lines 11 and 12 are located at the two edges of the annulus, while the line 13 is located in the middle (e.g., down the center) of the annulus. In some embodiments, the line 13 may separate the annulus evenly into two similar or identical sections. By forming such a pattern at the pupil plane 10 (e.g., the placement of the lines 11, 12, and 13), an optical lattice is generated at the image plane. The lines 11, 12, and 13 may be spaced such that different optical lattices are generated on the image plane. For example, the lines 11, 12, and 13 may be spaced to generate a square lattice or a hex lattice. In some embodiments, the pattern may comprise additional and / or alternative lines (e.g., three parallel lines, four parallel lines, etc.). The positioning of the three lines 11, 12, and 13 may generate an un-dithered lattice. As previously mentioned, the three parts of the optical lattice (e.g., the light represented by the three lines 11, 12, and 13) may be integrated in time to provide an un-dithered lattice, rather than utilizing an SLM at the image plane.

[0027] Figure 2 illustrates an exemplary embodiment of the present disclosure which shows the minimum optics required to generate a lattice lightsheet. An input laser beam 21 is sent through an array of cylindrical lenses 22. As the laser beam 21 passes through the cylindrical lenses 22, the cylindrical lenses 22 may bend, curve, or otherwise focus the light into an annulus mask 23. In some embodiments, the cylindrical lenses may focus the light into the annulus mask 23 such that the lines 11, 12, and 13 form on the annulus mask 23. In other words, the image of the light from the annulus mask 23 may have the form of the pupil plane shown in Figure 1. In some embodiments, multiple cylindrical lenses (e.g., two, three, four, etc.) may be used. The image is relayed using an optical relay 24 to the pupil plane of an objective 25. The optical relay 24 may be a series of mirrors or other optical elements (e.g., axicons) configured to take the shape of the light focused into the annulus mask 23 and replicate or pass the shape to the pupil plane of the objective 25.

[0028] In some embodiments, the minimum optics may further comprise multiple annuli (e.g., a plurality of annuli) placed at the pupil plane (not shown). The multiple annuli may include additional or alternative thicknesses and / or diameters, which may allow the system to be changed, altered, or otherwise reconfigured to generate different lattice shapes and sizes depending on, for example, a specific experiment or other application. In some embodiments, similar or the same effect of changing the thickness and / or diameter of the annulus using a variable magnification relay or an SLM.

[0029] Figure 3 shows an image of the optical lattice as generated by the setup of Figure 2. The lattice depicted in Figure 3 is identical to those generated by other methods and closely matches the theory.

[0030] Figure 4 illustrates the pattern of illumination as in Figure 1 and the modifications used to generate a smeared sheet. A smeared sheet may be a lattice sheet with a less clear lattice structure. To break the fine lattice structure seen in Figure 3 in the horizontal direction, the coherence between the three lines may be disrupted. A waveplate 41 is added to a beam (e.g., the beam represented by the line 11), which removes the coherence of the beam with the other two beams on the pupil plane. A variable waveplate or moving shear plates 42 is added to another beam (e.g., the beam represented by line 12) to remove the coherence with the center beam 43, which is left unchanged. In some embodiments, the smearing of the sheet may create a similar or the same lattice as a dithered SLM at an image plane. For instance, the three beams (e.g., lines 11, 12, and 13) may be integrated in time to create a sheet identical to a dithered SLM lattice.

[0031] Figure 5 illustrates an exemplary embodiment of the present disclosure as in Figure 2, with additional optics to generate a smeared sheet. The beam is split into three lines using the cylindrical lenses 51. After the annulus mask, 52, separate optics are added to the now separate beam paths. A waveplate 53 is added to one path (which may be similar to the waveplate 41). A variable waveplate 54 is added to another path. The variable waveplate 54 may be an adjustable device to adjust the coherence of the light passing therethrough with the other lines passing through the annulus. The use of the waveplate 53 and / or the variable waveplate 54 gives a smeared lightsheet out of the objective 55.

[0032] Figure 6 shows an image of the smeared optical lattice as generated by the setup of Figure 5. The broken coherence changes the lattice from Figure 3 to be a smeared lightsheet.

[0033] Figure 7 illustrates the pattern of illumination as in Figure 1 and the modifications needed to generate a smeared sheet with combined side beams. The top and bottom beams 71 may remain unchanged from the embodiment shown in Figure 1 (e.g., the top and bottom beams 71 may be similar to line 13 from Figure 1). The two side beams 11 and 12, however, are combined into a single beam 72. To break the fine structure seen in Figure 3 in the horizontal direction, a waveplate 73 is added to the single beam 72, which removes the coherence of the single beam 72 with the other two beams 71.

[0034] Figure 8 illustrates an exemplary embodiment of the present disclosure as in Figure 7, with optics combining the side beams. The beam is sent through the cylindrical lenses 81. After passing through the mask 82, three separate beams are created. A waveplate 83 is added to the middle beam. A relay 84 images the mask onto the pupil plane. This gives a smeared lightsheet out of the objective 85. Figure 9 illustrates the pattern of illumination to generate a "hex" lattice at the pupil plane 90 of the excitation objective. The lattice shown in Figure 9 is similar to Figure 1, but with the spacing of the three beams changed. In particular, the three beams are spaced closer together. This spacing of the three beams (91, 92 and 93) causes the resulting lattice to change to a hexagonal pattern used in, for example, SIM applications.

Claims

1. An illumination system for a lightsheet microscope for generating a lattice, the illumination system comprising: means to generate two or more parallel lines focused on a pupil plane, the means comprising a set of cylindrical lenses (22, 51, 81), and means to confine light from the two or more parallel lines (11, 12, 13) to the clear aperture of an annulus, the means comprising: a pair of axicons configured to further confine the light to the annulus, wherein a spacing of the parallel lines and the diameter and thickness of the annulus are chosen to generate the lattice.

2. The illumination system of claim 1, wherein the means to generate the two or more parallel lines (11, 12, 13) further comprises: a mask.

3. The illumination system of claim 1, wherein a variable optical relay (24) is configured to change a spacing of the two or more parallel lines (11, 12, 13).

4. The illumination system of claim 1, wherein the means to confine the emitted light to the annulus further comprises one or more of the following: an annulus mask (23, 52), and a phase-plate to shape light into an annulus ring.

5. The illumination system of claim 1, wherein a variable optical relay (24) is configured to change the diameter of the annulus.

6. The illumination system of claim 1, further comprising means to remove coherence between multiple beams output from the illumination system.

7. The illumination system of claim 6, wherein the means to remove the coherence comprises one or more of the following: a waveplate (41, 53, 54, 73, 83) disposed in a path of one or more of two or more output beams of the emitted light, and a liquid crystal waveplate.

8. The illumination system of claim 1, further comprising: an objective (25, 55, 85) disposed near a first end of an optical mask such that the apertured, focused two or more parallel lines (11, 12, 13) are focused on the pupil plane of the objective.