Engineered liquid crystal shutter as dynamic long pass optical filter

Abstract

The present disclosure relates to a novel liquid crystal assembly for use in a multispectral imaging system. In particular, the liquid crystal assembly includes an engineered polarizer and a liquid crystal. In various aspects, the liquid crystal assemblies utilize disadvantages found in conventional polarizers and liquid crystals that require complex temperature regulation.

Description

Technical Field

[0001] The present disclosure generally relates to an engineered liquid crystal shutter for multispectral imaging. Specifically, the shutter relies on previously undesirable limitations of optical polarizers to increase the ratio of transmitted light to one or more optical sensors that can be used for imaging and analysis at video rate switching speeds. Background Art

[0002] Polychromatic or "multispectral" imaging is a method of surveying information from a wide range of different wavelengths of light and displaying it within a relatively narrow range of visible light. When processing spectral information, it is often necessary to apply different image analysis techniques to the different wavelengths of light in order to appropriately weight their values ​​before recombining them. To allow images of different colors to be processed independently, the spectrum is often separated into multiple spectral bands, and images from those spectral bands collected independently. Hardware for imaging a wide spectrum may include multiple image sensors responsible for collecting their respective spectral bands.

[0003] Liquid crystal shutters are commonly used in liquid crystal displays and are used to implement video rate switching in digital light projection (DLP) systems, such as in movie theaters. Typically, when a linear polarizer is used for a liquid crystal shutter, the spectral range of the effective shutter is largely limited by the polarization extinction ratio of the polarization filter and the limited transmittance of the liquid crystal itself. Summary of the invention

[0004] The present disclosure generally relates to a liquid crystal assembly for use with an imaging system having a light source. The assembly includes a polarizer disposed in the path of light from the light source. The polarizer is configured to have a light cutoff within a range outside its polarization range. The assembly also includes: a liquid crystal disposed in the path of light transmitted through the polarizer; and a drive voltage source, the drive voltage source configured to apply a first voltage and a second voltage to the liquid crystal. The first voltage and the second voltage implement a high electric field and a low electric field on the liquid crystal. The first voltage and the second voltage cause the liquid crystal to switch between an open state and a closed state. In one aspect, applying one of the first voltage or the second voltage causes the liquid crystal to switch to the open state. Applying one of the first voltage or the second voltage causes the liquid crystal to switch to the closed state.

[0005] In various aspects, the polarizer is configured to be transparent above the light cutoff. In one aspect, the light cutoff includes the longest wavelength of the spectral range, and the polarizer is configured to be inactive and transparent above the longest wavelength. In some aspects, the liquid crystal includes a twisted nematic (TN) cell, a Pi cell, a thermotropic liquid crystal and a lyotropic liquid crystal, a liquid crystal display (LCD), a liquid crystal with photoconductor properties, or a liquid crystal that is developed and adjusted under an electric field. In some aspects, the second voltage causes a backflow in the liquid crystal.

[0006] In various aspects, one of the first voltage or the second voltage is 0V. In another aspect, the second voltage is a plurality of voltages including a bipolar square wave having an average value of 0V. In one aspect, the first voltage, the second voltage, or both the first voltage and the second voltage include a plurality of voltages that produce a bipolar square wave having an average value of 0V. In another aspect, the bipolar square wave includes an amplitude of + / -0.1 to + / -5Vp-p, + / -3 to + / -30Vp-p, + / -16Vp-p, + / -24Vp-p, or + / -30Vp-p, or an amplitude between + / -16V (32Vp-p) and + / -30V (60Vp-p), including any value, or an amplitude between + / -30V (60Vp-p) and + / -100V (200Vp-p), including any value. In another aspect, the second voltage is a bipolar square wave having an amplitude of + / -0.1 to + / -5 Vp-p, + / -3 to + / -30 Vp-p, + / -16 Vp-p, + / -24 Vp-p or + / -30 Vp-p, or an amplitude between + / -16 V (32 Vp-p) and + / -30 V (60 Vp-p), including any values, or an amplitude between + / -30 V (60 Vp-p) and + / -100 V (200 Vp-p), including any values.

[0007] In various aspects, the bipolar square wave amplitude is modulated at the video capture rate of the imaging system. In another aspect, the amplitude of the bipolar square wave is modulated from 500 Hz to 2000 Hz. In some aspects, the drive voltage source causes the liquid crystal to switch between the first voltage and the second voltage for a duration of 1 millisecond to 40 milliseconds inclusive, or 2 milliseconds to 60 milliseconds inclusive, 60 milliseconds to 100 milliseconds inclusive, and up to 300 milliseconds inclusive. In some aspects, the drive voltage source causes the liquid crystal to switch between the first voltage and the second voltage for a video rate sampling duration of approximately between 1 ms and 10 ms, between 2 ms and 20 ms, or 6 ms.

[0008] In various aspects, the liquid crystal is a twisted nematic (TN) liquid crystal. In one aspect, the liquid crystal is heated to a range of 30°C to 50°C. In another aspect, the liquid crystal is enclosed in a housing of the imaging system and the liquid crystal is heated by waste heat generated by other components of the imaging system. In one aspect, the transition speed between the open state and the closed state is determined by the temperature of the liquid crystal and the voltage source drive level.

[0009] In various aspects, the polarizer is an engineered dye polarizer. In one aspect, the polarizer is configured to have a light cutoff in a range that is optimal for a fluorescence imaging process. In some aspects, the polarizer is selected to lose effectiveness or cutoff between 650 nm and 800 nm, or between 600 nm and about 800 nm, or between about 700 nm to about 800 nm, between about 800 nm to about 950 nm, between about 800 nm to about 880 nm, between about 775 nm to about 795 nm, or at about 785 nm. In another aspect, the polarizer is selected to lose effectiveness or cut off at 700 nm, 725 nm, 750 nm, 775 nm, 780 nm, 785 nm, 790 nm, 795 nm, 800 nm, 805 nm, 810 nm, 815 nm, 820 nm, 825 nm, 830 nm, 835 nm, 840 nm, 850 nm, 855 nm, 860 nm, 865 nm, 870 nm, 875 nm, 880 nm, 885 nm, 890 nm, 895 nm or 900 nm.

[0010] In various aspects, further include a beam splitter. In one aspect, the beam splitter has a front surface and a rear surface. In some aspects, the front surface has a front surface coating, and the rear surface has a rear surface coating. In another aspect, the front surface coating has a P polarization reflection higher than the rear surface coating. In one aspect, the front surface coating and the rear surface coating produce a P polarized light front surface to rear surface reflectivity ratio of at least 2:1, at least 4:1, at least 6:1, at least 8:1, at least 10:1, at least 12:1, at least 14:1, at least 16:1, at least 18:1 or at least 20:1. In one aspect, the S polarization reflection of the front surface and the rear surface of the beam splitter is minimized by the front surface coating and the rear surface coating. In another aspect, the polarizer is configured to block or attenuate ghosting or secondary reflection reflected from the beam splitter. In one aspect, the polarizer is configured to block the S polarized light from the beam splitter. In another aspect, the polarizer is configured to allow P-polarized light to pass through the liquid crystal assembly and reach the imaging lens and / or camera when the liquid crystal is in the open state. In one aspect, the first polarizer allows a front-to-back reflectivity ratio of the beam splitter of at least 2:1, 4:1, 6:1, 8:1, 10:1, 12:1, 15:1, 17:1, or 20:1 to pass therethrough and reach the imaging lens and / or camera. In some aspects, the total reflectivity of the beam splitter is about 11% or less, about 10% or less, about 9% or less, about 8% or less, about 7% or less, about 6% or less, about 5% or less, about 4% or less, or about 3% or less.

[0011] In various aspects, the liquid crystal assembly includes two polarizers or more polarizers. In another aspect, the liquid crystal assembly is suitable for a medical device including a surgical microscope, a confocal microscope, a fluoroscope, an exoscope, an endoscope, or a surgical robot. In another aspect, the liquid crystal assembly is optionally suitable for the medical device via the use of an optical gasket. In some aspects, the medical device is a KINEVO system (e.g., KINEVO 900), an OMPIPENTERO system (e.g., PENTERO 900, PENTERO 800), or a Leica FL800 system. In another aspect, the liquid crystal assembly is used in a method for imaging emitted light emitted by a fluorophore.

[0012] The present disclosure also generally relates to a method for imaging emission light emitted by a fluorophore at an imaging system including a liquid crystal assembly. The method includes allowing or directing visible light to reach a sample; directing excitation light to the sample; and directing the emission light and reflected visible light from the sample to the liquid crystal assembly. In various aspects of the foregoing method, the liquid crystal assembly includes an engineered polarizer disposed in the path of light from the sample; the polarizer is configured to have a light cutoff in a range between 600 nm and 900 nm. The liquid crystal assembly also includes: a liquid crystal disposed in the path of light transmitted through the polarizer; and a drive voltage source, the drive voltage source configured to apply a first voltage and a second voltage to the liquid crystal; wherein the first voltage and the second voltage cause the liquid crystal to switch between an open state and a closed state. The method further includes directing the emission light and the reflected visible light through the engineered polarizer, wherein the reflected visible light having a wavelength below the light cutoff passes through the polarizer with about 50% attenuation, and the emission light having a wavelength above the light cutoff passes through the polarizer with minimal attenuation. The method further includes: directing polarized visible light and non-polarized emission light to the liquid crystal; applying the first voltage to the liquid crystal to assume the on state for a first video rate sampling duration; and detecting the polarized light at an imaging sensor. The method further includes: applying the second voltage to the liquid crystal to assume the off state and block the polarized light from passing to the imaging sensor; and detecting the non-polarized emission light at the imaging sensor during the off state for a second video rate sampling duration.

[0013] In various aspects of the disclosed method, the minimum attenuation of the emitted light having a wavelength above the light cutoff is 0% attenuation to 15% attenuation. In one aspect, the reflected visible light having a wavelength below the light cutoff passes through the polarizer with an attenuation of about 50% or less. In another aspect, the first video rate sampling duration is between 1 ms and 10 ms, between 2 ms and 20 ms, or 6 ms. In some aspects, the second video rate sampling duration is between 1 ms and 10 ms, between 2 ms and 20 ms, or 6 ms. In some aspects, the first voltage and the second voltage achieve a high electric field and a low electric field on the liquid crystal, respectively.

[0014] In various aspects of the disclosed method, the first voltage and the second voltage switch the liquid crystal between an open state and a closed state. In one aspect, the first voltage switches the liquid crystal to the open state and the second voltage switches the liquid crystal to the closed state, or vice versa. In another aspect, one of the first voltage or the second voltage is 0V. In one aspect, the second voltage is a plurality of voltages including a bipolar square wave having an average value of 0V. In another aspect, the first voltage, the second voltage, or both the first voltage and the second voltage include a plurality of voltages that produce a bipolar square wave having an average value of 0V. In one aspect, the bipolar square wave has an amplitude of + / -0.1 to + / -5 Vp-p, + / -3 to + / -30 Vp-p, + / -16 Vp-p, + / -24 Vp-p or + / -30 Vp-p, or an amplitude between + / -16 V (32 Vp-p) and + / -30 V (60 Vp-p), including any values, or an amplitude between + / -30 V (60 Vp-p) and + / -100 V (200 Vp-p), including any values. In another aspect, the second voltage is a bipolar square wave having an amplitude of + / -0.1 to + / -5 Vp-p, + / -3 to + / -30 Vp-p, + / -16 Vp-p, + / -24 Vp-p, or + / -30 Vp-p, or an amplitude between + / -16 V (32 Vp-p) and + / -30 V (60 Vp-p), any values ​​inclusive, or an amplitude between + / -30 V (60 Vp-p) and + / -100 V (200 Vp-p), any values ​​inclusive. In some aspects, the bipolar square wave amplitude is modulated at the video capture rate of the imaging system. In another aspect, the first voltage is a bipolar square wave having an amplitude of + / -0.1 to + / -5 Vp-p, + / -3 to + / -30 Vp-p, + / -16 Vp-p, + / -24 Vp-p or + / -30 Vp-p, or an amplitude between + / -16 V (32 Vp-p) and + / -30 V (60 Vp-p), including any values, or an amplitude between + / -30 V (60 Vp-p) and + / -100 V (200 Vp-p), including any values.

[0015] In various aspects of the disclosed method, the amplitude is modulated from 500 Hz to 2000 Hz. According to various aspects, the disclosed method further includes heating the liquid crystal using waste heat from the imaging system. In some aspects, the polarizer is a dye polarizer. In another aspect, the polarizer is configured to have a light cutoff in a range that is optimal for a fluorescence imaging process. In various aspects, the polarizer in the liquid crystal assembly is configured to have a light cutoff between about 600 nm and about 850 nm, or in a range of about 700 nm to about 850 nm, about 800 nm to about 950 nm, about 800 nm to about 880 nm, about 775 nm to about 795 nm, or at about 785 nm. In another aspect, the polarizers in the liquid crystal assembly are selected to lose effectiveness or cut off at 700 nm, 725 nm, 750 nm, 775 nm, 780 nm, 785 nm, 790 nm, 795 nm, 800 nm, 805 nm, 810 nm, 815 nm, 820 nm, 825 nm, 830 nm, 835 nm, 840 nm, 850 nm, 855 nm, 860 nm, 865 nm, 870 nm, 875 nm, 880 nm, 885 nm, 890 nm, 895 nm, or 900 nm. In other aspects, the polarizers in the liquid crystal assembly further include two polarizers or more polarizers.

[0016] According to various aspects of the disclosed method, the excitation light is designed to enhance the signal-to-noise ratio (SNR) of the NIR image so that the peak intensity and duration of the excitation light produce a controlled energy excitation that drops below a safety threshold. According to various aspects, the disclosed method further includes directing the emitted light and the reflected visible light to a beam splitter before directing the emitted light and the reflected light to the liquid crystal assembly. In one aspect, the beam splitter has a front surface and a rear surface. In another aspect, the front surface has a front surface coating and the rear surface has a rear surface coating. In one aspect, the front surface coating has a higher P-polarized reflectivity than the rear surface coating. In another aspect, the front surface coating and the rear surface coating produce a P-polarized light front surface to rear surface reflectivity ratio of at least 2:1, at least 4:1, at least 6:1, at least 8:1, at least 10:1, at least 12:1, at least 14:1, at least 16:1, at least 18:1 or at least 20:1. In another aspect, S-polarized reflections of the front and rear surfaces of the beam splitter are minimized by the front surface coating and the rear surface coating. In another aspect, the polarizer is configured to block or attenuate ghosting or secondary reflections reflected from the beam splitter. In one aspect, the first polarizer is configured to block S-polarized light from the beam splitter. In another aspect, the polarizer is configured to allow P-polarized light to pass through the liquid crystal assembly and reach an imaging lens or camera when the liquid crystal is in the open state. In one aspect, the first polarizer allows a front-to-back reflectivity ratio of the beam splitter to be at least 2:1, 4:1, 6:1, 8:1, 10:1, 12:1, 15:1, 17:1 or 20:1 to pass therethrough and reach an imaging lens and camera. In some aspects, the total reflectivity of the beam splitter is about 11% or less, about 10% or less, about 9% or less, about 8% or less, about 7% or less, about 6% or less, about 5% or less, about 4% or less, or about 3% or less.

[0017] The present disclosure also relates to a liquid crystal assembly comprising any of the features described herein, alone or in combination with any other features, in any configuration.

[0018] The present disclosure also relates to a method for imaging emission light from a fluorophore at an imaging system, the imaging system comprising a liquid crystal assembly comprising any of the features described herein in any configuration, alone or in combination with any other features.

[0019] The present disclosure also relates to a liquid crystal assembly for use with an imaging system having a light source for providing excitation light. The liquid crystal assembly includes: a first polarizer, the first polarizer is arranged in the path of light from the light source; the first polarizer is configured to have a light cutoff in a range outside its polarization range; a liquid crystal, the liquid crystal is arranged in the path of light transmitted through the first polarizer; and a second polarizer, the second polarizer is arranged in the path of light transmitted through the liquid crystal. The second polarizer is configured to have a light cutoff in a range outside its polarization range. The liquid crystal assembly also includes a driving voltage source, the driving voltage source is configured to apply a first voltage and a second voltage to the liquid crystal. The first voltage and the second voltage switch the liquid crystal between an open state and a closed state.

[0020] In various aspects, the first polarizer and the second polarizer are cross-polarized. In one aspect, when one of the first voltage or the second voltage is applied to the liquid crystal, the liquid crystal is in the open state. In another aspect, one of the first voltage or the second voltage is 0V. In some aspects, when one of the first voltage or the second voltage is applied to the liquid crystal, the liquid crystal is in the closed state. In another aspect, the second voltage is a bipolar square wave. In one aspect, the bipolar square wave has an amplitude of + / -0.1 to + / -5Vp-p, + / -3 to + / -30Vp-p, + / -16Vp-p, + / -24Vp-p, or + / -30Vp-p.

[0021] In various aspects, the liquid crystal assembly further includes: an imaging lens located in the path of light transmitted through the second polarizer; and a camera located in the path of light transmitted through the imaging lens. In another aspect, when the liquid crystal is in the open state, the liquid crystal rotates the polarization of polarized visible light transmitted through the first polarizer, thereby allowing the polarized visible light to pass through the second polarizer and reach the imaging lens with an attenuation of about 50% or less. In some aspects, when the liquid crystal is in the closed state, the liquid crystal does not rotate the polarization of the polarized visible light, and the polarized visible light transmitted through the first polarizer is blocked by the second polarizer.

[0022] In various aspects, the light cutoff range of the first polarizer and the second polarizer is a range where the polarizer extinction ratio is poor or minimal. In another aspect, light outside the light cutoff range of the first polarizer and the second polarizer passes through the first polarizer and the second polarizer without attenuation or with minimal attenuation and reaches the imaging lens. In one aspect, the minimum attenuation is 15% or less. In some aspects, the light cutoff range of the first polarizer is about 700nm to about 800nm ​​or about 700nm, 725nm, 750nm, 775nm, 780nm, 785nm, 790nm, 795nm, 800nm, 805nm, 810nm, 815nm, 820nm, 825nm, 830nm, 835nm, 840nm, 850nm, 855nm, 860nm, 865nm, 870nm, 875nm, 880nm, 885nm, 890nm, 895nm or 900nm. In another aspect, the light cutoff range of the second polarizer is about 700nm to about 800nm ​​or about 700nm, 725nm, 750nm, 775nm, 780nm, 785nm, 790nm, 795nm, 800nm, 805nm, 810nm, 815nm, 820nm, 825nm, 830nm, 835nm, 840nm, 850nm, 855nm, 860nm, 865nm, 870nm, 875nm, 880nm, 885nm, 890nm, 895nm or 900nm. In another aspect, the light cutoff range of the first polarizer and the second polarizer is higher than the longest wavelength of the visible spectrum.

[0023] According to various aspects, when the liquid crystal is in the open state, the liquid crystal assembly is configured to view visible light. In another aspect, when the liquid crystal is in the closed state, the liquid crystal assembly is configured to block visible light. In one aspect, the blocked visible light allows fluorescence imaging from the fluorophore. In some aspects, the fluorophore provides emission light emitted by the fluorophore. In one aspect, the fluorophore provides excitation light. In some aspects, the excitation light is white light, NIR light, IR light, or any other type of excitation light. In one aspect, the excitation light is near infrared light provided by a laser diode.

[0024] In various aspects, the liquid crystal assembly further includes a beam splitter. In some aspects, the beam splitter has a front surface and a rear surface. In one aspect, the front surface has a front surface coating, and the rear surface has a rear surface coating. In some aspects, the front surface coating has a P polarization reflection higher than the rear surface coating. In another aspect, the front surface coating and the rear surface coating produce a P polarized light front surface to rear surface reflectivity ratio of at least 2:1, at least 4:1, at least 6:1, at least 8:1, at least 10:1, at least 12:1, at least 14:1, at least 16:1, at least 18:1 or at least 20:1. In one aspect, the S polarization reflection of the front surface and the rear surface of the beam splitter is minimized by the front surface coating and the rear surface coating. In another aspect, the first polarizer is configured to block or attenuate ghosting or secondary reflections reflected from the beam splitter. In some aspects, the first polarizer is configured to block S polarized light from the beam splitter. In another aspect, the first polarizer and the second polarizer are configured to allow P-polarized light to pass through the liquid crystal assembly and reach an imaging lens or camera when the liquid crystal is in the open state. In one aspect, the first polarizer allows a front-to-back reflectivity ratio of the beam splitter of at least 2:1, 4:1, 6:1, 8:1, 10:1, 12:1, 15:1, 17:1, or 20:1 to pass therethrough and reach an imaging lens and camera. In another aspect, the total reflectivity of the beam splitter is about 11% or less, about 10% or less, about 9% or less, about 8% or less, about 7% or less, about 6% or less, about 5% or less, about 4% or less, or about 3% or less.

[0025] In various aspects, the imaging lens and the camera are configured to view fluorescent images using a video rate sampling duration of between 1 ms and 10 ms, between 2 ms and 20 ms, or 6 ms. In one aspect, the fluorescent images have reduced motion blur.

[0026] The present disclosure also relates to a method for imaging emission light emitted by a fluorophore at an imaging system including a liquid crystal assembly. The method includes: allowing or directing visible light to reach a sample; directing excitation light to the sample; and directing the emission light and reflected visible light to the liquid crystal assembly. The liquid crystal assembly includes a first engineered polarizer located in the path of light from the sample. The first engineered polarizer is configured to have a light cutoff between 600nm and 900nm or in the range of 800nm ​​to 880nm. The liquid crystal assembly also includes a liquid crystal, which is arranged in the path of light transmitted through the first engineered polarizer. The liquid crystal assembly also includes a second engineered polarizer located in the path of light transmitted through the liquid crystal. The second engineered polarizer is configured to have a light cutoff between 600nm and 900nm or in the range of 800nm ​​to 880nm. In one aspect, the first engineered polarizer and the second engineered polarizer are cross-polarized. The liquid crystal assembly also includes a driving voltage source configured to apply a first voltage and a second voltage to the liquid crystal; wherein the first voltage and the second voltage switch the liquid crystal between an open state and a closed state. In one aspect, the disclosed method also includes directing the emitted light and the reflected visible light through the first engineered polarizer, wherein the first engineered polarizer polarizes the reflected visible light below the light cutoff range, thereby generating polarized reflected visible light. The disclosed method further includes: directing the emitted light and the polarized reflected visible light through the liquid crystal; applying the first voltage to the liquid crystal to present the open state within a first video rate sampling duration; and using the liquid crystal in the open state to rotate the polarization of the polarized reflected visible light. The disclosed method further includes directing the emitted light and the polarized reflected visible light through the second engineered polarizer, wherein the second engineered polarizer allows the polarized reflected visible light to pass through the second engineered polarizer. The disclosed method further includes detecting at an imaging sensor a first portion of the polarized reflected visible light that passes through the second engineered polarizer during the open state. The disclosed method further includes: applying the second voltage to the liquid crystal to present the off state for a second video rate sampling duration; allowing the emitted light and the polarized reflected visible light to pass through the liquid crystal without rotating the emitted light and the polarized reflected visible light; blocking the polarized reflected visible light with the second engineered polarizer; and detecting a portion of the emitted light during the off state at the imaging sensor.

[0027] In various aspects of the disclosed method, the method further comprises blocking secondary reflections of the reflected visible light using the first polarizer. In one aspect, the excitation light is provided to the sample via a laser diode.

[0028] In various aspects of the disclosed method, the method further includes: turning off the laser diode to stop providing excitation light; providing the second voltage to the liquid crystal to present the off state within a third video rate sampling duration; detecting a dark background image of the sample; and subtracting the dark background image of the sample from the detected portion of the emitted light to produce a fluorescent image.

[0029] In various aspects of the disclosed method, the first video rate sample duration is about 2 ms to about 10 ms. In one aspect, the second video rate sample duration is about 8 ms to about 16 ms. In another aspect, the third video rate sample duration is about 8 ms to about 16 ms.

[0030] In various aspects of the disclosed method, one of the first voltage or the second voltage is about 0V. In another aspect, the second voltage is a bipolar square wave. In one aspect, the bipolar square wave has an amplitude of + / -0.1 to + / -5Vp-p, + / -3 to + / -30Vp-p, + / -16Vp-p, + / -24Vp-p, or + / -30Vp-p, or an amplitude between + / -16V (32Vp-p) and + / -30V (60Vp-p), including any value, or an amplitude between + / -30V (60Vp-p) and + / -100V (200Vp-p), including any value.

[0031] In various aspects of the disclosed method, the fluorescent image has reduced motion blur. In one aspect, the second video rate sampling duration and the third video rate sampling duration allow post-processing using digital gain. In another aspect of the disclosed method, the disclosed method further includes orienting the first engineered polarizer to block undesired polarized light. In one aspect, the second video rate sampling duration and the third video rate sampling duration enhance the signal-to-noise ratio (SNR) and contrast-to-noise ratio (CNR) of the fluorescent image. In one aspect, the enhanced CNR achieves a fast frame rate. In another aspect, the disclosed method provides real-time viewing of the reflected visible light and fluorescent images.

[0032] In various aspects of the disclosed method, the disclosed method further includes directing the emitted light and the reflected light to a beam splitter before directing the emitted light and the reflected light to the liquid crystal assembly. In one aspect, the beam splitter has a front surface and a rear surface. In another aspect, the front surface has a front surface coating and the rear surface has a rear surface coating. In some aspects, the front surface coating has a higher P-polarized reflection than the rear surface coating. In one aspect, the front surface coating and the rear surface coating produce a P-polarized light front surface to rear surface reflectivity ratio of at least 2:1, at least 4:1, at least 6:1, at least 8:1, at least 10:1, at least 12:1, at least 14:1, at least 16:1, at least 18:1 or at least 20:1. In another aspect, the S-polarized reflection of the front surface and the rear surface of the beam splitter is minimized by the front surface coating and the rear surface coating. In another aspect, the first polarizer is configured to block or attenuate ghosting or secondary reflections reflected from the beam splitter. In one aspect, the first polarizer is configured to block S-polarized light from the beam splitter. In another aspect, the first polarizer and the second polarizer are configured to allow P-polarized light to pass through the liquid crystal assembly and reach the imaging lens and / or camera when the liquid crystal is in the open state. In some aspects, the first polarizer allows a front-to-back reflectivity ratio of the beam splitter of at least 2:1, 4:1, 6:1, 8:1, 10:1, 12:1, 15:1, 17:1, or 20:1 to pass therethrough and reach the imaging lens and camera. In various aspects, the total reflectivity of the beam splitter is about 11% or less, about 10% or less, about 9% or less, about 8% or less, about 7% or less, about 6% or less, about 5% or less, about 4% or less, or about 3% or less.

[0033] The present disclosure also relates to a method of imaging abnormal tissue, cancer, tumor, vasculature or structure in a sample from a subject, the method comprising generating an image of the vasculature or structure by imaging fluorescence using the liquid crystal assembly disclosed herein.

[0034] The present disclosure also relates to a method of imaging abnormal tissue, cancer, tumor, vasculature, or structure in a sample from a subject according to any of the methods disclosed herein, the method comprising generating an image of the abnormal tissue, cancer, tumor, vasculature, or structure by fluorescence imaging using an imaging system including a liquid crystal assembly.

[0035] In various aspects, the fluorescence imaged is autofluorescence, a contrast or imaging agent, a chemical agent, a radiolabeled agent, a radiosensitizer, a photosensitizer, a fluorophore, a therapeutic agent, an imaging agent, a diagnostic agent, a protein, a peptide, a nanoparticle or a small molecule, or any combination thereof, or any combination thereof. In one aspect, the fluorescence imaged is autofluorescence, a contrast or imaging agent, a chemical agent, a radiolabeled agent, a radiosensitizer, a photosensitizer, a fluorophore, a therapeutic agent, an imaging agent, a diagnostic agent, a protein, a peptide, a nanoparticle or a small molecule, or any combination thereof. In one aspect of the disclosed method, the method further comprises administering a contrast or imaging agent to the subject.

[0036] The present disclosure also relates to a method of imaging abnormal tissue, cancer, tumor, vasculature or structure in a fluorophore from a subject using any of the disclosed liquid crystal assemblies. The method comprises: administering a contrast or imaging agent to the subject; and generating an image of the abnormal tissue, cancer, tumor, vasculature or structure by imaging the contrast or imaging agent using an imaging system.

[0037] The present disclosure also relates to a method of imaging abnormal tissue, cancer, tumor, vasculature, or structure in a fluorophore from a subject according to any disclosed method. The disclosed method further comprises: administering a contrast or imaging agent to the subject; and generating an image of the abnormal tissue, cancer, tumor, vasculature, or structure by imaging the contrast or imaging agent using an imaging system. In one aspect, the contrast or imaging agent comprises a dye, a fluorophore, a fluorescent biotin compound, a luminescent compound, a chemiluminescent compound, or any combination thereof.

[0038] In various aspects of the disclosed method, the contrast or imaging agent further comprises a protein, a peptide, an amino acid, a nucleotide, a polynucleotide or any combination thereof. In one aspect, the contrast or imaging agent further comprises tozuleristide. In another aspect, the contrast or imaging agent absorbs a wavelength between about 200nm and about 900nm.In other aspects, the contrast or imaging agent includes: DyLight-680, DyLight-750, VivoTag-750, DyLight-800, IRDye-800, VivoTag-680, Cy5.5 or indocyanine green (ICG) and any derivatives of the foregoing; fluorescein and fluorescent dyes (e.g., fluorescein isothiocyanate or FITC, naphthyl fluorescein, 4′,5′-dichloro-2′,7′-dimethoxyfluorescein, 6-carboxyfluorescein or FAM, etc.), carbocyanine, merocyanine, styryl dye, oxonol dye, phycoerythrin, erythrosine, eosin, rhodamine dye (e.g., tetramethylcarboxy- Rhodamine or TAMRA, carboxyrhodamine 6G, carboxy-X-rhodamine (ROX), lissamine rhodamine B, rhodamine 6G, rhodamine green, rhodamine red, tetramethylrhodamine (TMR), etc.), coumarin and coumarin dyes (e.g., methoxycoumarin, dialkylaminocoumarin, hydroxycoumarin, aminomethylcoumarin (AMCA), etc.), Oregon green dyes (e.g., Oregon Green 488, Oregon Green 500, Oregon Green 514, etc.), Texas Red, Texas Red-X, Spectrum Red, Spectrum Green, cyanine dyes (e.g., CY-3, Cy-5, CY-3.5, CY-5.5, etc.), ALEXAFLUOR dyes (e.g., ALEXA FLUOR 350, ALEXA FLUOR 488, ALEXA FLUOR 532, ALEXA FLUOR546, ALEXA FLUOR 568, ALEXA FLUOR 594, ALEXA FLUOR 633, ALEXA FLUO R660, ALEXA FLUOR 680, etc.), BODIPY dyes (e.g., BODIPY FL, BODIPY R6G, BODIPY TMR, BODIPY TR, BODIPY530 / 550, BODIPY 558 / 568, BODIPY 564 / 570, BODIPY 576 / 589, BODIPY 581 / 591, BODIPY630 / 650, BODIPY 650 / 665, etc.), IRD dyes (e.g., IRD40, IRD 700, IRD 800, etc.), 7-aminocoumarin, dialkylaminocoumarin reactive dyes, 6,8-difluoro-7-hydroxycoumarin fluorophores, hydroxycoumarin derivatives, alkoxycoumarin derivatives, succinimide esters, pyrene succinimide esters, pyridyl oxazole derivatives, aminonaphthyl dyes, dansyl chloride, dapoxyl dyes, Dapoxyl sulfonyl chloride, amine-reactive Dapoxyl succinimide esters, carboxylic acid-reactive Dapoxyl (2-aminoethyl) sulfonamide), bimane dyes, bimane thioglycolic acid, NBD dyes, QsY 35, or any combination thereof.

[0039] In various aspects of the disclosed methods, administration includes intravenous administration, intramuscular administration, subcutaneous administration, intraocular administration, intraarterial administration, peritoneal administration, intratumoral administration, intradermal administration, or any combination thereof. In one aspect, the imaging includes tissue imaging, ex vivo imaging, intraoperative imaging, or any combination thereof. In one aspect, the sample is in an in vivo sample, an in situ sample, an ex vivo sample, or an intraoperative sample. In another aspect, the sample is an organ, an organ substructure, a tissue, or a cell. In various aspects, the sample is autofluorescent. In one aspect, the autofluorescence of the sample includes an ocular fluorophore, tryptophan, or a protein present in a tumor or malignancy. In one aspect, the method is used to visualize vascular flow or vascular patency.

[0040] In various aspects of the disclosed method, the abnormal tissue, cancer, tumor, vasculature or structure comprises a blood vessel, lymphatic vasculature, neuronal vasculature or CNS structure. In one aspect, the imaging is angiography, arteriography, lymphangiography or cholangiography. In another aspect, the imaging comprises detecting a vascular abnormality, a vascular malformation, a vascular lesion, an organ or an organ substructure, a cancer or a diseased area, a tissue, a structure or a cell. In another aspect, the vascular abnormality, vascular malformation or vascular lesion is an aneurysm, an arteriovenous malformation, a cavernous vascular malformation, a venous malformation, a lymphatic malformation, a telangiectasia, a mixed vascular malformation, a spinal dural arteriovenous fistula or a combination thereof. In one aspect, the organ or organ substructure is the brain, heart, lung, kidney, liver or pancreas.

[0041] In various aspects of the disclosed method, the disclosed method further comprises performing surgery on the subject. In one aspect, the surgery comprises angioplasty, cardiovascular surgery, aneurysm repair, valve replacement, aneurysm surgery, arteriovenous malformation or cavernous malformation surgery, venous malformation surgery, lymphatic malformation surgery, telangiectasia surgery, mixed vascular malformation surgery or dural arteriovenous fistula surgery, repair or bypass, arterial bypass, organ transplantation, plastic surgery, ophthalmic surgery, reproductive system surgery, stent insertion or replacement, plaque ablation, removal of the cancer or diseased area, tissue, structure or cells of the subject, or any combination thereof.

[0042] In various aspects of the disclosed method, the imaging includes imaging of vascular abnormalities, cancer or diseased areas, tissues, structures or cells of the subject after surgery. In various aspects, the disclosed method further includes treating cancer in the subject. In one aspect, the disclosed method further includes repairing intracranial CNS vascular defects, spinal CNS vascular defects; peripheral vascular defects; removal of abnormal vascularized tissue; eye imaging and repair; anastomosis; reconstruction or plastic surgery; ablation or treatment of atherosclerotic plaques or restenosis; repair or resection (including selective resection), retention (including selective retention) of important organs or structures such as nerves, kidneys, thyroid, parathyroid, liver segments or ureters; identification and management during surgery (sometimes retention, sometimes selective resection); diagnosis and treatment of limb ischemia; or treatment of chronic wounds. BRIEF DESCRIPTION OF THE DRAWINGS

[0043] Figure 1 is a diagram of a liquid crystal shutter assembly shown with a camera according to one aspect.

[0044] Figure 2 The spectral response of an exemplary camera according to one aspect is shown.

[0045] Figure 3 The spectral response of an illustrative camera to light attenuated by a long pass filter is shown according to one aspect.

[0046] Figure 4 The change in the extinction ratio of light in the open state and the closed state of the liquid crystal cell according to one aspect is shown.

[0047] Figure 5 The off-state transmittance of a liquid crystal cell in combination with various polarizers according to one aspect is shown.

[0048] Figure 6 The liquid crystal cell transition speed in response to different applied voltages according to one aspect is shown.

[0049] Figure 7 Liquid crystal cell transition speeds at different temperatures according to one aspect are shown.

[0050] Figure 8 Optical blocking in a liquid crystal cell in response to an applied voltage is shown according to one aspect.

[0051] Fig. 9 Shown is the average power applied by pulsed or controlled excitation energy applied to a tissue sample according to one aspect versus the average power of exemplary clinical levels of energy applied to the tissue sample.

[0052] Fig.10An exemplary signal-to-noise ratio obtained using a pulsed or controlled excitation light source according to one aspect compared to an exemplary clinical laser excitation light is shown.

[0053] Fig.11 A liquid crystal shutter assembly according to one aspect is shown.

[0054] Fig.12 The transmittance of a liquid crystal medium according to one aspect is shown in the open state and in the closed state.

[0055] Fig.13 A diagram showing a liquid crystal shutter assembly shown with a camera according to one aspect.

[0056] Fig.14A A visible light image captured using a liquid crystal shutter assembly in an open state according to one aspect is shown.

[0057] Fig. 14B A visible light image captured using a liquid crystal shutter assembly in a closed state according to one aspect is shown.

[0058] Fig.15 A contrast-to-noise ratio using a driving voltage supplied to a liquid crystal assembly according to one aspect is shown.

[0059] Fig.16 An exposure ratio necessary for post-processing via digital gain using a driving voltage supplied to a liquid crystal assembly at a contrast-to-noise ratio of 5 according to one aspect is shown.

[0060] Fig.17 A diagram illustrating a method for shortening the integration time of a fluorescent image using a liquid crystal shutter assembly according to one aspect.

[0061] Fig.18A An image of a fluorescent cube moving on a linear stage taken using bright background subtraction according to one aspect is shown.

[0062] Fig.18B An image of a fluorescent cube moving on a linear stage captured using a liquid crystal shutter assembly according to one aspect is shown.

[0063] Fig.19 Shown is the use of a liquid crystal shutter assembly configured to visualize a peptide-fluorophore conjugate (tocillin peptide).

[0064] FIG. 20A to FIG. 20B shows a front-to-back reflectivity ratio (FBR) of approximately 10:1 in the visible spectrum ( Fig. 20A ) and the total reflectivity ( Fig. 20BIf the FBR of the beam splitter designed to be filtered by the LC shutter is for a different FBR (e.g., an FBR of about 20:1), the FBR curve will still vary with wavelength, but will average 20 over the visible spectrum.

[0065] Fig. 20A The FBR of a beam splitter designed to be filtered by an LC shutter in the visible spectrum (i.e., between 400nm and 700nm wavelengths) is shown for S polarized light (FBR S) or P polarized light (FBR P), respectively. Note that in this example, the minimum FBR is 10:1.

[0066] Fig. 20B The effect of the polarizer on the total reflectivity ("total reflectivity") of a beam splitter designed to be filtered by an LC shutter within the visible spectrum (i.e., between 400nm and 700nm wavelengths) is shown compared to the minimum reflectivity (11%) without using a polarization filter. DETAILED DESCRIPTION

[0067] Multispectral imaging is an important technology for observing the spectral characteristics of light in a single image. The most common example of multispectral imaging is color vision. The rods and cones of the human eye have three different chemical compositions that allow different colors of light to be captured. The three colors: red, green and blue, are then displayed as a monochrome image in the human mind. Digital color cameras simulate the biological characteristics of the eye by using different color filters for each pixel of the sensor. Dedicating different pixels to different colors effectively trades spatial resolution for spectral information. The advantages of using color bands to add information to the image can be applied to imaging technology. In addition, the capabilities of multispectral imaging can be extended to wavelengths of light that the naked eye cannot directly see. As the range of spectral information expands, more and more information can be compressed into a visible image. An interesting application of multispectral imaging is to mix visible light (VIS) images and near infrared (NIR) images. This combination is supported by CCD (charge coupled device) and CMOS (complementary metal oxide semiconductor) silicon cameras with good sensitivity in the visible range and the NIR range. The two spectral bands can be mixed using multiple cameras or using a single camera with a dedicated NIR pixel filter on the sensor, for example. In addition, in some embodiments, there are one or more sensors in the system that detect these changing spectral bands. Alternatively, time multiplexing allows a single image sensor to capture full-resolution images in each spectral band, but at the expense of time, because each image must be captured continuously. Filter wheels or motorized mirrors are traditionally used to switch filters, but are usually too slow for the optimal video rate switching speed required for real-time imaging. Time multiplexing can also be applied to multiple imaging sensors in a system (e.g., a multi-camera or multi-sensor system), but the same time cost and video rate limitations also apply.

[0068] Multiplexing spectral bands is also used for fluorescence imaging. Fluorescence is an energy transfer mechanism where light of one color (wavelength) is absorbed and subsequently released at a different color (wavelength). The emitted photons are typically lower in energy and therefore longer in wavelength than the absorbed photons (Stokes shift). Fluorescence is typically observed by the human eye when the excitation wavelength is in the UV and therefore not perceptible to the human eye, but then emitted in the visible. Fluorescence is also observed by instruments and sensors where the excitation wavelength is in any part of the spectrum that the instrument or sensor cannot perceive, but then emits a wavelength that is within the range of the instrument or sensor designed to receive a detectable fluorescence emission signal.

[0069] Fluorophores are molecules that absorb light and then emit it, so specific fluorophores are responsible for the excitation and emission spectral bands. For example, one class of fluorophores, including the cyanine dyes and indocyanine green dye family (ICG), absorbs and emits light in the near-infrared spectrum. These NIR fluorophores are particularly important when imaging biological samples, in part because their absorption and emission spectra are both located near 830nm, which is considered the main "biological window" for transmission in tissues. ICG can also be used in medical and diagnostic applications. Combining fluorescent images with visible spectrum images can be an insightful tool for such biological applications. Infrared and near-infrared are wavelength ranges that are particularly beneficial for medical imaging, but other fluorescent dyes and parts can be used in detection systems using the liquid crystal shutter assembly described herein.

[0070] In addition, video image processing can be used to capture fluorescence imaging in real time or near real time to detect changes over time. In many cases, video rate switching of NIR fluorescence with visible wavelengths may require multiple sensors or cameras, while in some cases a single camera system can be used. In addition, when used during NIR fluorescence, the excitation of the fluorophore can be turned on and off to control a known time state, thereby allowing background subtraction methods. The liquid crystal ("LC") shutter is an assembly consisting of two linear polarization filters with a liquid crystal medium between the two filters. The polarization filter element is capable of blocking light with a polarization different from the orientation of the filter. The liquid crystal material does not block light, but is capable of rotating the polarization of light passing through the material. The orientation of the crystals in the LC material will change the degree of polarization rotation of the light, so whether the polarization of the light passing through is rotated or not, the orientation of the crystals will change. The combination of the two filters and the LC medium and an applied electric field can be assembled to produce a shutter that will block light or transmit light depending on the applied electric field. When the polarization orientation of the light passing through the LC material is aligned with the exit polarizer, the light will pass with minimal attenuation. This situation is considered to be the open state of the shutter. When the electric field is adjusted to change the polarization orientation of the light so that it is rotated 90° relative to the exit polarizer, the light will be blocked. This is considered the closed state of the LC shutter assembly.

[0071] Due to the spectral bandwidth limitation of the polarization element incorporated into the liquid crystal shutter assembly, light outside the high extinction ratio range of the polarization element (e.g., outside the spectral bandwidth limitation, also referred to as out-of-band light) is conventionally blocked or redirected by other spectral elements (filters) before being incident on the first polarization element. By using these additional spectral filters, the extinction ratio of the liquid crystal shutter can be guaranteed.

[0072] In the method and system of the present disclosure, instead of diverting out-of-band incident light away from the liquid crystal assembly as would normally occur, out-of-band light is intentionally allowed to be incident on the polarizing element and thus on the liquid crystal shutter assembly. The out-of-band light is not affected by the orientation of the polarizer or liquid crystal medium and therefore passes through the shutter. Traditionally, designing the optical system in this way would produce a poor or minimal extinction ratio, thus destroying the function of the liquid crystal shutter assembly. In the method and system of the present disclosure, the spectral characteristics of the polarizing element are selected so that the light leaking through the shutter assembly is designed to be aligned with the NIR emission of the fluorophore, so the leaking light is intentional and beneficial. This configuration is contrary to traditional good practices for liquid crystal shutter applications. The liquid crystal shutter is used to attenuate light in a controllable manner by adjusting the level of the applied electric field. Liquid crystal shutters used in this manner have a limited effective spectral range, which is most commonly limited by the polarization technology of the polarization filter. According to various aspects, the liquid crystal shutter assembly of the present disclosure utilizes various wavelength ranges that cannot be polarized by the polarization filter. Alternatively, a polarization beam splitter can be used in conjunction with a polarization filter or in place of a polarization filter.

[0073] Wavelengths of light that are not successfully polarized by a linear polarization filter are typically considered to be outside the operating range of a liquid crystal shutter assembly. Rather than operating within the typical operating range of a liquid crystal shutter assembly, the disclosed liquid crystal shutter assembly utilizes various wavelength ranges outside the effective range of a linear polarizer, thus utilizing a spectral range that was previously considered detrimental in typical liquid crystal applications.

[0074] The terms "liquid crystal," "LC," "LC cell," "LC medium," "LC material," "liquid crystal cell," "liquid crystal material," or "liquid crystal medium" and the like are used interchangeably herein to refer to non-limiting liquid crystal materials, media, or cells, including, for example, materials, media, or cells comprising thermotropic and lyotropic liquid crystals, Pi cells, twisted nematic ("TN") cells, liquid crystal displays (LCDs), liquid crystals having photoconductor properties, or liquid crystals that are easily developed and adjusted under an electric field. It should be understood that liquid crystals can exist in states between a crystalline (solid) state and an isotropic (liquid) state. There are many types of liquid crystal states, depending on the amount of order in the liquid crystal material.

[0075] The terms "liquid crystal shutter," "liquid crystal assembly," "liquid crystal shutter assembly," "LC shutter," "LC assembly," or "LC shutter assembly" are used interchangeably herein to refer to a liquid crystal associated with, used in conjunction with, used with, including, or housing one or more polarizers. Exemplary LC shutters are described herein.

[0076] The terms "polarizing," "polarizer," or "polarizing," particularly when used in describing a component or element, should be understood to describe various components or elements that are functionally interchangeable and can be used to transmit light of a specific polarization orientation. For example, the LC assemblies described herein can use various polarizing component elements, including but not limited to one or more polarizing beam splitters, polarizing filters, alone or in combination with other polarizing components or elements.

[0077] The terms "excitation energy" and "excitation light" are used interchangeably in the present disclosure and are intended to provide excitation light to a fluorophore, or to produce emission light from a fluorophore. Excitation energy or excitation light is also used to illuminate a fluorophore or other substance or molecule and cause emission light.

[0078] In the context of images, videos, frame rates and video rates, the terms "video rate", "video rate imaging", "video frame rate", "frame rate" or "real time" and "real time", particularly when used to describe the speed, visualization and clarity of an image or imaging system, refer to the number of unique or different frames or images in the time used. Video rate is usually measured in frames per second ("fps"), which defines how many frames or images there will be in a video in any given second. Video rate can also be measured in bit rate standards. Real-time frame rates can be about 25fps, or between 30 and 50fps. Real-time video frame rates can also be between 16 and 120fps (including the end values). Video rate is a measurement that affects the system's responsiveness to motion. For example, the higher the video rate, the clearer the image on the moving image. A higher frame rate means that more images are displayed in one second, thereby producing smoother and clearer video quality, and may provide clearer edges for the image.

[0079] The terms "video rate sampling duration" and "exposure time" may be used interchangeably in the present disclosure. The video rate sampling duration or exposure time may be the duration for which an imaging sensor or camera images a sample. The video rate sampling duration may also be described with respect to the LC shutter and LC assembly described herein, for example, when describing the switching rate between an open state and a closed state (i.e., the period between an open state and a closed state) or when two voltage sources or drive voltage sources are switched in a system. The voltage supplied to the system may cause the LC shutter to switch between an open state and a closed state with a video rate sampling duration ranging between 1 ms and 10 ms, between 2 ms and 20 ms, or about 1 ms, 2 ms, 3 ms, 4 ms, 5 ms, 6 ms, 7 ms, 8 ms, 9 ms, 10 ms, 11 ms, 12 ms, 13 ms, 14 ms, 15 ms, 16 ms, 17 ms, 18 ms, 19 ms, 20 ms. The shorter switching time between the open state and the closed state in the LC shutter and LC assembly described herein allows for a shorter video rate sampling duration, effectively increasing the video rate, thereby displaying more images in one second, thus resulting in smoother and clearer video quality. The shorter video rate sampling duration results in a higher video rate, which also results in improved video quality, reduced motion blur, clearer images, and clear image boundaries, and reduced artifacts.

[0080] The liquid crystal shutter assemblies and methods disclosed herein may be used in various imaging systems and methods. By way of example and not limitation, the liquid crystal shutter assembly of the present disclosure may be incorporated into one or more systems and methods for fluorescence imaging disclosed in the following cases: PCT / US2014035203, entitled “Systems and methods for recording simultaneously visible light image and infrared light image from fluorophores,” published as WO 2014 / 176375 on October 30, 2014; PCT / US2019 / 024689, entitled “Systems and methods for simultaneous near-infrared light and visible light imaging,” published as WO 2019 / 191497 on October 3, 2019; PCT / US2020 / 053746, entitled “Systems and methods for vascular and structural imaging,” published as WO 2021 / 067563 on April 8, 2021; and PCT / US2020 / 053747, entitled “Systems and methods for vascular and structural imaging,” published as WO 2021 / 067563 on December 30, 2021. 2021 / 263159, entitled “Systems and methods for simultaneous near-infrared light and visible light imaging” and PCT / US2021 / 039177 (collectively, the “Blaze Bioscience system”), each of which is incorporated herein by reference in its entirety. It should also be understood that the liquid crystal shutter assembly of the present disclosure may be incorporated into any one or more systems and methods for fluorescence imaging known in the art.

[0081] In one aspect of the present disclosure, it is desirable to use the liquid crystal shutter assembly 100 in conjunction with at least one sensor or camera 102 having sufficient sensitivity in the visible and NIR spectral ranges. Figure 1 As shown, the LC shutter assembly 100 further includes a polarizer 104 and a liquid crystal medium 106 within the LC shutter assembly 100 .

[0082] According to one embodiment, two polarizers 104 are integrated into the LC shutter 100, such as Figure 1106. In other embodiments, one or more polarizers 104 may be separate components located in the same optical path and aligned on either side of the LC medium cell 106. In still other embodiments, multiple LC medium cells 106, each associated with one or more polarizers 104, may be used to form the LC assembly 100. Such stacking of polarizers and liquid crystal media may be used to improve the extinction ratio compared to a single-cell LC shutter assembly. Alternatively, one or more LC assemblies 100 including one or more LC cells 106 and one or more polarizers 104 may be used in combination with any combination of additional LC cells 106 and / or polarizers 104.

[0083] According to one embodiment, the two polarizers 104 may each include a substrate layer 1102(a), 1102(b), such as Fig.11 As shown. The first substrate layer 1102 (a) of the first polarizer and the second substrate layer 1102 (b) of the second polarizer may be glass substrate layers. The glass substrate layers may be configured to allow light to pass through. Fig.11 As shown, a first polarizing film 1101(a) can be adhered to a first substrate layer 1102(a), and a second polarizing film 1101(b) can be adhered to a second substrate layer 1102(b). The first polarizing film 1101(a) and the second polarizing film 1101(b) can be cross-polarized (e.g., have opposite orientations).

[0084] In one aspect, the first substrate layer 1101(a) may have a conductive layer 1103(a) on a side of the substrate layer 1102(a) opposite the first polarizing film 1101(a) and adjacent to the LC medium 106. The second substrate layer 1102(b) may have a conductive layer 1103(b) on a side of the substrate layer 1102(b) opposite the second polarizing film 1101(b) and adjacent to the LC medium 106.

[0085] In one aspect, both the first conductive layer 1103(a) and the second conductive layer 1103(b) may include indium tin oxide ("ITO"). As used herein, it is understood that a transparent conductive film or layer includes any thin film of an optically transparent and conductive material. In other examples, the first conductive layer 1103(a) may include any transparent conductive film or layer, including a transparent conductive film or layer including: indium tin oxide ("ITO"), a low or high resistance transparent conductive film, a transparent conductive oxide ("TCO"), indium zinc oxide ("IZO"), other tin oxide (Sno2) based films (including undoped tin dioxide without indium), zinc oxide (ZnO) films doped with Cu and Ga ("GZO"), aluminum zinc oxide ("AZO"), and cadmium stannate ("CTO"). The second conductive layer 1103(b) may include any transparent conductive film or layer, including transparent conductive films or layers including: indium tin oxide ("ITO"), low or high resistance transparent conductive films, transparent conductive oxides ("TCO"), indium zinc oxide ("IZO"), other tin oxide (Sno2) based films (including undoped tin dioxide without indium), zinc oxide (ZnO) films doped with Cu and Ga ("GZO"), aluminum zinc oxide ("AZO"), and cadmium stannate ("CTO"). In some examples, the first conductive layer 1103(a) and the second conductive layer 1103(b) may include the same material. In other examples, the first conductive layer 1103(a) and the second conductive layer 1103(b) may include different materials. The first conductive layer 1103(a) may be soldered to the first substrate layer 1102(a). The second conductive layer 1103(b) may be soldered to the second substrate layer 1102(b).

[0086] In another aspect, the first conductive layer 1103(a) and the second conductive layer 1103(b) can be kept thin to maintain high transmission of NIR light through the LC shutter 100. In some embodiments, the conductive layer (whether the first layer or the second layer) can be 10-50 nm thick (inclusive), less than 25 nm thick, less than 50 nm thick, 50 nm to 250 nm thick, 75 nm to 165 nm thick, or more than 250 nm thick, depending on the material used for the transparent conductive film or layer. In some aspects, the transparent conductive film or layer is 10-30 nm thick, 30-50 nm thick, 50-70 nm thick, 70-90 nm thick, 90-110 nm thick, 110-130 nm thick, 130-150 nm thick, 150-170 nm thick, 170-190 nm thick, 190-210 nm thick, 210-230 nm thick, or 230-250 nm thick. In some aspects, the transparent conductive film or layer is 20nm thick (+ / -5nm), 30nm thick (+ / -5nm), 40nm thick (+ / -5nm), 50nm thick (+ / -5nm), 60nm thick (+ / -5nm), 70nm thick (+ / -5nm), 80nm thick (+ / -5nm), 90nm thick (+ / -5nm), 100nm thick (+ / -5nm), 110nm thick (+ / -5nm), 120nm thick (+ / -5nm), 130nm thick (+ / -5nm), 140nm thick (+ / -5nm), 150nm thick (+ / -5nm), 160nm thick (+ / -5nm), or 170nm thick (+ / -5nm). In some embodiments, the conductive film or layer may be heated or not heated.

[0087] In another aspect, the LC medium 106 can be located between the first conductive layer 1103(a) and the second conductive layer 1103(b), such as Fig.11As shown. The LC medium 106 may be sealed in the LC shutter 100 by the first conductive layer 1103 (a), the second conductive layer 1103 (b), the first spacer 1104 (a) and the second spacer 1104 (b). The LC shutter 100 may include a first electrical connector 1106 (a) and a second electrical connector 1106 (b). The first electrical connector 1106 (a) may be connected to the first conductive layer 1103 (a). The second electrical connector 1106 (b) may be connected to the second conductive layer 1103 (b). The first electrical connector 1106 (a) may be configured to supply a voltage from a driving voltage source to the first conductive layer 1103 (a). The second electrical connector 1106 (b) may be configured to supply a voltage from a driving voltage source to the second conductive layer 1103 (b). When the voltage is supplied to the first conductive layer 1103 (a) and the second conductive layer 1103 (b), an electric field may be applied to the LC medium 106. The voltage supplied by the driving voltage source to the first conductive layer 1103 ( a ) and the second conductive layer 1103 ( b ) may be a bipolar square wave.

[0088] When no electric field (e.g., no voltage) is applied to the LC medium 106, the LC medium 106 may rotate the polarization of light passing therethrough. When the first polarizing film 1101(a) and the second polarizing film 1101(b) are cross-polarized (e.g., have opposite orientations), this state of the LC shutter 100 is referred to as an open state. By rotating the polarization of light, the light may pass through the second polarizing film 1101(b) because the light will have the same polarization as the second polarizing film 1101(b) (e.g., the light is polarized by the first polarizing film 1101(a), and then the LC medium 106 rotates the light to the same polarization as the second polarizing film 1101(b)). In this example, the two polarizing films may appear to be aligned with each other. When no electric field is supplied to the LC medium 106, the LC shutter 100 may appear transparent (e.g., no light attenuation). When an electric field is applied to the LC medium 106, the LC medium 106 does not rotate the light passing therethrough. When the LC medium 106 does not rotate the light, when the light reaches the second polarizing film 1101 (b), the light has the polarization of the first polarizing film 1011 (a), and is therefore blocked by the cross-oriented second polarizing film 1101 (b). This state of the LC shutter 100 is the closed state. The supplied voltage can switch the LC shutter between the open state and the closed state.

[0089] In another aspect, the first polarizing film 1101(a) and the second polarizing film 1101(b) may have the same polarization (e.g., the same orientation). In this example, when no electric field is supplied (e.g., no voltage), the LC medium 106 may rotate the polarization of light passing therethrough. In this example, by rotating the polarization of the light, the light may be blocked by the second polarizing film 1101(b) because the rotated polarization of the light has a polarization opposite to that of the second polarizing film 1101(b). In this example, when an electric field is supplied to the LC medium 106, the LC medium 106 does not rotate the light. In this example, when the LC medium 106 does not rotate the light, the light has the polarization of the first polarizing film 1101(a), so the polarization of the second polarizing film 1101(b) allows the light to pass through the second polarizing film 1101(b). In this example, the effect of the voltage supply (e.g., no voltage provides a closed state and the supply of voltage provides an open state) can be reversed by changing the orientation of the polarizing films 1101(a), 1101(b).

[0090] In various aspects, camera 102 incorporates one or more color filters that are highly transmissive in the NIR spectrum. Figure 2 As shown in FIG. 2 , which illustrates the spectral response 200 of an exemplary camera, the camera 102 may include blue, green, and red filters (generally indicated by 202, 204, and 206, respectively) on the sensor pixels to isolate incident light in the aforementioned wavelengths. According to various aspects, the camera 102 is a multispectral camera. In other aspects, the camera is configured for hyperspectral imaging. In various aspects, the polarizer 104 and the LC cell 106 effectively act as a long pass filter, where shorter wavelengths of light in the visible spectrum can be blocked or allowed to pass based on an applied electric field, thereby controlling the light received at the camera 102. In various aspects, the unattenuated visible light and the emitted light can be directed to the LC shutter 100.

[0091] like Figure 3As shown, a multispectral imaging system typically includes a NIR long pass filter for separating visible light and NIR light. The long pass filter allows only NIR light from the light source to pass through and reach the imaging sensor, such as camera 102. In various aspects of multispectral imaging, the use of a NIR long pass filter is desirable because all three color pixels 202-206 depicted behave more like a monochrome camera that carries the full resolution of the sensor. In order to time-multiplex the visible light and NIR images, the long pass filter is typically moved in and out of the image path. In practice, a rotating filter wheel or a rotating solenoid can be used to achieve moving the physical filter in and out of the image path, but these methods are difficult to accomplish accurately at video speeds and are prone to synchronization errors, audible and subaudible noise, thermal noise, and vibration (i.e., any of the above is generally referred to as noise, or specifically as the context requires) and mechanical failure. Therefore, the liquid crystal shutter assembly of the present disclosure can act as a long pass filter that is used to address these limitations in order to obtain images at video speeds and reduce or eliminate the aforementioned problems, including reducing or eliminating synchronization errors, noise, and mechanical failures. According to various aspects of the present disclosure, the reduced extinction ratio at long wavelengths actually converts the liquid crystal assembly into a long pass filter with variable attenuation in short wavelength light within the effective range of the polarizer. In many cases, linear polarization filters have undesirable disadvantages. The first disadvantage is that at least 50% of the unpolarized light is lost due to absorption or reflection, so that only polarized light remains. In the case of a liquid crystal assembly using such a polarizer, the light is lost due to the first polarizer. In addition, in low light applications, such as those found during NIR fluorescence imaging, signal attenuation of about 50% or more may reduce the signal-to-noise ratio (SNR) to the point that an increased exposure time is required to capture useful data. Therefore, the need for longer exposure times is generally not conducive to capturing data at video rates.

[0092] The second disadvantage of using these types of polarizers is that they are not effective for all wavelengths of incident light. It is easy to understand that the polarizer becomes effectively transparent outside its polarization range. In other words, the polarizer may not be able to effectively polarize light outside its polarization wavelength range. Therefore, at a certain wavelength, the polarizer cannot polarize the incident light.

[0093] Contrary to popular belief, according to various aspects, the systems and methods of the present disclosure take advantage of these previously undesirable features or disadvantages. The LC shutter assembly 100 is capable of operating in three states. These states include "state 1", also referred to as the "open state", in which non-polarized light within the functional range of the linear polarizers is partially attenuated by the first linear polarizer based on the polarization rotation of the liquid crystal, but is generally allowed to pass through the second polarizer. The open state can be used to image visible light (e.g., reflected visible light) from the sample. "State 2" is also referred to as the "closed state", in which non-polarized light is partially attenuated by the first linear polarizer and then almost completely blocked or attenuated by the second linear polarizer due to the alternating rotation of the liquid crystal phase state. The ability to attenuate light in this closed state depends on the effectiveness of the linear polarizer to fully polarize the light. In various aspects, when comparing the open state to the closed state, the characteristics of the polarization filter allow the LC shutter to achieve an extinction ratio of two to four orders of magnitude within the spectral range of the polarization filter. In an exemplary embodiment of visible light and NIR spectral imaging, the polarization filter in the systems and methods of the present disclosure is selected to be effective in the visible spectrum, so the open state and the closed state only affect visible light. Alternatively, a polarizing beam splitter may be used instead of the polarizing filter.

[0094] Finally, "State 3" refers to a state in which light passing through an LC shutter assembly (such as, but not limited to, LC assembly 100) has spectral characteristics outside the effective operating range of the polarizer and therefore passes through the shutter assembly with very little attenuation. In the State 3 spectral range, unpolarized light is not polarized or attenuated, so the LC phase state does not matter. In this spectral range, transmittances in excess of 90% can be achieved using appropriate anti-reflection coatings on the polarization filter surface. State 3, which was traditionally considered a failure state due to low extinction ratio, becomes a highly transmissive state regardless of the liquid crystal phase state.

[0095] According to one aspect, the LC shutter assembly 100 uses incident light polarized by the polarizer 104 (state 1) and incident light that is not polarized by the filter and reaches the LC cell 106 with little attenuation to be filtered through the LC shutter 100 in an open state or a closed state (states 1 and 2). Figure 4 As shown, the extinction ratio (indicated as 400) of light entering the LC shutter 100 between the open and closed states of the shutter decreases as the polarizer fails to attenuate light in the NIR spectrum. Fig.12As shown, when the LC shutter 100 is in an open state 404 (state 1), light in the visible spectrum (e.g., having a wavelength between 400 nm and 700 nm) may be transmitted through the LC shutter 100. When the LC shutter 100 is in a closed state 406 (state 2), light in the visible spectrum (e.g., having a wavelength between 400 nm and 700 nm) may not be transmitted through the LC shutter 100. The same general approach is also applicable to other optical systems including light sources with similar spectral characteristics. For systems with different spectral contributions to their light sources, other polarizers may be selected to achieve appropriate light blocking / attenuation.

[0096] As shown, the attenuation caused by polarizer 104 fails at longer wavelengths. Due to the failed 50% attenuation at these longer wavelengths, LC cell 106 no longer receives approximately 50% of the incident (unpolarized) light. In some examples, the failed attenuation is the result of a light cutoff range that is specifically designed to observe desired light (e.g., fluorescence). Therefore, the contrast ratio (generally represented as 402) from the open state 404 to the closed state 406 is also reduced.

[0097] According to various aspects of the present disclosure, when the visible light image is superimposed on the NIR fluorescence image, the contrast reduction becomes a beneficial feature during State 3 operation. The resulting superimposed composite image and / or video provides medical personnel with greater clarity, detail, and information regarding the presence or absence of disease states (e.g., including the presence of tumors, cancers, vascular malformations, inflammation, or other diseases), changes in tissue and subtissue structures, anatomical structures of interest, and surrounding tissue or samples of the above (whether examined in vivo or ex vivo).

[0098] In yet another aspect, the amount of visible light can be adjusted by operating the LC phase between state 1 (on state) and state 2 (off state). These intermediate states are actually controlled by adjusting the applied voltage and therefore the electric field of the LC cell and allow variable attenuation of the light without affecting the light in the failed state (state 3).

[0099] According to various aspects, the LC shutter assembly of the present disclosure (e.g., LC shutter assembly 100) can be used in various states, and can additionally be operated between various open and closed states to allow selection of desired wavelengths in a failed state, depending on the application of the system or method of the present disclosure. In another aspect, based on the extinction ratio of the filter, an "intermediate" operating state has at least 50% attenuation and up to 100% maximum attenuation. Operating between the open state and the closed state allows the ratio of visible light to NIR light to be adjusted or customized. The intensity of the NIR signal is typically much lower than the visible light present in the system, so the image sensor typically requires a different gain setting or exposure time. By adjusting the amount of visible light attenuation between the open state and the closed state, the visible light level can be scaled to match the weak NIR fluorescence signal, thus allowing the same camera settings to be used for both spectral ranges.

[0100] In another aspect, the video rate sampling duration (i.e., exposure time) may be different for imaging a visible light image and emission light from a fluorophore (e.g., a fluorescence or fluorescent image). The liquid crystal shutter assembly may be placed in an open state to image a visible light image. The liquid crystal shutter assembly may remain in an open state for about 2 ms to about 6 ms during the video rate sampling duration to image a visible light image. The liquid crystal shutter assembly may be placed in a closed state to image emission light from a fluorophore. The liquid crystal shutter assembly may remain in a closed state for about 8 ms to about 16 ms during the video rate sampling duration to image emission light from a fluorophore. In other examples, the video rate sampling duration of a visible light image may be about 1 ms, about 2 ms, about 3 ms, about 4 ms, about 5 ms, about 6 ms, about 7 ms, about 8 ms, about 9 ms, about 10 ms, about 11 ms, about 12 ms, about 13 ms, about 14 ms, about 15 ms, about 16 ms, or longer. In another example, the video rate sampling duration for imaging emission light from a fluorophore may be about 5 ms, about 6 ms, about 7 ms, about 8 ms, about 9 ms, about 10 ms, about 11 ms, about 12 ms, about 13 ms, about 14 ms, about 15 ms, about 16 ms, about 17 ms, about 18 ms, about 19 ms, about 20 ms, about 25 ms, about 30 ms, or longer.

[0101] Engineered Polarizers

[0102] To take advantage of the deactivated polarizer state (i.e., when the incident wavelength falls outside the polarization range of the polarizer), the polarizer 104 can be specially manufactured or designed to cut off (e.g., optical cutoff) at a specific desired wavelength. For example, when using indocyanine green (ICG) dye for fluorescence imaging, the spectral emission is in the range of 750nm-950nm, or in the range of 800nm-880nm, and the peak is around 830nm. For example, in some embodiments, the polarizer 104 of the liquid crystal shutter assembly 100 is selected to lose effectiveness between 650nm and 800nm, or between 600nm and about 850nm, or between about 700nm to about 850nm, between about 800nm ​​to about 950nm, between about 800nm ​​to about 880nm, between about 775nm to about 795nm, or at about 785nm. In additional embodiments, the polarizer 104 (or both polarizers) is configured to lose effectiveness or have a light cutoff range of about 600 nm to about 620 nm, about 620 nm to about 640 nm, about 640 nm to about 660 nm, about 660 nm to about 680 nm, about 680 nm to about 700 nm, about 700 nm to about 720 nm, about 720 nm to about 740 nm, about 740 nm to about 760 nm, about 760 nm to about 780 nm, about 780 nm to about 800 nm, about 800 nm to about 820 nm, about 820 nm to about 840 nm, about 840 nm to about 860 nm, about 860 nm to about 880 nm, or about 880 nm to about 900 nm. In other embodiments, the polarizer 104 (or both polarizers) can be configured to lose effectiveness or have a light cutoff range below about 100 nm to about 150 nm, about 150 nm to about 200 nm, about 200 nm to about 250 nm, about 250 nm to about 300 nm, about 300 nm to about 350 nm, about 350 nm to about 400 nm, about 400 nm to about 450 nm, or about 450 nm to about 500 nm. In other embodiments, the liquid crystal shutter assembly is selected to lose effectiveness at 800 nm.

[0103] According to one aspect of the LC shutter assembly 100, approximately 50% of the fluorescent emission may be lost due to the polarizer 500, such as Figure 5. In contrast, according to various embodiments, a specially configured or designed polarizer 104A may be used. The engineered polarizer 104A is specifically selected and / or manufactured so that the cut-off wavelength of the polarizer can be adjusted or designed to a specific wavelength. For example, if ICG is a detectable fluorophore of interest, the polarizer can be adjusted to cut off at a desired wavelength in the range of 790nm to 900nm. In another example, another ICG fluorophore or any other fluorophore suitable for fluorescence imaging in the near infrared range (~780nm to 2500nm) can be used in conjunction with a polarizer 104 designed according to a specific desired cut-off wavelength. Figure 5 The different NIR transmission levels between the engineered polarizer 104A and the polymer polarizer 500 in combination with a closed-state LC shutter are shown.

[0104] By way of example and not limitation, polarizer 104 can be a dye polarizer, which is very efficient in the visible range but falls off rapidly in the NIR wavelength range. According to various aspects, dye polarizers are preferred because they have a high damage threshold suitable for absorbing a large amount of light in the visible spectrum.

[0105] In one aspect, the liquid crystal shutter assembly provides minimal attenuation of light outside the light cutoff range of the polarizer in state 3. In some instances, the light outside the light cutoff range may be near infrared light (NIR) or infrared light (IR). In other instances, the light outside the light cutoff range may be light with a wavelength above the visible spectrum, light with a wavelength within the visible spectrum, or light with a wavelength below the visible spectrum. Minimum attenuation allows optimal imaging of fluorescence from fluorophores by providing maximum intensity emission light to an imaging sensor and / or camera. In some instances, the emission light from the fluorophore may have an attenuation of 0% to 5% when passing through the liquid crystal shutter assembly 100. In other instances, the emission light from the fluorophore may have an attenuation of about 0% to about 5%, about 5% to about 10%, or about 10% to about 15% when passing through the liquid crystal shutter assembly 100.

[0106] Liquid crystal unit

[0107] According to various aspects, any of a variety of liquid crystal materials may be used in the LC shutter 100. For example, the LC shutter 100 may include a Pi cell or a twisted nematic ("TN") cell. Typically, a Pi LC cell has faster opening and closing times than a twisted nematic ("TN") cell. In addition, a Pi cell may cause a color shift in the image hue. Pi cells are commonly used in DLP projectors to provide video rate switching. However, DLP projectors require temperature compensated voltage control to avoid color fluctuations. Although effective, the use of Pi cells may not be suitable for non-cooled and simplified imaging systems. In one aspect of a twisted nematic liquid crystal, applying a non-zero voltage causes a backflow in the liquid crystal, causing the cell to be momentarily "opened". In yet another aspect, the backflow defines a partially open transmission state.

[0108] According to other aspects of the LC shutter assembly 100, TN cells are preferred. Although TN cell shutters are generally slower than equivalent Pi cells, TN cells have more desirable spectral characteristics because their absorption is relatively neutral in the operating spectrum. Therefore, the color natural characteristics of TN cells are more desirable for compact, uncooled imaging systems. The strong spectral characteristics of TN cells are a great benefit. The result of TN cells is a slower shutter open time. The time can improve with temperature.

[0109] According to various aspects, the closed time of a TN cell can be increased by increasing the applied voltage of the TN cell above its normal operating voltage. In one example, a TN cell (e.g., LC medium 106) that can be used in the LC shutter 100 can be configured to open when no electric field is applied. In this example, a TN cell shutter can achieve very fast shutter speeds and high contrast extinction ratios. Another characteristic of a TN cell that may generally be considered a disadvantage is that when driven with a high voltage, the TN cell will "backflow" during the opening period. Specifically, the high voltage can cause the TN cell to backflow, causing the LC shutter to open briefly, close completely, and finally open completely.

[0110] For example, Figure 6TN cells driven with two different voltages are shown, i.e., a lower voltage drive 600 and a higher voltage drive 602. The manufacturer recommends using the lower voltage drive 600 to avoid backflow. As shown, the 6 ms box 604 shows an exemplary video rate sampling duration (i.e., exposure time) for video rate imaging. Although the lower voltage drive 600 causes more open states within the 6 ms duration, it is necessary to stop the voltage drive earlier to close the cell due to the slow closing time. In contrast, the higher voltage drive 602 causes a faster initial opening before the backflow closing 606 occurs. Although the total transmittance of light using the higher voltage drive 602 within the 6 ms window is less than the total transmittance of the lower voltage drive 600, the TN cell performance can be further modified by heating the LC cell 106.

[0111] refer to Figure 7 , the opening time of a TN LC shutter (i.e., an LC shutter configured with a TN cell as a liquid crystal cell) can vary with temperature. Specifically, the opening times of the TN LC shutter at 20°C (700), 25°C (702), 35°C (704), 45°C (706), 55°C (708), 65°C (710), and 75°C (712) are shown. The opening time of the Pi cell (denoted as 714) is shown for comparison. As shown, heating the LC cell 106 reduces the viscosity of the liquid crystal material in the cell. As a result, the LC shutter response speed increases, thereby shortening the duration of the reflow phenomenon and ultimately increasing the speed at which the LC shutter 100 fully opens. In some instances, the transition speed between the open state and the closed state is determined by the temperature of the liquid crystal and the driving voltage source. In some embodiments, the LC cell 106 is heated, for example, between thirty and fifty degrees Celsius.

[0112] According to one aspect, the LC shutter may be heated by a dedicated heater that can be controlled to directly adjust the efficiency of the TN cell. According to another aspect, the LC shutter 100 may be disposed within a closed housing, whereby the elevated temperature is provided by waste heat generated by other components of the imaging system. Figure 6 As shown, the reduced backflow transmission coupled with the partially open state in response to the heated environment is high enough for real-time visible light imaging. By mounting the LC cell 106 or the entire LC shutter assembly 100 within a closed environment (such as, but not limited to, the housing of the imaging system disclosed in the aforementioned Blaze Bioscience system), the elevated internal temperature generated by the other components of the imaging system helps to mitigate the backflow associated with the TN cell. By way of example and not limitation, the LC cell 106 can be maintained at around 35°C to 40°C. In many aspects, an additional benefit of driving the TN cell at a higher voltage is an increase in optical blocking, such as Figure 8As shown. For example, a TN cell is typically driven with 0V in the on state and with some nominal low voltage in the off state. However, a higher voltage for a fast off time also helps improve the blocking capability of the cell. As shown, a higher peak-to-peak ("pp") drive voltage is advantageous for the application. The LC cell 106 can be driven by a DC drive voltage or an AC drive voltage.

[0113] According to one aspect, the LC cell 106 is driven with a high voltage in a range between + / -3Vp-p and + / -60Vp-p [peak to peak]. In an exemplary aspect, the LC shutter cell is driven by + / -24V (48Vp-p). In an exemplary aspect, the LC shutter cell is driven in a range between + / -16V (32Vp-p) and + / -30V (60Vp-p) (including any value). In an exemplary aspect, the LC shutter cell is driven in a range between + / -30V (60Vp-p) and + / -100V (200Vp-p) (including any value). According to various aspects, the LC shutter is driven by a square wave voltage with a period of 1-2kHz and a duty cycle of 50%. In this example, the square wave voltage ranges from a minimum value of 0V in the open state to a maximum value of + / -24V (48Vp-p) in the closed state. In another aspect, the square wave drive voltage is provided as a DC drive voltage ranging from -24V to 24V, so that the voltage drive is about 48V peak-to-peak. In another aspect, the amplitude of the bipolar square wave has a range of + / -3 to + / -30Vp-p. In an exemplary aspect, the amplitude of the bipolar square wave is in the range between + / -16V (32Vp-p) and + / -30V (60Vp-p), including the end value at any value. In an exemplary aspect, the amplitude of the bipolar square wave is in the range between + / -30V (60Vp-p) and + / -100V (200Vp-p), including the end value at any value. In another aspect, the second voltage includes a plurality of voltages that produce a bipolar square wave voltage with an average mean of 0V. In this aspect, for an off-state video rate image, the voltage is maintained at full 24V. In this implementation, there is no switching of the polarity of the voltage bias during frame exposure, so the LC shutter maintains a higher level of optical attenuation for the closed state. The voltage polarity for the next frame will flip, which requires the closed state to maintain the health and reliability of the LC shutter.

[0114] According to various aspects, the LC shutter may have a first voltage that switches the LC shutter to an open state and a second voltage that switches the LC shutter to a closed state. In some instances, the first voltage may be 0V. Applying the first voltage may allow the liquid crystal shutter assembly to assume an open state. In other instances, the first voltage may be a bipolar square wave with an amplitude in the range of + / -0.1Vp-p to + / -5Vp-p. When the liquid crystal is a twisted nematic liquid crystal cell, the first voltage may be 0V. When the liquid crystal is a Pi cell, the first voltage may be a bipolar square wave with an amplitude in the range of + / -0.1Vp-p to + / -5Vp-p. In other instances, the first voltage may be a bipolar square wave with an amplitude in the range of + / -3Vp-p to + / -60Vp-p. In other examples, the first voltage may be a bipolar square wave having an amplitude of about + / -12 Vp-p to + / -15 Vp-p, about + / -15 Vp-p to + / -18 Vp-p, about + / -18 Vp-p to + / -21 Vp-p, about + / -21 Vp-p to + / -24 Vp-p, about + / -24 Vp-p to + / -27 Vp-p, about + / -30 Vp-p The second voltage may be a bipolar square wave having an amplitude in the range of + / -3 Vp-p to + / -60 Vp-p. Applying the second voltage may allow the liquid crystal shutter assembly to assume a closed state. In other examples, the second voltage may be a bipolar square wave having an amplitude of about + / -12 Vp-p to + / -15 Vp-p, about + / -15 Vp-p to + / -18 Vp-p, about + / -18 Vp-p to + / -21 Vp-p, about + / -21 Vp-p to + / -24 Vp-p, about + / -24 Vp-p to + / -27 Vp-p, about + / -30 Vp-p In some embodiments, the second voltage may be a bipolar square wave having an amplitude in the range of + / -0.1 Vp-p to + / -5 Vp-p, about + / -0.1 Vp-p to + / -5 Vp-p, about + / -0.2 Vp-p to + / -6 Vp-p, about + / -0.5 Vp-p to + / -7 Vp-p, about + / -10 Vp-p to + / -11 Vp-p, about + / -11 Vp-p to + / -12 Vp-p, about + / -12 Vp-p to + / -13 Vp-p, about + / -13 Vp-p to + / -14 Vp-p, about + / -14 Vp-p to + / -15 Vp-p, about + / -15 Vp-p to + / -16 Vp-p, about + / -16 Vp-p to + / -17 Vp-p, about + / -18 Vp-p to + / -19 Vp-p, about + / -20 Vp-p to + / -21 Vp-p, about + / -22 Vp-p to + / -23 Vp-p, about + / -24 Vp-p to + / -25 Vp-p, about + / -26 Vp-p to + / -27 Vp-p, about + / -28 Vp-p to + / -29 Vp-p

[0115] In one aspect, the bipolar square wave can be modulated from 500 Hz to 2000 Hz. In some examples, the driving voltage source switches the liquid crystal between the first voltage (e.g., open state) and the second voltage (e.g., closed state) for a duration of 2 ms to 40 ms (inclusive), 1 ms to 40 ms (inclusive), or 2 ms to 60 ms (inclusive), 60 ms to 100 ms (inclusive), and up to 300 ms (inclusive). In other examples, the driving voltage source causes the liquid crystal to switch between a first voltage (e.g., an open state) and a second voltage (e.g., a closed state) with a duration of about 2 ms to about 4 ms, about 4 ms to about 6 ms, about 6 ms to about 8 ms, about 8 ms to about 10 ms, about 10 ms to about 12 ms, about 12 ms to about 14 ms, about 14 ms to about 16 ms, about 16 ms to about 18 ms, about 18 ms to about 20 ms, about 20 ms to about 22 ms, about 22 ms to about 24 ms, about 24 ms to about 26 ms, about 26 ms to about 28 ms, about 28 ms to about 30 ms, about 30 ms to about 32 ms, about 32 ms to about 34 ms, about 34 ms to about 36 ms, about 36 ms to about 38 ms, or about 38 ms to about 40 ms.

[0116] With respect to the first voltage and the second voltage described herein, it should be understood that the first voltage and the second voltage may be applied or present in any order or configuration in the liquid crystal assembly described herein (e.g., with an applied voltage of 0V or a + / - voltage regardless of whether the first voltage or the second voltage is in an off state or an off state, as described herein). For example, the polarizers within the LC assembly may also be configured (e.g., oriented) to achieve an applied voltage of 0V or a + / - voltage regardless of whether the first voltage or the second voltage is in an on state or an off state, as described herein. For example, when the polarizers are cross-polarized, a 0V voltage may be applied to present an on state (e.g., the LC cell rotates the polarization of light transmitted from the first polarizer so that the light passes through the second polarizer). When the polarizers are cross-polarized, a + / - voltage may be applied to present an off state (e.g., the LC cell does not rotate the polarization of light transmitted from the first polarizer so that the light is blocked or attenuated by the second polarizer). For example, when the polarizers have the same polarization (e.g., the same orientation), a + / - voltage may be applied to present an open state (e.g., the LC cell does not rotate the polarization of light transmitted from the first polarizer and the light passes through the second polarizer). When the polarizers have the same polarization (e.g., the same orientation), a 0V voltage may be applied to present a closed state (e.g., the LC cell rotates the polarization of light transmitted from the first polarizer and the light is blocked or attenuated by the second polarizer).

[0117] According to one aspect of the LC shutter assembly 100, the dye polarizer 104 is paired with a twisted nematic (TN) liquid crystal cell 106, which can be tuned to a desired separation between the visible and NIR spectral bands. When used as the LC cell 106 with the TN cell, the dye polarizer 104 achieves high attenuation in the visible light band, while having only about 15% attenuation in the ICG emission band when used for ICG fluorescence imaging. Thus, the engineered dye polarizer provides about 85% transmission of the ICG emission band regardless of whether the LC shutter 100 is in the closed or open state. This increased emission light transmittance is particularly useful when creating a composite image by superimposing visible light on available NIR fluorescence.

[0118] Adjusting fluorescent dyes

[0119] In many aspects, the cutoff wavelength of the polarizer can be adjusted or designed to a specific wavelength optimized for a specific fluorescent molecule that is being detected by the system using the liquid crystal shutter assembly of the present invention. In such embodiments, the polarizer can be configured to have a light cutoff that is adjusted to the emission range of the applicable fluorescent portion. In some instances, the light cutoff range of the polarizer may include the longest wavelength of the visible spectrum range. In some instances, the light cutoff range of the polarizer may be within the visible spectrum range. The polarizer may be ineffective and transparent at a wavelength above the longest wavelength of the visible spectrum range. In another instance, the polarizer may be ineffective and transparent at a wavelength below the shortest wavelength of the visible spectrum range. For example, the systems and methods of the present disclosure can be configured or optimized for fluorophores in the NIR range and / or fluorophores outside the NIR range. For example, using ICG dye, the polarizer can be configured to have a cutoff in a long wavelength spectral range suitable for excitation light, such as between about 600 nm and about 850 nm, or between about 700 nm and 800 nm, about 800 nm to about 950 nm, about 800 nm to about 880 nm, about 775 nm to about 795 nm, or about 785 nm. In addition, using ICG dye, the polarizer can be configured to have a wavelength cutoff within a certain range, can be configured to have a wavelength cutoff at 800nm, or have a wavelength cutoff when reaching the following wavelengths: 600nm, 625nm, 650nm, 675nm, 700nm, 725nm, 750nm, 775nm, 780nm, 785nm, 790nm, 795nm, 800nm, 805nm, 810nm, 815nm, 820nm, 825nm, 830nm, 835nm, 840nm, 850nm, 855nm, 860nm, 865nm, 870nm, 875nm, 880nm, 885nm, 890nm, 895nm or 900nm. A polarizer with an effective cutoff minimum enables emission bands (e.g., from excited fluorophores) to pass unpolarized wavelengths to be collected, for example, in the range of 800 nm to 880 nm, where the polarizer cutoff occurs at 800 nm. In other embodiments, fluorophores and / or fluorescent dyes that emit at wavelengths in the infrared range or above, or in the visible range, or below the visible range can be used with engineered polarizers and LC shutters driven by voltages suitable for capturing light at video switching rates based on the corresponding emission spectra.

[0120] For example, the fluorescent molecule may include an ultraviolet (UV) dye, a blue dye, or both. Exemplary UV and blue dyes for fluorophores include: ALEXA FLUOR 350 and AMCA dyes (e.g., AMCA-X dyes), derivatives of 7-aminocoumarin dyes, dialkylaminocoumarin reactive versions of ALEXA FLUOR 350 dye, ALEXA FLUOR 430 (and reactive UV dyes absorbing 400-450 nm with significant fluorescence beyond 500 nm in aqueous solution), Marina Blue and Pacific Blue dyes (based on the 6,8-difluoro-7-hydroxycoumarin fluorophore) that exhibit bright blue fluorescence emission around 460 nm, hydroxycoumarin and alkoxycoumarin derivatives, Zenon ALEXA FLUOR 350, Zenon ALEXA FLUOR430 and Zenon Pacific Blue, succinimidyl esters of Pacific Orange dyes, Cascade Blue acetyl azide and other pyrene derivatives, ALEXA FLUOR 405 and its derivatives, pyrene succinimidyl ester ...30 and its derivatives, pyrene succinimidyl esters, Cascade Blue acetyl azide and other pyrene derivatives, ALEXA FLUOR 405 and its derivatives, pyrene succinimidyl esters, Cascade Blue acetyl azide and other pyrene derivatives, Cascade Blue acetyl azide and other pyrene derivatives, Cascade Blue acetyl azide and other pyrene derivatives, Cascade Blue acetyl azide and other pyrene derivatives, Cascade Blue Yellow dye, PyMPO and pyridyl oxazole derivatives, aminonaphthyl dyes and dansyl chloride, dapoxyl dyes (e.g., Dapoxyl sulfonyl chloride, amine-reactive Dapoxyl succinimide esters, carboxylic acid-reactive Dapoxyl (2-aminoethyl) sulfonamide), bimane dyes (e.g., bimane thioglycolic acid) and derivatives thereof, NBD dyes and derivatives thereof, QsY 35 dyes and derivatives thereof, fluorescein and derivatives thereof. Fluorescent molecules may include infrared dyes, near-infrared dyes, or both.Exemplary infrared and near infrared dyes for fluorophores include: DyLight-680, DyLight-750, VivoTag-750, DyLight-800, IRDye-800, VivoTag-680, Cy5.5, or indocyanine green (ICG) and any derivatives thereof, cyanine dyes, acridine orange or yellow, ALEXA FLUOR and any derivatives thereof, 7-actinomycin D, 8-anilinonaphthalene-1-sulfonic acid, ATTO dye and any derivatives thereof, auramine-rhodamine stain and any derivatives thereof, benzanthrone, bimane, 9-10-bis(phenylethynyl)anthracene, 5,12-bis(phenylethynyl)anthracene, bisbenzimides, brain rainbow, calcein, carboxyfluorescein and any derivatives thereof, 1-chloro-9,10-bis(phenylethynyl)anthracene and any derivatives thereof, DAPI, DiOC6, DyLight Fluor and any derivative thereof, epicocconone, ethidium bromide, FlAsH-EDT2, Fluo dye and any derivative thereof, FluoProbe and any derivative thereof, fluorescein and any derivative thereof, pyrimidine and any derivative thereof, GelGreen and any derivative thereof, GelRed and any derivative thereof, fluorescent protein and any derivative thereof, m subtype protein and any derivative thereof (such as mCherry), hetamethine dye and any derivative thereof, hoeschst colorant, iminocoumarin, Indian yellow, indo-1 and any derivative thereof, la urdan, fluorescent yellow and any derivatives thereof, fluorescein and any derivatives thereof, luciferase and any derivatives thereof, merocyanine and any derivatives thereof, nile dye and any derivatives thereof, perylene, phloxine, phyco dye and any derivatives thereof, propidium iodide, pyranine, rhodamine and any derivatives thereof, ribogreen, RoGFP, rubrene, stilbene and any derivatives thereof, sulforhodamine and any derivatives thereof, SYBR and any derivatives thereof, synapto-pHluorin, tetraphenylbutadiene, tetrasodium tris, Texas Red, dardan yellow, TSQ, umbelliferone, anthrone violet, yellow fluorescent protein and YOYO-1.Other suitable fluorescent dyes include, but are not limited to, fluorescein and fluorescein dyes (e.g., fluorescein isothiocyanate or FITC, naphthyl fluorescein, 4′,5′-dichloro-2′,7′-dimethoxyfluorescein, 6-carboxyfluorescein or FAM, etc.), carbocyanines, merocyanines, styryl dyes, oxonol dyes, phycoerythrin, erythrosine, eosin, rhodamine dyes (e.g., tetramethylcarboxy-rhodamine or TAMRA, carboxyrhodamine 6G, carboxy-X-rhodamine (ROX), lissamine rhodamine B, rhodamine 6G, rhodamine green, rhodamine red, tetramethylrhodamine (TMR), etc.), coumarins and coumarin dyes (e.g., methoxycoumarins, dialkylaminocoumarins, hydroxycoumarins, aminomethylcoumarins (AMCA), etc.), Oregon green dyes (e.g., Oregon Green 488, Oregon Green 500, Oregon Green 514, etc.), Texas Red, Texas Red-X, Spectrum Red, Spectrum Green, cyanine dyes (e.g., CY-3, Cy-5, CY-3.5, CY-5.5, etc.), ALEXA FLUOR dyes (e.g., ALEXA FLUOR 350, ALEXA FLUOR 488, ALEXA FLUOR 532, ALEXA FLUOR 546, ALEXA FLUOR568, ALEXA FLUOR 594, ALEXA FLUOR 633, ALEXA FLUOR 660, ALEXA FLUOR 680, etc.), BODIPY dyes (e.g., BODIPY FL, BODIPY R6G, BODIPY TMR, BODIPY TR, BODIPY 530 / 550, BODIPY558 / 568, BODIPY 564 / 570, BODIPY 576 / 589, BODIPY 581 / 591, BODIPY 630 / 650, BODIPY 650 / 665, etc.), IR dyes (e.g., IRD 40, IRD Additional suitable detectable agents are known and described in International Patent Application No. PCT / US2014 / 056177, which is incorporated herein by reference in its entirety.

[0121] The systems and methods herein used to detect fluorescent molecules in a sample may include one or more dyes, two or more, three, four, five, and up to ten or more such dyes in a given sample using any type of dye (e.g., ultraviolet (UV) dyes, blue dyes, infrared dyes, or near infrared dyes) in any combination. In addition to fluorescence, the disclosed systems and methods are effective in photoluminescence applications. The band gap associated with aluminum nitride has an emission similar to that of visible spectrum fluorophores, while the band gap associated with aluminum gallium arsenide has an emission similar to that of NIR fluorophores. Photoluminescence imaging can be applied to, but is not limited to, defect detection in optoelectronic semiconductor processing and organic and inorganic LED displays. Using the systems and methods herein, autofluorescence in an organ, organ substructure, tissue, target, cell, or sample can be detected.

[0122] Using controlled excitation energy for liquid crystal shutters

[0123] According to various aspects, the integration of LC shutters (such as but not limited to LC shutter 100) in various combined visible and NIR light systems, combined with appropriate adjustment of spectral filters (such as but not limited to polarizer 104), enables the capture of NIR images without the significant impact of visible light. These improvements can be further enhanced when paired with techniques for reducing the effects of light contamination within the collected emission band.

[0124] Fluorescence and photoluminescence optical pumping rely on strong light sources to excite the relevant materials. Specifically, increasing the excitation power can lead to enhanced signals. This signal enhancement is effective until the saturation point of the fluorophore or the non-radiative recombination loss of the photoluminescent junction.

[0125] Typically, increasing the excitation energy produces a traditionally undesirable effect, namely, adding heat to the sample being tested. Undesirable effects can be mitigated by shortening the duration of the excitation so that the average wattage incident on the sample being tested is maintained below a predetermined level or maintained at a safety level required for the application. For example, in in vivo applications where the systems and methods disclosed herein are used to image organs, organ substructures, tissues, targets, cells, or samples, the excitation energy applied to the tissue must be below known safety standards and thresholds to minimize heating and / or damage. In various aspects, controlling the peak intensity and duration of the excitation energy provides controlled excitation energy. The controlled excitation energy can be further adjusted so that the signal-to-noise ratio (SNR) of the NIR image can be enhanced while retaining boundaries due to damage thresholds or safety issues.

[0126] refer to Fig. 9, patient safety criteria can be quantified as the area under a curve of energy applied to tissue plotted over time or duration. In physiological applications, there are safe energy levels that can enter tissue at which the tissue is not damaged. As shown, the area depicted under A clinical 900 shows an example of a "safe" energy exposure (e.g., to living tissue), as indicated by the area under the curve formed by the energy of the threshold level or intensity depicted at 902, which energy can be administered for a short period of time or a long period of time (e.g., an extended amount of time). The present system can use higher intensity energy for a short period of time to achieve a safe level of energy or a lower level of energy, which has the advantage of enhancing the photons available for imaging while still meeting or exceeding patient safety in clinical applications at which tissue is not damaged. As Fig. 9 As shown in the figure, the burst-controlled strong excitation energy identified as A flash 906 shows the strong excitation energy pulse that can be applied in a short time using the systems and methods disclosed herein. As shown in the figure, the area under the curve (A flash 906) is limited by the controlled higher intensity excitation energy indicated by 908, and the flash duration is indicated by 910. As shown in the figure, although the flash energy 908 intensity is much higher than the clinical energy intensity 902, because the flash duration 910 is much shorter than the clinical duration 904, the area under the curve (A flash 906) is equal to or less than the area under the curve A clinical 900, which meets or exceeds the safety standard that the tissue is not damaged. Fig. 9 As shown, the area under the curve of A flash 906 is approximately equal to or less than the area under the curve of A clinical 900, so that the total energy applied to the tissue by the controlled excitation energy is reduced below the applicable safety threshold. In certain embodiments used herein, the systems and methods of the present invention are configured (e.g., by adjusting the intensity of the energy during the duration of the excitation energy pulse) so that the area under the curve (AUC) of A flash 906 is equal to or less than the AUC of A clinical 900.

[0127] It should be understood that the methods described herein using controlled excitation energy can be applied to methods and systems using the LC shutter assembly of the present disclosure and / or used with the Blaze Bioscience system.

[0128] In various aspects of the foregoing method, the liquid crystal assembly includes an engineered polarizer disposed in the path of light from the sample, wherein the polarizer is configured to have an effective spectral range such that the long wavelength cut-off range is between 600 nm and 900 nm or about 800 to 880 nm. The method may further include directing the emitted light and the reflected visible light through the liquid crystal assembly, wherein the reflected visible light (having a wavelength within the effective spectral range of the engineered polarizer) may be blocked or allowed to pass through the liquid crystal assembly depending on the state of the liquid crystal. Emitted light (having a wavelength outside the effective spectral range of the engineered polarizer) incident on the liquid crystal will pass through with little attenuation, regardless of the state of the liquid crystal.

[0129] Furthermore, in specific applications of video rate imaging, the inherent ordering of individual frames time-division multiplexed with the LC shutter will pair well with controlled and / or pulsed energy excitation. In some instances, the controlled and / or pulsed energy excitation may be white light, NIR light, IR light, or any other type of excitation light. Controlled excitation energy has the additional advantage that, in addition to producing more intensive excitation with respect to ambient light contamination in the emission spectral band, the controlled excitation energy also increases the signal to background or ambient light ratio, thereby allowing shorter exposure times (i.e., video rate sampling durations). According to various aspects, the shorter exposure times allowed by the use of controlled excitation energy reduce the amount of thermal noise integrated into the image from the sensor, thereby having the indirect benefit of improving image SNR. Specifically, the SNR increases significantly as the laser pulse width decreases and the corresponding integration time decreases. Reference Fig.10 , the SNR of the pulsed or controlled excitation energy 1000 is greater than the SNR of the lower intensity energy pulse 1002. It is further shown that as the laser duration is shortened, the signal-to-noise ratio continues to increase.

[0130] One type of SNR used in image analysis is the contrast-to-noise ratio (CNR). The CNR of an image can be improved when the object you want to detect is bright and the surrounding area is dark. When the object you want to detect is fluorescent, the amount of light available is much lower than the ambient light. In addition, visible illumination added to assist in detail work (such as surgery) is usually much brighter than typical ambient light. Therefore, the fluorescence to be detected may be several orders of magnitude lower than the ambient background light, and the CNR of the fluorescent image is below the usable range. In order to improve the CNR, the fluorescence can be increased. Two known methods of increasing fluorescence are to increase the amount of fluorescent material or to increase the excitation energy. However, due to physiological barriers, increasing the amount of fluorescent material may not be feasible in biological materials. In addition, increasing the amount of excitation energy may not be feasible due to the possible thermal damage to the biological material.

[0131] Traditionally, CNR is improved by reducing background light (e.g., visible light) to make fluorescence more apparent. Ideally, the background light source is intermittently removed or turned off. When the background light source cannot be removed or turned off, the excitation source is modulated and dark frames are collected to subtract the background light from the signal. Using this approach results in the detection of fluorescence, but at the expense of a large portion of the camera's dynamic range being filled with bright background light, which must be removed by subtraction. The images have high noise levels and require long sequence times, resulting in reduced video frame rates.

[0132] Although the frame subtraction system is an improvement over previous systems, reducing the sequence time (e.g., the necessary video rate sampling duration) by using the LC shutter 100 can significantly increase the video frame rate, block interfering light (e.g., background light), improve and increase CNR, and enhance SNR in bright ambient light environments. The enhanced CNR and / or SNR can achieve fast or increased frame rates. The LC shutter 100 can be used to intermittently block background light, such as Fig.17 As shown. When the LC shutter is in an open state (e.g., a first voltage of 0V is applied), bright background light (e.g., visible light reflected from the sample) is allowed to pass through the LC shutter to image the visible light (e.g., a first video rate sampling duration). Imaging the visible light may be performed during the first video rate sampling duration (e.g., when the LC shutter is in an open state). In some examples, the first video rate sampling duration (i.e., the first exposure time) may be about 1 ms to about 2 ms, about 2 ms to about 3 ms, about 3 ms to about 4 ms, about 4 ms to about 5 ms, about 5 ms to about 6 ms, about 6 ms to about 7 ms, about 7 ms to about 8 ms, about 8 ms to about 9 ms, or about 9 ms to about 10 ms. In one example, the first video rate sampling duration may be about 2 ms to about 6 ms.

[0133] When the LC shutter is in a closed state (e.g., a second voltage is applied), bright background light (e.g., visible light) from the camera can be blocked, thereby allowing only NIR to pass during a second video rate sampling duration (e.g., a second exposure time) and allowing fluorescence from the sample to be imaged. When the LC shutter 100 is in a closed state, the laser diode can be turned on to provide excitation energy to the fluorophore, thereby allowing fluorescence (i.e., emitted light from the fluorophore) to be imaged. In some examples, the second video rate sampling duration (i.e., the second exposure time) can be about 5 ms to about 6 ms, about 6 ms to about 7 ms, about 7 ms to about 8 ms, about 8 ms to about 9 ms, about 9 ms to about 10 ms, about 10 ms to about 11 ms, about 11 ms to about 12 ms, about 12 ms to about 13 ms, about 13 ms to about 14 ms, about 14 ms to about 15 ms, about 15 ms to about 16 ms, or about 16 ms to about 17 ms. In some examples, the second video rate sample duration may be about 12 ms to about 16 ms or about 13 ms to about 17 ms.

[0134] After imaging the fluorescence, the laser diode may be turned off for a short period of time (e.g., a third video rate sampling duration) while the LC shutter remains in a closed state to allow imaging of a dark background image. When the laser diode is turned off, the laser diode stops providing excitation energy. In some examples, the third video rate sampling duration (i.e., the third exposure time) may be about 5 ms to about 6 ms, about 6 ms to about 7 ms, about 7 ms to about 8 ms, about 8 ms to about 9 ms, about 9 ms to about 10 ms, about 10 ms to about 11 ms, about 11 ms to about 12 ms, about 12 ms to about 13 ms, about 13 ms to about 14 ms, about 14 ms to about 15 ms, about 15 ms to about 16 ms, or about 16 ms to about 17 ms. In some examples, the third video rate sampling duration may be about 12 ms to about 16 ms or about 13 ms to about 17 ms.

[0135] In one aspect, a dark background image can be subtracted from a fluorescent image, thereby providing a clear fluorescent image. The LC shutter 100 in the closed state blocks any light outside the emission band and spectrally blocks interfering light (bright background light) from being captured. Removing bright background light (e.g., visible light) allows a higher analog gain to be used to fill the camera dynamic range during image capture. Increasing the camera gain can greatly shorten the integration time. In this embodiment, the LC shutter 100 acts as a transient spectral filter, allowing fluorescence to be observed in the closed state and visible light to be observed in the open state. The integration time required for the minimum CNR is shortened, enabling real-time video of the fluorescent image even in the presence of bright background light.

[0136] like Figure 8As shown, the blocking of visible light increases with increasing driving voltage of the LC medium 106. When visible light is blocked, the fluorescent image hidden under the visible light image is revealed. The CNR value of the fluorescent image increases with increasing driving voltage of the LC medium, thereby allowing better imaging of the fluorescent image, such as Fig.15 shown.

[0137] The increased CNR achieved by using the LC shutter 100 allows post-processing of the fluorescence image using digital gain, which is different from requiring longer exposure times. Post-processing via digital gain and shorter exposure times can increase the frame rate of the fluorescence image. Fig.16 As shown, the exposure time required to maintain a CNR of 5 decreases as the voltage increases. When a voltage of about 13 V or higher is supplied to the LC shutter 100, an exposure time of less than 5 milliseconds can be achieved.

[0138] like Fig.18A As shown in Figure 1-B, when the LC shutter assembly is in the closed state, the brightness of the dark image is reduced, reducing artifacts in the subtracted image, as described herein. In addition, the exposure time (i.e., video rate sampling duration) of the dark frame (laser off) is reduced to minimize motion blur in the image. Furthermore, due to the reduced brightness of the dark image, the use of the LC shutter assembly reduces the intensity of the dark frame and the ratio of dark frames to NIR frames. As shown in Figure 1-B, when the LC shutter assembly is in the closed state, the brightness of the dark image is reduced, reducing artifacts in the subtracted image, as described herein. In addition, the exposure time (i.e., video rate sampling duration) of the dark frame (laser off) is reduced to minimize motion blur in the image. In addition, due to the reduced brightness of the dark image, the use of the LC shutter assembly reduces the intensity of the dark frame and the ratio of dark frames to NIR frames. Fig.18A As shown in FIG. 1 , by using bright background subtraction, a fluorescence image taken on a linear stage (e.g., one that is rotating or starting to move) results in an image with unclear boundaries because the subtracted image shows errors in the translation of the dark image relative to the NIR image. Fig.18A In , long dark frame exposure times result in images that appear to capture motion and are not accurate depictions of the location of the fluorescence. Fig.18B As shown, by using the LC shutter 100 to block visible light, a fluorescence image captured on a linear stage (e.g., being rotated or starting to move) results in an image with sharp boundaries. By using the LC shutter 100 to block visible light and shortening the exposure time, a clear fluorescence image can be produced, thereby providing the precise location of the fluorescence. Clear fluorescence images are critical in surgical operations, where surgeons require the precise location of various parts of the body to perform safe, efficient, and effective operations.

[0139] Using the LCD shutter to reduce ghosting or unwanted light

[0140] like Fig.13 As shown, the imaging system may include a beam splitter 1302. The function of the beam splitter is to split the incident light into a reflected beam and a transmitted beam. An undesirable feature of some types of beam splitters is the generation of secondary reflections. The secondary reflections may produce a "ghost" image. Fig.13As shown, light from object plane 1301 is partially reflected off the front surface of beam splitter 1302 to form front surface reflection 1304. The rear surface of the beam splitter will produce rear surface reflection 1305 (e.g., a "ghost" image). The remaining light is transmitted light 1303 without a "ghost" image. The front surface reflection 1304 and the rear surface reflection 1305 of the beam splitter 1302 both have an S-dominant polarization relative to the orientation of the beam splitter surface. Conventionally, the rear surface coating will attempt to minimize the rear surface reflection 1305 using an anti-reflection (AR) coating, while the front surface coating will produce the front surface reflection 1304 (i.e., primary reflection) of the beam splitter 1302. Conventionally, the reflectivity of the front surface reflection 1304 must be at least 10 times that of the rear surface reflection 1305 to make the rear surface reflection 1305 inconspicuous or "invisible" relative to the front surface reflection 1304. In other words, it is considered a desirable result for the front reflectivity (e.g., the reflectivity of the front surface reflection 1304) to be 10 times higher than the back reflectivity (e.g., the reflectivity of the back surface reflection 1305) or to have a 10:1 ratio (front reflection / back reflection) to reduce ghosting in a system containing the beam splitter 1302. It is technically difficult to suppress S-polarized reflection and achieve a back surface reflectivity (e.g., the reflectivity of the back surface reflection 1305) below 1% over the entire visible spectrum, so the reflectivity of the front surface reflection 1304 is intentionally increased as the only mechanism to achieve a 10:1 reflectivity ratio. This limits the minimum total reflectivity of the beam splitter (by adding the front and back percentage reflections) to 11%. In a system that attempts to maintain a very low total reflectivity, this 11% lower limit is undesirable.

[0141] Compared to S polarization, it is easier to reduce and control the P polarization of the front surface reflection 1304 or the rear surface reflection 1305 to a value close to zero. Within the effective spectral range of the polarizer, the disclosed LC shutter provides the advantage that only linearly polarized light passes through the shutter. Therefore, the LC shutter can be oriented so that it will block S polarized light reflected from the beam splitter. By orienting the polarizer of the LC shutter to block S polarized light, only P polarized light is imaged. The front surface coating and the rear surface coating of the beam splitter can be configured to minimize S polarization reflections on the front and rear surfaces of the beam splitter. By adding only a small amount of P polarization reflection on the front surface coating, a reflectivity ratio of 10:1 or more can be easily achieved using the described LC shutter while only imaging P polarized light. The S polarized light is then simply designed to the lowest reasonable level. By appropriately designing the coating, the ratio of the front surface reflection 1304 to the rear surface reflection 1305 in P polarization can exceed 10:1 while keeping the total reflectivity of the beam splitter 1302 below the minimum value of 11%. The front-to-back ratio of S polarization does not reach a 10:1 ratio, but this light is then filtered out using appropriately positioned polarization filters of the LC shutter of the present disclosure. The first polarizing film 1101(a) of the LC shutter 100 can be oriented to block S polarized light from the back surface reflection 1305 and the front surface reflection 1304 and pass only P polarized light. The intensity ratio of the front surface reflection 1304 and the back surface reflection 1305 can increase as the P polarized light is passed to the imaging lens 1307 and then to the camera 1308. By removing the S polarized light and imaging only the P polarized (relative to the angle of the beam splitter) light, various front-to-back reflectance ratios can be achieved in applications where low emissivity values ​​are desired by designing the reflection of the P polarized light.

[0142] In some aspects, a front-to-back reflectivity ratio of at least 2:1, at least 4:1, at least 6:1, at least 8:1, at least 10:1, at least 11:1, at least 12:1, at least 14:1, at least 15:1, at least 16:1, at least 17:1, or at least 20:1 front-to-back reflectivity ratio inclusive (i.e., for any ratio between 2:1 and 20:1) can thus be achieved, any of which can be used to reduce ghosting. This technique of designing the beam splitter coating to utilize linearly polarized light is an advantage of the LC shutter, namely, that the total reflectivity of the beam splitter 1302 can be removed or reduced to well below the nominal minimum of 11% that can be achieved without the use of a polarizer. The system can be configured so that the total reflectivity of the beam splitter is 11% or less, 10% or less, 9% or less, 8% or less, 7% or less, 6% or less, 5% or less, 4% or less, or 3% or less. Fig. 20A An example of a front to back reflectivity ratio (FBR) of approximately 10:1 across the visible spectrum is illustrated. Fig. 20AAs shown, the P-polarized light FBR 2000 can maintain a FBR of 10:1 or greater across the entire visible spectrum. Fig. 20A As shown, the S-polarized FBR 2002 is below the desired FBR 10:1 in the visible spectrum, however, the S-polarized light is attenuated or blocked by the first polarizing film 1101(a). Fig. 20B As shown, using an LC shutter to filter S-polarized light out of the beam splitter can result in a total reflectivity of less than 11% over the visible spectrum.

[0143] like Fig. 14B As shown, removing the S polarized light from the rear surface reflection 1305 removes the "ghost" image 1400. Fig.14A As shown, when the LC medium 106 is oriented so that the first polarizer attenuates P polarized light, a ghost image 1400 may be present on the lower edge of the image. Fig. 14B As shown, when the LC medium 106 is oriented so that the first polarizer attenuates S-polarized light, the ghost image is removed. In some instances, the first polarizing film 1101 (a) may be designed so that S-polarized light is 100% attenuated or blocked. By blocking S-polarized light and designing the beam splitter to have a reflectivity ratio of 10:1 or greater for P-polarized light (front surface reflection to rear surface reflection), ghost images are absent or significantly reduced, resulting in a clearer image. In other instances, the first polarizing film 1101 (a) may be oriented to block other types of polarized light so that undesirable polarized light may be filtered before being transmitted through the liquid crystal shutter assembly. As Fig.14A As shown in Figure 1400, since the S polarization is attenuated, removing the ghost image results in reduced signal strength (e.g., signal strength drops from μ=212 to μ=131). The resulting image may lose some intensity; however, without the "ghost" image 1400, the image also defines clear boundaries, which is critical when detailed imaging is required.

[0144] In some embodiments, the polarizer 104 of the LC shutter 100 can be configured to improve image quality in any situation where the desired light can have a different polarization than the undesired light. In one example, a laser illuminator can be used in the imaging system. The laser illuminator can produce heavily polarized light. In some examples, it may be beneficial to block the laser light to allow other light (such as fluorescence) to be observed. Typically, a spectrally specific filter is used to block the light from the laser illuminator. Since the polarizer has at least partial efficacy at the laser wavelength, the use of polarization can help block the laser light. In some examples, a typical spectral filter can be configured to block polarized laser light and only allow fluorescence to be observed. A typical spectral filter can be configured to block polarized laser light outside the desired fluorescence range and allow fluorescence at a specific wavelength adjusted to the fluorescent molecule to be preferably observed. For example, such configurations can be used to visualize fluorescent molecules described herein, including fluorescent molecules or moieties within the following NIR spectra: between 650 nm and 850 nm, or between 600 nm and about 850 nm, or between about 700 nm and about 850 nm, between about 800 nm and about 950 nm, between about 800 nm and about 880 nm, between about 775 nm and about 795 nm, or about 785 nm, 700 nm, 725 nm, or about 785 nm. , 750nm, 775nm, 780nm, 785nm, 790nm, 795nm, 800nm, 805nm, 810nm, 815nm, 820nm, 825nm, 830nm, 835nm, 840nm, 845nm, 850nm, 855nm, 860nm, 865nm, 870nm, 875nm, 880nm, 885nm, 890nm, 895nm or 900nm. For example, the first polarizing film 1101 (a) can be configured to block polarized laser light having a wavelength of about 780nm to about 790nm while allowing the emitted light from the fluorescent molecules to pass through with minimal attenuation. It should be understood that various laser illuminators can be used in the system, including gas lasers, solid-state lasers, semiconductor lasers, chemical lasers, dye lasers, metal vapor lasers, or other types of lasers. It should be understood that the first polarizing film 1101(a) can be oriented so that different undesired polarized light from various laser illuminators can be blocked. In addition, any undesired polarized light can be blocked by adjusting the orientation of the first polarizing film 1101(a).

[0145] The first polarizing film 1101(a) and the second polarizing film 1101(b) can be configured to block heavily polarized laser light while allowing fluorescence to pass through the LC shutter 110, thereby improving the signal-to-noise ratio of the resulting fluorescence image. In one aspect, the first polarizing film 1101(a) and the second polarizing film 1101(b) can be configured to be effective in the wavelength of light from the laser illuminator. In one example, the first polarizing film 1101(a) can be configured to have an effective range of 420nm to 790nm, and thus can be used to help block laser light of 780nm to 790nm.

[0146] Fig.19 The use of a liquid crystal shutter assembly described herein configured to visualize a specific peptide-fluorophore conjugate (tocillin peptide) is shown. In this example, the liquid crystal shutter assembly is configured to use a first polarizing film 1101 (a) to block the heavily polarized light of the laser illuminator. The first polarizing film 1101 (a) can be effective in a portion of the visible spectrum 1900 and the NIR spectrum 1902. In order to block the heavily polarized laser light 1906 from the laser illuminator, the first polarizing film 1101 (a) is oriented to attenuate the heavily polarized laser light 1906 from the laser illuminator. The heavily polarized laser light 1906 can have a wavelength of about 780nm to about 790nm. The first polarizing film 1101 (a) can be configured to be effective up to about 800nm, and then lose effectiveness at wavelengths exceeding 800nm. Visualization of fluorescent molecules (e.g., peptide-fluorophore conjugates (tocillin)) with wavelengths of approximately 800 nm to 840 nm is optimized by blocking heavily polarized laser light 1906. The fluorescence can be observed regardless of the state of the liquid crystal shutter assembly because the fluorescence 1908 is outside the light cutoff range (e.g., effective range) of the polarizer.

[0147] like Fig.19 As shown, visible light (e.g., wavelengths of about 420 nm to 700 nm) can be completely attenuated (e.g., blocked) by the liquid crystal shutter assembly in the closed state 406. In the open state 404, the liquid crystal shutter assembly can be configured to allow visible light to pass through the liquid crystal shutter assembly with about 50% attenuation, allowing imaging of visible light. In the visible spectrum 1900, undesirable light of polarization (e.g., ghost images from S-polarized light exiting the beam splitter, as described herein) can be attenuated or blocked by orienting the first polarizing film 1101(a) of the first polarizer to block undesirable light of polarization. Blocking or attenuating undesirable polarization light during visible light imaging may result in a clearer visible image (e.g., reduced shadows or ghost images).

[0148] Detection Methods

[0149] In another aspect, disclosed herein is a method for imaging a sample, the method comprising: emitting infrared or near-infrared light by a light source to induce fluorescence from the sample; directing the infrared or near-infrared light to the sample by a plurality of optical devices; receiving fluorescence from the sample at a detector by the plurality of optical devices, wherein the infrared or near-infrared light is directed to the sample substantially coaxially with the fluorescence received from the sample so as to reduce shadows; and forming a fluorescence image of the sample and a visible light image of the sample on the detector. In some embodiments, the method herein comprises using an imaging system disclosed herein. In some embodiments, the sample is an organ, an organ substructure, a tissue, or a cell. In some embodiments, the method for imaging an organ, an organ substructure, a tissue, or a cell comprises imaging an organ, an organ substructure, a tissue, or a cell with an imaging system herein. In some embodiments, the method further comprises detecting cancer or a diseased area, tissue, structure, or cell. In some embodiments, the method further comprises performing surgery on a subject. In some embodiments, the method further comprises treating cancer. In some embodiments, the method further comprises removing cancer or a diseased area, tissue, structure, or cell from the subject. In some embodiments, the method further includes imaging the cancer or diseased area, tissue, structure or cell of the subject after surgical removal. In some embodiments, detection is performed using fluorescence imaging. In some embodiments, fluorescence imaging detects a detectable agent, which includes a dye, a fluorophore, a fluorescent biotin compound, a luminescent compound or a chemiluminescent compound. In some embodiments, the detectable agent includes any one or more fluorophores described herein. In some embodiments, the detectable agent includes ICG or tocilizumab.

[0150] In another aspect, disclosed herein is a method for treating or detecting a subject in need, the method comprising administering a companion diagnostic agent, a therapeutic agent or an imaging agent, wherein the companion diagnostic agent or imaging agent is detected by the system and method described herein. In another embodiment, the method of administering a companion diagnostic includes any of the various methods of using the system described herein. In another embodiment, the diagnostic agent or imaging agent includes a chemical agent, a radiolabeled agent, a radiosensitizer, a fluorophore, an imaging agent, a photosensitizer, an acoustic sensitizer, a diagnostic agent, a protein, a peptide, a nanoparticle or a small molecule. In another embodiment, the system is combined with radiology or fluorescence, including X-ray radiography, magnetic resonance imaging (MRI), ultrasound, endoscopy, elastic imaging, tactile imaging, thermal imaging, flow cytometry, medical photography, nuclear medicine functional imaging technology, positron emission tomography (PET), single photon emission computed tomography (SPECT), surgical instruments, surgical microscopes, confocal microscopes, fluoroscopes, exoscopes or surgical robots. In another embodiment, the system and method are used to detect the safety and physiological effects of therapeutic agents or evaluators. In another embodiment, the safety and physiological effects detected by the systems and methods are measurement of bioavailability, uptake, concentration, presence, distribution and clearance, metabolism, pharmacokinetics, localization, blood concentration, tissue concentration, ratio, concentration in blood and / or tissue, assessment of therapeutic window, range and optimization of the agent.

[0151] In various aspects, the imaging system disclosed herein can be used in a method for imaging abnormal tissue, cancer, tumor, vasculature, or structure in a fluorophore from a subject. In one aspect, the method includes generating an image of the vasculature or structure by using an imaging system provided herein to image the fluorescence. In another aspect, the method includes generating an image of abnormal tissue, cancer, tumor, vasculature, or structure by using an imaging system to image the fluorescence, and the imaging system includes a liquid crystal assembly system and method disclosed herein. In some examples, the fluorescence imaged may be autofluorescence, a contrast or imaging agent, a chemical agent, a radiolabeling agent, a radiosensitizer, a photosensitizer, a fluorophore, a therapeutic agent, an imaging agent, a diagnostic agent, a protein, a peptide, a nanoparticle, or a small molecule, or any combination thereof, or any combination thereof. In some examples, the method may further include administering a contrast or imaging agent to the subject. In some examples, the contrast or imaging agent may be a dye, a fluorophore, a fluorescent biotin compound, a luminescent compound, a chemiluminescent compound, or any combination thereof.

[0152] In various aspects, the contrast or imaging agent further comprises a protein, a peptide, an amino acid, a nucleotide, a polynucleotide or any combination thereof. In some instances, the contrast or imaging agent may be tocilizumab. In some instances, the contrast or imaging agent may absorb wavelengths between about 200 nm and about 900 nm.In some examples, the contrast or imaging agent may include: DyLight-680, DyLight-750, VivoTag-750, DyLight-800, IRDye-800, VivoTag-680, Cy5.5 or indocyanine green (ICG) and any derivatives of the foregoing; fluorescein and fluorescent dyes (e.g., fluorescein isothiocyanate or FITC, naphthyl fluorescein, 4′,5′-dichloro-2′,7′-dimethoxyfluorescein, 6-carboxyfluorescein or FAM, etc.), carbocyanine, merocyanine, styryl dye, oxonol dye, phycoerythrin, erythrosine, eosin, rhodamine dye (e.g. , tetramethylcarboxy-rhodamine or TAMRA, carboxyrhodamine 6G, carboxy-X-rhodamine (ROX), lissamine rhodamine B, rhodamine 6G, rhodamine green, rhodamine red, tetramethylrhodamine (TMR), etc.), coumarin, coumarin dyes (e.g., methoxycoumarin, dialkylaminocoumarin, hydroxycoumarin, aminomethylcoumarin (AMCA), etc.), Oregon green dyes (e.g., Oregon Green 488, Oregon Green 500, Oregon Green 514, etc.), Texas Red, Texas Red-X, Spectrum Red, Spectrum Green, cyanine dyes (e.g., CY-3, Cy-5, CY-3.5, CY-5.5, etc.), ALEXA FLUOR dyes (for example, ALEXA FLUOR 350, ALEXAFLUOR 488, ALEXA FLUOR 532, ALEXA FLUOR 546, ALEXA FLUOR 568, ALEXA FLUOR 594, ALEXA FLUOR 633, ALEXA FLUO R660, ALEXA FLUOR 680, etc.), BODIPY dyes (for example, BODIPY FL, BODIPY R6G, BODIPY TMR, BODIPY TR, BODIPY 530 / 550, BODIPY 558 / 568, BODIPY 564 / 570, BODIPY 576 / 589, BODIPY 581 / 591, BODIPY 630 / 650, BODIPY 650 / 665, etc.), IRD dyes (e.g., IRD40, IRD 700, IRD 800, etc.), 7-aminocoumarin, dialkylaminocoumarin reactive dyes, 6,8-difluoro-7-hydroxycoumarin fluorophores, hydroxycoumarin derivatives, alkoxycoumarin derivatives, succinimide esters, pyrene succinimide esters, pyridyl oxazole derivatives, aminonaphthyl dyes, dansyl chloride, dapoxyl dyes, Dapoxyl sulfonyl chloride, amine-reactive Dapoxyl succinimide esters, carboxylic acid-reactive Dapoxyl (2-aminoethyl) sulfonamide), bimane dyes, bimane thioglycolic acid, NBD dyes, QsY 35, or any combination thereof.

[0153] In various aspects, administering a contrast or imaging agent to a subject can include intravenous administration, intramuscular administration, subcutaneous administration, intraocular administration, intraarterial administration, peritoneal administration, intratumoral administration, intradermal administration, or any combination thereof.

[0154] In another aspect, the imaging may include tissue imaging, ex vivo imaging, intraoperative imaging, or any combination thereof. In some instances, the sample may be an in vivo sample, an in situ sample, an ex vivo sample, or an intraoperative sample. In another instance, the sample may be an organ, an organ substructure, a tissue, or a cell. In some instances, the sample is autofluorescent or exhibits autofluorescence. In one example, autofluorescence may include an ocular fluorophore, tryptophan, or a protein present in a tumor or malignancy. In some instances, the method may be used to visualize vascular flow or vascular patency.

[0155] In other aspects, the abnormal tissue, cancer, tumor, vasculature or structure may include blood vessels, lymphatic, vasculature, neuronal vasculature or CNS structures. In some instances, imaging may be angiography, arteriography, lymphangiography or cholangiography. In one example, imaging may include detecting vascular abnormalities, vascular malformations, vascular lesions, organs or organ substructures, cancer or diseased areas, tissues, structures or cells. In other instances, the vascular abnormalities, vascular malformations or vascular lesions may be aneurysms, arteriovenous malformations, venous malformations, lymphatic malformations, capillary malformations, mixed vascular malformations, spinal dural arteriovenous fistulas or combinations thereof.

[0156] In various aspects, the organ or organ substructure may be the brain, heart, lung, kidney, liver, breast, skin or pancreas. In some aspects, the method may include performing surgery on the subject. In one example, the surgery may include angioplasty, cardiovascular surgery, aneurysm repair, valve replacement, aneurysm surgery, arteriovenous malformation or cavernous vascular malformation surgery, venous malformation surgery, lymphatic malformation surgery, telangiectasia surgery, mixed vascular malformation surgery or dural arteriovenous fistula surgery, repair or bypass, arterial bypass, organ transplantation, plastic surgery, ophthalmic surgery, reproductive system surgery, stent insertion or replacement, plaque ablation, removal of the cancer or diseased area, tissue, structure or cell of the subject, or any combination thereof. In some instances, the imaging may include imaging of the subject's vascular abnormality, cancer or diseased area, tissue, structure or cell after surgery. In some aspects, the method may include treating cancer in the subject.

[0157] In various aspects, the method may include repair of intracranial CNS vascular defects, spinal CNS vascular defects; peripheral vascular defects; removal of abnormal vascularized tissue; eye imaging and repair; anastomosis; reconstruction or plastic surgery; ablation or treatment of atherosclerotic plaques or restenosis; repair or resection (including selective resection), preservation (including selective preservation) of important organs or structures such as nerves, kidneys, thyroid, parathyroid, liver segments or ureters; identification and management during surgery (sometimes preservation, sometimes selective resection); diagnosis and treatment of limb ischemia; or treatment of chronic wounds.

[0158] Adaptation of LC assembly to microscope

[0159] In some embodiments, the imaging system herein is stereoscopic. In some embodiments, the imaging system herein is not stereoscopic. In some embodiments, the imaging system herein is an operating microscope, a confocal microscope, a fluoroscope, an exoscope, an endoscope, or a surgical robot or an add-on (e.g., an imaging head) to any of the above.

[0160] In some aspects, at least one of the microscope, confocal microscope, fluoroscope, exoscope, surgical instrument, endoscope or surgical robot includes a KINEVO system (e.g., KINEVO 900), a QEVO system, a CONVIVO system, an OMPIPENTERO system (e.g., PENTERO 900, PENTERO 800), an INFRARED 800 system, a FLOW 800 system, a YELLOW 560 system, a BLUE 400 system, an OMPI LUMERIA system, an OMPI Vario system (e.g., OMPI Vario and OMPI VARIO 700), an OMPIPico system, an OPMI Sensera, an OPMI Movena, an OPMI 1FC, an EXTARO 300, a TREMON 3DHD system, a CIRRUS system (e.g., CIRRUS 6000 and CIRRUS HD-OCT), a CLARUS system (e.g., CLARUS 500 and CLARUS 700), a PRIMUS200, a PLEX Elite9000, AngioPlex, VISUCAM 524, VISUSCOUT 100, ARTEVO 800 (and any other surgical microscopes, confocal microscopes, fluoroscopes, exoscopes, endoscopes, ophthalmoscopes, retinal camera systems, optical coherence tomography (OCT) systems, and surgical robotic systems purchased from Carl Zeiss);PROVido system, ARvido system, GLOW800 system, Leica ARveo system, Leica M530 system (e.g., Leica M530 OHX, Leica M530 OH6), Leica M720 system (e.g., Leica M720 OHX5), Leica M525 system (e.g., Leica M525 F50, Leica M525 F40, Leica M525 F20, Leica M525 OH4), Leica M844 system, Leica HD C100 system, Leica FL system (e.g., Leica FL560, Leica FL400, Leica FL800), Leica DI C500, Leica ULT500, Leica rotatable beam splitter, Leica M651 MSD, LIGHTENING, Leica TCS, and SP8 system (e.g., Leica TCS SP8, SP8 FALCON, SP8 DIVE, LeicaTCS SP8 STED, Leica TCS SP8 DLS, LeicaTCS SP8 X, Leica TCS SP8 CARS, Leica TCS SPE), Leica HyD, Leica HCS A, Leica DCM8, Leica EnFocus, Leica Proveo 8, Leica Envisu C2300, Leica PROvido and any other surgical microscopes, confocal microscopes, fluoroscopes, exoscopes, endoscopes, ophthalmoscopes, retinal camera systems, OCT systems and surgical robotic systems; Haag-Streit 5-1000 system, Haag-Streit 3-1000 system, Haag-Streit HI-R NEO 900, Haag-Streit Allegra 900, Haag-Streit Allegra 90, Haag-Streit EIBOS2 and any other surgical microscopes, confocal microscopes, fluoroscopes, exoscopes, endoscopes, and surgical robotic systems; Intuitive Surgical da Vinci surgical robotic system, and any other surgical microscopes, confocal microscopes, fluoroscopes, exoscopes, endoscopes, ophthalmoscopes, retinal camera systems, OCT systems, and surgical robotic systems purchased from Intuitive Surgical;Heidelberg Engineering Spectralis OCT system, and any other surgical microscope, confocal microscope, fluoroscope, exoscope, endoscope, ophthalmoscope, retinal camera system, OCT system, and surgical robotic system purchased from Heidelberg Engineering; Topcon 3D OCT 2000, DRIOCT Triton, TRC systems (e.g., TRC 50DX, TRC-NW8, TRC-NW8F, TRC-NW8F Plus, TRC-NW400), IMAGEnet Stingray systems (e.g., Stingray, Stingray Pike, Stingray Nikon), IMAGEnet Pike systems (e.g., Pike, Pike Nikon), and any other surgical microscope, confocal microscope, fluoroscope, exoscope, endoscope, ophthalmoscope, retinal camera system, OCT system, and surgical robotic system purchased from Topcon; Canon CX-1, CR-2AF, CR-2PLUS purchased from Canon AF, and any other surgical microscope, confocal microscope, fluoroscope, exoscope, endoscope, ophthalmoscope, retinal camera system, OCT system, and surgical robotic system; Welch Allyn 3.5V system (e.g., 3.5V, 3.5V Autostep), CenterVue DRS, Insight, PanOptic, RetinaVue system (e.g., RetinaVue 100, RetinaVue 700), Elite, Binocular Indirect, PocketScope, Prestige coaxial-plus, and any other surgical microscope, confocal microscope, fluoroscope, exoscope, endoscope, ophthalmoscope, retinal camera system, OCT system, and surgical robotic system available from Welch Allyn; Metronic INVOS system, and any other surgical microscope, confocal microscope, fluoroscope, exoscope, endoscope, ophthalmoscope, retinal camera system, OCT system, and surgical robotic system available from Medtronic; Karl Storz ENDOCAMELEON, IMAGE1 systems (e.g. IMAGE1 S, IMAGE1S 3D, with or without OPAL1 NIR imaging module), SILVER SCOPE series instruments (e.g. gastroscopes, duodenoscopes, colonoscopes) and any other surgical microscopes, confocal microscopes, fluoroscopes, exoscopes, endoscopes, ophthalmoscopes, retinal camera systems, OCT systems and surgical robotic systems; or any combination thereof. ;

[0161] Combining or integrating the systems of the present invention into existing surgical microscopes, confocal microscopes, fluoroscopes, exoscopes, endoscopes, or surgical robots can be achieved by adapting an attachment containing an imaging system, a common housing (completely or partially), combining one or more aspects or components of the disclosed system into an existing system, or integrating one or more aspects or components of the disclosed system into an existing system. Such combinations can enhance image clarity and sharpen image boundaries; enhance video rates and real-time imaging; reduce shadows, motion effects, and / or ghosting; utilize improved, enhanced imaging, increased image clarity, optimized imaging, and improved surgical workflow, and other features of the systems and methods disclosed herein. In addition, such combinations or integrations can utilize an LC shutter assembly, an LC cell, a polarizer, a first voltage and a second voltage that switches the liquid crystal between an open state and a closed state, a light cutoff criterion, or any other feature of the systems disclosed herein, or any combination thereof.

[0162] The system of this article has been successfully adapted to one or more surgical microscopes, confocal microscopes, fluoroscopes, exoscopes, endoscopes or surgical robots by using optical gaskets to form a light-tight interface between two mating optical bases, mounting hardware and fixtures to allow light to pass from the applicable existing device to the system described herein. For example, the system of this article has been successfully adapted to at least one KINEVO system (e.g., KINEVO900), OMPIPENTERO system (e.g., PENTERO 900, PENTERO 800) using such an interface, and may be successfully adapted to other Zeiss systems or Leica FL800, etc. using such an interface.

[0163] Although certain embodiments, aspects, and examples are provided in the foregoing description, the inventive subject matter extends beyond the specifically disclosed embodiments or aspects to other alternative embodiments, aspects, and / or uses, and to modifications and equivalents thereof. Therefore, the appended claims are not limited to any specific embodiment or aspect described below. For example, in many of the methods or processes disclosed herein, the actions or operations of the methods or processes may be performed in any suitable order and are not necessarily limited to any particular disclosed order. Various operations may be described as multiple discrete operations in sequence in a manner that helps to understand certain embodiments; however, the order of description should not be understood to imply that these operations are order-dependent. In addition, the structures, systems, and / or devices described herein may be embodied as integrated components or separate components.

Claims

1. A liquid crystal assembly for use with an imaging system having a light source, the assembly comprising: a polarizer disposed in a path of light from the light source; The polarizer is configured to have a light cutoff outside its polarization range; a liquid crystal disposed in a path of light transmitted through the polarizer; as well as A driving voltage source is configured to apply a first voltage and a second voltage to the liquid crystal.

2. The liquid crystal assembly of claim 1, wherein the first voltage and the second voltage implement a high electric field and a low electric field on the liquid crystal.

3. The liquid crystal assembly as claimed in claim 1 or 2, wherein the first voltage and the second voltage cause the liquid crystal to switch between an open state and a closed state.

4. The liquid crystal assembly of any one of claims 1 to 3, wherein applying one of the first voltage or the second voltage switches the liquid crystal to the open state.

5. The liquid crystal assembly of any one of claims 1 to 4, wherein applying one of the first voltage or the second voltage switches the liquid crystal to the closed state.

6. The liquid crystal assembly of any one of claims 1-5, wherein the polarizer is configured to be transparent above the light cutoff.

7. The liquid crystal assembly of any one of claims 1-6, wherein the light cutoff includes the longest wavelength of a spectral range, and the polarizer is configured to be inactive and transparent above the longest wavelength.

8. A liquid crystal assembly as described in any one of claims 1 to 7, wherein the liquid crystal includes a twisted nematic (TN) unit, a Pi unit, a thermotropic liquid crystal and a lyotropic liquid crystal, a liquid crystal display (LCD), a liquid crystal with photoconductor properties, or a liquid crystal that is developed and adjusted under an electric field.

9. The liquid crystal assembly of any one of claims 1 to 8, wherein the second voltage induces a backflow in the liquid crystal.

10. The liquid crystal assembly according to any one of claims 1 to 9, wherein one of the first voltage or the second voltage is 0V.

11. The liquid crystal assembly of any one of claims 1 to 10, wherein the second voltage is a plurality of voltages including a bipolar square wave having an average value of 0V.

12. The liquid crystal assembly of any one of claims 1-11, wherein the first voltage, or the second voltage, or both the first voltage and the second voltage comprise a plurality of voltages that generate a bipolar square wave having an average value of 0V.

13. A liquid crystal assembly as described in any one of claims 11-12, wherein the bipolar square wave includes an amplitude of + / -0.1 to + / -5 Vp-p, + / -3 to + / -30 Vp-p, + / -16 Vp-p, + / -24 Vp-p or + / -30 Vp-p, or an amplitude between + / -16 V (32 Vp-p) and + / -30 V (60 Vp-p), including the end values ​​at any value, or an amplitude between + / -30 V (60 Vp-p) and + / -100 V (200 Vp-p), including the end values ​​at any value.

14. The liquid crystal assembly of any one of claims 1-13, wherein the second voltage is a bipolar square wave having an amplitude of + / -0.1 to + / -5 Vp-p, + / -3 to + / -30 Vp-p, + / -16 Vp-p, + / -24 Vp-p or + / -30 Vp-p, or an amplitude between + / -16 V (32 Vp-p) and + / -30 V (60 Vp-p), including any end value, or an amplitude between + / -30 V (60 Vp-p) and + / -100 V (200 Vp-p), including any end value.

15. The liquid crystal assembly of any one of claims 11 to 14, wherein the amplitude of the bipolar square wave is modulated at a video capture rate of the imaging system.

16. The liquid crystal assembly of any one of claims 13 to 15, wherein the amplitude of the bipolar square wave is modulated from 500 Hz to 2000 Hz.

17. A liquid crystal assembly as described in any one of claims 1-16, wherein the driving voltage source causes the liquid crystal to switch between the first voltage and the second voltage with a duration of 1 millisecond to 40 milliseconds inclusive, or 2 milliseconds to 60 milliseconds inclusive, 60 milliseconds to 100 milliseconds inclusive, and up to 300 milliseconds inclusive.

18. The liquid crystal assembly of any one of claims 1-17, wherein the drive voltage source causes the liquid crystal to switch between the first voltage and the second voltage with a video rate sampling duration of approximately between 1 ms and 10 ms, between 2 ms and 20 ms, or 6 ms.

19. The liquid crystal assembly according to any one of claims 1 to 18, wherein the liquid crystal is a twisted nematic (TN) liquid crystal.

20. The liquid crystal assembly of any one of claims 1 to 19, wherein the liquid crystal is heated to a temperature in the range of 30°C to 50°C.

21. The liquid crystal assembly of any one of claims 1-20, wherein the liquid crystal is enclosed in a housing of the imaging system, and wherein the liquid crystal is heated by waste heat generated by other components of the imaging system.

22. The liquid crystal assembly of any one of claims 4-21, wherein a transition speed between the open state and the closed state is determined by a temperature of the liquid crystal and a voltage source driving level.

23. The liquid crystal assembly of any one of claims 1-22, wherein the polarizer is an engineered dye polarizer.

24. The liquid crystal assembly of any one of claims 1-23, wherein the polarizer is configured to have a light cutoff in a range that is optimal for a fluorescence imaging process.

25. The liquid crystal assembly of any one of claims 1-24, wherein the polarizer is selected to lose effectiveness or cut off between 650 nm and 800 nm, or between 600 nm and about 800 nm, or between about 700 nm to about 800 nm, between about 800 nm to about 950 nm, between about 800 nm to about 880 nm, between about 775 nm to about 795 nm, or at about 785 nm.

26. A liquid crystal assembly as described in any of claims 1-25, wherein the polarizer is selected to lose effectiveness or cut off at 700nm, 725nm, 750nm, 775nm, 780nm, 785nm, 790nm, 795nm, 800nm, 805nm, 810nm, 815nm, 820nm, 825nm, 830nm, 835nm, 840nm, 850nm, 855nm, 860nm, 865nm, 870nm, 875nm, 880nm, 885nm, 890nm, 895nm or 900nm.

27. The liquid crystal assembly of any one of claims 1 to 26, further comprising a beam splitter.

28. The liquid crystal assembly of claim 27, wherein the beam splitter has a front surface and a rear surface.

29. The liquid crystal assembly of claim 28, wherein the front surface has a front surface coating, and the rear surface has a rear surface coating.

30. The liquid crystal assembly of claim 29, wherein the front surface coating has a higher P-polarization reflectance than the rear surface coating.

31. A liquid crystal assembly as described in claim 30, wherein the front surface coating and the rear surface coating produce a front surface to rear surface reflectivity ratio of P-polarized light of at least 2:1, at least 4:1, at least 6:1, at least 8:1, at least 10:1, at least 12:1, at least 14:1, at least 16:1, at least 18:1 or at least 20:

1.

32. The liquid crystal assembly of any one of claims 29-31, wherein S-polarized reflections of the front surface and the rear surface of the beam splitter are minimized by the front surface coating and the rear surface coating.

33. A liquid crystal assembly as described in any one of claims 1-32, wherein the polarizer is configured to block or attenuate ghost images or secondary reflections from the beam splitter.

34. The liquid crystal assembly of any one of claims 27-33, wherein the polarizer is configured to block S-polarized light from the beam splitter.

35. The liquid crystal assembly of claims 27-34, wherein the polarizer is configured to allow P-polarized light to pass through the liquid crystal assembly and reach an imaging lens and / or camera when the liquid crystal is in the open state.

36. A liquid crystal assembly as described in any of claims 27-35, wherein the first polarizer allows a front-to-back reflectivity ratio of the beam splitter to be at least 2:1, 4:1, 6:1, 8:1, 10:1, 12:1, 15:1, 17:1 or 20:1 to pass therethrough and reach the imaging lens and / or camera.

37. A liquid crystal assembly as described in any of claims 27-36, wherein the total reflectivity of the beam splitter is about 11% or less, about 10% or less, about 9% or less, about 8% or less, about 7% or less, about 6% or less, about 5% or less, about 4% or less, or about 3% or less.

38. The liquid crystal assembly of any one of claims 1-37, further comprising two or more polarizers.

39. The liquid crystal assembly of any one of claims 1-38, wherein the liquid crystal assembly is suitable for a medical device including a surgical microscope, a confocal microscope, a fluoroscope, an exoscope, an endoscope or a surgical robot.

40. The liquid crystal assembly of claim 39, wherein the liquid crystal assembly is adapted for the medical device, optionally through the use of optical spacers.

41. The liquid crystal assembly of claims 39-40, wherein the medical device is a KINEVO system (e.g., KINEVO900), an OMPIPENTERO system (e.g., PENTERO 900, PENTERO 800), or a Leica FL800 system.

42. A method for imaging emission from a fluorophore using a liquid crystal assembly as claimed in any one of claims 1 to 41.

43. A method for imaging emission light from a fluorophore at an imaging system including a liquid crystal assembly; the method comprising: Allowing or directing visible light to reach the sample; directing excitation light toward the sample; directing the emitted light and the reflected visible light from the sample to the liquid crystal assembly; The liquid crystal assembly comprises: an engineered polarizer disposed in the path of light from the sample; the polarizer configured to have a light cutoff in a range between 600 nm and 900 nm; a liquid crystal disposed in a path of light transmitted through the polarizer; and A driving voltage source configured to apply a first voltage and a second voltage to the liquid crystal; wherein the first voltage and the second voltage cause the liquid crystal to switch between an open state and a closed state; directing the emitted light and the reflected visible light through the engineered polarizer, wherein the reflected visible light having a wavelength below the light cutoff passes through the polarizer with approximately 50% attenuation, and the emitted light having a wavelength above the light cutoff passes through the polarizer with minimal attenuation; directing polarized visible light and non-polarized emitted light to the liquid crystal; applying the first voltage to the liquid crystal to exhibit the open state for a first video rate sampling duration; detecting the polarized light at an imaging sensor; applying the second voltage to the liquid crystal to assume the closed state and block the polarized light from passing to the imaging sensor; and The unpolarized emitted light is detected at the imaging sensor during the off state within a second video rate sampling duration.

44. The method of claim 43, wherein the minimum attenuation of the emitted light having a wavelength above the optical cutoff is 0% attenuation to 15% attenuation.

45. The method of claim 43 or 44, wherein the reflected visible light having a wavelength below the optical cutoff passes through the polarizer with an attenuation of about 50% or less.

46. ​​The method of any of claims 43-45, wherein the first video rate sample duration is between 1 ms and 10 ms, between 2 ms and 20 ms, or is 6 ms.

47. The method of any of claims 43-46, wherein the second video rate sampling duration is between 1 ms and 10 ms, between 2 ms and 20 ms, or is 6 ms.

48. The method of any one of claims 43-47, wherein the first voltage and the second voltage respectively achieve a high electric field and a low electric field on the liquid crystal.

49. The method of any one of claims 43-48, wherein the first voltage and the second voltage cause the liquid crystal to switch between an open state and a closed state.

50. The method of any one of claims 43-49, wherein the first voltage causes the liquid crystal to switch to the open state and the second voltage causes the liquid crystal to switch to the closed state, or vice versa.

51. The method of any one of claims 43-50, wherein one of the first voltage or the second voltage is 0V.

52. The method of any one of claims 43-51, wherein the second voltage is a plurality of voltages comprising a bipolar square wave having an average value of 0V.

53. The method of any one of claims 43-52, wherein the first voltage, or the second voltage, or both the first voltage and the second voltage comprise a plurality of voltages that produce a bipolar square wave having an average value of 0V.

54. A method as described in any of claims 52-53, wherein the bipolar square wave has an amplitude of + / -0.1 to + / -5 Vp-p, + / -3 to + / -30 Vp-p, + / -16 Vp-p, + / -24 Vp-p or + / -30 Vp-p, or an amplitude between + / -16 V (32 Vp-p) and + / -30 V (60 Vp-p), including any values, or an amplitude between + / -30 V (60 Vp-p) and + / -100 V (200 Vp-p), including any values.

55. The method of any one of claims 52-54, wherein the second voltage is a bipolar square wave having an amplitude of + / -0.1 to + / -5 Vp-p, + / -3 to + / -30 Vp-p, + / -16 Vp-p, + / -24 Vp-p or + / -30 Vp-p, or an amplitude between + / -16 V (32 Vp-p) and + / -30 V (60 Vp-p), including any values, or an amplitude between + / -30 V (60 Vp-p) and + / -100 V (200 Vp-p), including any values.

56. The method of any one of claims 52-55, wherein the bipolar square wave amplitude is modulated at a video capture rate of the imaging system.

57. The method of any one of claims 43-56, wherein the first voltage is a bipolar square wave having an amplitude of + / -0.1 to + / -5 Vp-p, + / -3 to + / -30 Vp-p, + / -16 Vp-p, + / -24 Vp-p or + / -30 Vp-p, or an amplitude between + / -16 V (32 Vp-p) and + / -30 V (60 Vp-p), including any values, or an amplitude between + / -30 V (60 Vp-p) and + / -100 V (200 Vp-p), including any values.

58. The method of any one of claims 54-57, wherein the amplitude is modulated from 500 Hz to 2000 Hz.

59. The method of any one of claims 43-58, further comprising: The liquid crystal is heated using waste heat from the imaging system.

60. The method of any one of claims 43-59, wherein the polarizer is a dye polarizer.

61. The method of any one of claims 43-60, wherein the polarizer is configured to have a light cutoff in a range optimal for a fluorescence imaging process.

62. The method of any one of claims 43-61, wherein the polarizer in the liquid crystal assembly is configured to have a light cutoff between about 600 nm and about 850 nm, or in a range of about 700 nm to about 850 nm, about 800 nm to about 950 nm, about 800 nm to about 880 nm, about 775 nm to about 795 nm, or at about 785 nm.

63. A method as described in any of claims 43-62, wherein the polarizer in the liquid crystal assembly is selected to lose effectiveness or cut off at 700nm, 725nm, 750nm, 775nm, 780nm, 785nm, 790nm, 795nm, 800nm, 805nm, 810nm, 815nm, 820nm, 825nm, 830nm, 835nm, 840nm, 850nm, 855nm, 860nm, 865nm, 870nm, 875nm, 880nm, 885nm, 890nm, 895nm or 900nm.

64. The method of any one of claims 43-63, wherein the polarizer in the liquid crystal assembly further comprises two polarizers or more polarizers.

65. The method of any one of claims 43-64, wherein the excitation light is designed to enhance the signal-to-noise ratio (SNR) of the NIR image such that the peak intensity and duration of the excitation light produce a controlled energy excitation that falls below a safety threshold.

66. The method of any one of claims 43-65, further comprising directing the emitted light and the reflected visible light to a beam splitter before directing the emitted light and the reflected light to the liquid crystal assembly.

67. The method of claim 66, wherein the beam splitter has a front surface and a rear surface.

68. The method of claim 67, wherein the front surface has a front surface coating and the back surface has a back surface coating.

69. The method of claim 68, wherein the front surface coating has a higher P-polarization reflectance than the back surface coating.

70. The method of claim 69, wherein the front surface coating and the rear surface coating produce a front surface to rear surface reflectivity ratio of P-polarized light of at least 2:1, at least 4:1, at least 6:1, at least 8:1, at least 10:1, at least 12:1, at least 14:1, at least 16:1, at least 18:1, or at least 20:

1.

71. The method of any one of claims 68-70, wherein S-polarized reflections of the front surface and the rear surface of the beam splitter are minimized by the front surface coating and the rear surface coating.

72. The method of any one of claims 43-71, wherein the polarizer is configured to block or attenuate ghost images or secondary reflections from a beam splitter.

73. The method of any one of claims 66-72, wherein the polarizer is configured to block S-polarized light from the beam splitter.

74. The method of any one of claims 66-73, wherein the polarizer is configured to allow P-polarized light to pass through the liquid crystal assembly and reach an imaging lens or camera when the liquid crystal is in the open state.

75. A method as described in any of claims 66-74, wherein the polarizer allows a front-to-back reflectivity ratio of the beam splitter to be at least 2:1, 4:1, 6:1, 8:1, 10:1, 12:1, 15:1, 17:1 or 20:1 to pass therethrough and reach the imaging lens and camera.

76. A method as described in any of claims 66-75, wherein the total reflectivity of the beam splitter is about 11% or less, about 10% or less, about 9% or less, about 8% or less, about 7% or less, about 6% or less, about 5% or less, about 4% or less or about 3% or less.

77. A liquid crystal assembly comprising any of the features described herein, alone or in combination with any other features, in any configuration.

78. A method for imaging emission light from a fluorophore at an imaging system, the imaging system comprising a liquid crystal assembly comprising any of the features described herein in any configuration, alone or in combination with any other features.

79. A liquid crystal assembly for use with an imaging system having a light source for providing excitation light, the assembly comprising: a first polarizer disposed in a path of light from the light source; The first polarizer is configured to have a light cutoff in a range outside of its polarization range; a liquid crystal disposed in a path of light transmitted through the first polarizer; a second polarizer disposed in a path of light transmitted through the liquid crystal; The second polarizer is configured to have a light cutoff in a range outside of its polarization range; as well as A driving voltage source is configured to apply a first voltage and a second voltage to the liquid crystal; wherein the first voltage and the second voltage cause the liquid crystal to switch between an open state and a closed state.

80. A liquid crystal assembly as described in claim 79, wherein the first polarizer and the second polarizer are cross-polarized.

81. A liquid crystal assembly as described in claim 79 or claim 80, wherein when one of the first voltage or the second voltage is applied to the liquid crystal, the liquid crystal is in the open state.

82. The liquid crystal assembly of any one of claims 79-81, wherein one of the first voltage or the second voltage is 0V.

83. The liquid crystal assembly of any one of claims 79-82, wherein when one of the first voltage or the second voltage is applied to the liquid crystal, the liquid crystal is in the closed state.

84. A liquid crystal assembly as described in any one of claims 79-83, wherein the second voltage is a bipolar square wave.

85. A liquid crystal assembly as described in claim 84, wherein the bipolar square wave has an amplitude of + / -0.1 to + / -5 Vp-p, + / -3 to + / -30 Vp-p, + / -16 Vp-p, + / -24 Vp-p or + / -30 Vp-p.

86. The liquid crystal assembly according to any one of claims 79 to 85, further comprising: an imaging lens located in a path of light transmitted through the second polarizer; as well as A camera is located in the path of light transmitted through the imaging lens.

87. A liquid crystal assembly as described in claim 86, wherein when the liquid crystal is in the open state, the liquid crystal rotates the polarization of polarized visible light transmitted through the first polarizer, thereby allowing the polarized visible light to pass through the second polarizer and reach the imaging lens with an attenuation of approximately 50% or less.

88. A liquid crystal assembly as described in claim 87, wherein when the liquid crystal is in the closed state, the liquid crystal does not rotate the polarization of the polarized visible light, and the polarized visible light transmitted through the first polarizer is blocked by the second polarizer.

89. The liquid crystal assembly of any one of claims 79-88, wherein the light cutoff range of the first polarizer and the second polarizer is a range in which the extinction ratio of the polarizer is poor or minimum.

90. A liquid crystal assembly as described in any one of claims 86-89, wherein light outside the light cutoff range of the first polarizer and the second polarizer passes through the first polarizer and the second polarizer without attenuation or with minimal attenuation and reaches the imaging lens.

91. A liquid crystal assembly as described in claim 90, wherein the minimum attenuation is 15% or less.

92. A liquid crystal assembly as described in any one of claims 79-91, wherein the light cutoff range of the first polarizer is about 700nm to about 800nm ​​or about 700nm, 725nm, 750nm, 775nm, 780nm, 785nm, 790nm, 795nm, 800nm, 805nm, 810nm, 815nm, 820nm, 825nm, 830nm, 835nm, 840nm, 850nm, 855nm, 860nm, 865nm, 870nm, 875nm, 880nm, 885nm, 890nm, 895nm or 900nm.

93. A liquid crystal assembly as described in any one of claims 79-92, wherein the light cutoff range of the second polarizer is about 700nm to about 800nm ​​or about 700nm, 725nm, 750nm, 775nm, 780nm, 785nm, 790nm, 795nm, 800nm, 805nm, 810nm, 815nm, 820nm, 825nm, 830nm, 835nm, 840nm, 850nm, 855nm, 860nm, 865nm, 870nm, 875nm, 880nm, 885nm, 890nm, 895nm or 900nm.

94. A liquid crystal assembly as described in any one of claims 79-93, wherein the light cutoff range of the first polarizer and the second polarizer is higher than the longest wavelength of the visible spectrum.

95. The liquid crystal assembly of any one of claims 79-94, wherein when the liquid crystal is in the open state, the liquid crystal assembly is configured to view visible light.

96. The liquid crystal assembly of any one of claims 79-95, wherein the liquid crystal assembly is configured to block visible light when the liquid crystal is in the closed state.

97. A liquid crystal assembly as described in any of claims 96, wherein the blocked visible light allows fluorescence imaging from the fluorophore.

98. A liquid crystal assembly as described in claim 97, wherein the fluorophore provides emission light emitted by the fluorophore.

99. A liquid crystal assembly as described in claim 97 or 98, wherein the excitation light is white light, NIR light, IR light or any other type of excitation light.

100. A liquid crystal assembly as described in any one of claims 79-99, wherein the excitation light is near-infrared light provided by a laser diode.

101. The liquid crystal assembly of any one of claims 79-100, further comprising a beam splitter.

102. A liquid crystal assembly as described in claim 101, wherein the beam splitter has a front surface and a rear surface.

103. A liquid crystal assembly as described in claim 102, wherein the front surface has a front surface coating and the rear surface has a rear surface coating.

104. A liquid crystal assembly as described in claim 103, wherein the front surface coating has a higher P-polarization reflectance than the rear surface coating.

105. A liquid crystal assembly as described in claim 103 or 104, wherein the front surface coating and the rear surface coating produce a front surface to rear surface reflectivity ratio of P-polarized light of at least 2:1, at least 4:1, at least 6:1, at least 8:1, at least 10:1, at least 12:1, at least 14:1, at least 16:1, at least 18:1 or at least 20:

1.

106. The liquid crystal assembly of any one of claims 103-105, wherein S-polarized reflections of the front surface and the rear surface of the beam splitter are minimized by the front surface coating and the rear surface coating.

107. A liquid crystal assembly as described in any of claims 79-106, wherein the first polarizer is configured to block or attenuate ghost images or secondary reflections from the beam splitter.

108. The liquid crystal assembly of claims 101-107, wherein the first polarizer is configured to block S-polarized light from the beam splitter.

109. The liquid crystal assembly of claims 101-108, wherein the first polarizer and the second polarizer are configured to allow P-polarized light to pass through the liquid crystal assembly and reach an imaging lens or camera when the liquid crystal is in the open state.

110. A liquid crystal assembly as described in any of claims 101-109, wherein the first polarizer allows a front-to-back reflectivity ratio of the beam splitter to be at least 2:1, 4:1, 6:1, 8:1, 10:1, 12:1, 15:1, 17:1 or 20:1 to pass therethrough and reach the imaging lens and camera.

111. A liquid crystal assembly as described in any of claims 101-110, wherein the total emissivity of the beam splitter is about 11% or less, about 10% or less, about 9% or less, about 8% or less, about 7% or less, about 6% or less, about 5% or less, about 4% or less or about 3% or less.

112. A liquid crystal assembly as described in any of claims 86, wherein the imaging lens and the camera are configured to view fluorescent images using a video rate sampling duration between 1 ms and 10 ms, between 2 ms and 20 ms, or 6 ms.

113. A liquid crystal assembly as described in any of claims 112, wherein the fluorescent image has reduced motion blur.

114. A method for imaging emission light from a fluorophore at an imaging system including a liquid crystal assembly, the method comprising: Allowing or directing visible light to reach the sample; directing excitation light toward the sample; directing the emitted light and the reflected visible light to the liquid crystal assembly; The liquid crystal assembly comprises: a first engineered polarizer located in a path of light from the sample; the first engineered polarizer configured to have a light cutoff between 600 nm and 900 nm or in a range of 800 nm to 880 nm; a liquid crystal disposed in a path of light transmitted through the first engineered polarizer; a second engineered polarizer located in a path of light transmitted through the liquid crystal; the second engineered polarizer being configured to have a light cutoff between 600 nm and 900 nm or in a range of 800 nm to 880 nm, wherein the first engineered polarizer and the second engineered polarizer are cross-polarized; and A driving voltage source configured to apply a first voltage and a second voltage to the liquid crystal; wherein the first voltage and the second voltage cause the liquid crystal to switch between an open state and a closed state; directing the emitted light and the reflected visible light through the first engineered polarizer, wherein the first engineered polarizer polarizes the reflected visible light below the light cutoff range, thereby producing polarized reflected visible light; directing the emitted light and the polarized reflected visible light through the liquid crystal; applying the first voltage to the liquid crystal to exhibit the open state for a first video rate sampling duration; rotating the polarization of the polarized reflected visible light using the liquid crystal in the open state; directing the emitted light and the polarized reflected visible light through the second engineered polarizer, wherein the second engineered polarizer allows the polarized reflected visible light to pass through the second engineered polarizer; detecting at an imaging sensor a first portion of the polarized reflected visible light that passes through the second engineered polarizer during the on state; applying the second voltage to the liquid crystal to exhibit the off state for a second video rate sampling duration; allowing the emitted light and the polarized reflected visible light to pass through the liquid crystal without rotating the emitted light and the polarized reflected visible light; blocking the polarized reflected visible light with the second engineered polarizer; and A portion of the emitted light during the off state is detected at the imaging sensor.

115. The method of claim 114, further comprising blocking secondary reflections of the reflected visible light using the first polarizer.

116. The method of claim 114 or 115, wherein the excitation light is provided to the sample via a laser diode.

117. The method of claim 116, further comprising: Turning off the laser diode to stop providing the excitation light; providing the second voltage to the liquid crystal to exhibit the off state during a third video rate sampling duration; detecting a dark background image of the sample; as well as The dark background image of the sample is subtracted from the detected portion of the emitted light to produce a fluorescent image.

118. The method of any one of claims 114-117, wherein the first video rate sample duration is about 2 ms to about 10 ms.

119. The method of any one of claims 114-118, wherein the second video rate sampling duration is about 8 ms to about 16 ms.

120. The method of any one of claims 117, wherein the third video rate sampling duration is approximately 8 ms to approximately 16 ms.

121. The method of any one of claims 114-120, wherein one of the first voltage or the second voltage is approximately 0V.

122. The method of any one of claims 114-121, wherein the second voltage is a bipolar square wave.

123. A method as claimed in claim 122, wherein the bipolar square wave has an amplitude of + / -0.1 to + / -5 Vp-p, + / -3 to + / -30 Vp-p, + / -16 Vp-p, + / -24 Vp-p or + / -30 Vp-p, or an amplitude between + / -16 V (32 Vp-p) and + / -30 V (60 Vp-p), including any values, or an amplitude between + / -30 V (60 Vp-p) and + / -100 V (200 Vp-p), including any values.

124. The method of claim 117, wherein the fluorescent image has reduced motion blur.

125. The method of any one of claims 114-124, wherein the second video rate sampling duration and the third video rate sampling duration allow for post-processing using digital gain.

126. The method of any one of claims 114-125, wherein the method further comprises orienting the first engineered polarizer to block undesired polarized light.

127. The method of claim 117, wherein the second video rate sampling duration and the third video rate sampling duration enhance the signal-to-noise ratio (SNR) and contrast-to-noise ratio (CNR) of the fluorescence image.

128. The method of claim 127, wherein the enhanced CNR achieves a fast frame rate.

129. The method of any one of claims 114-128, wherein the method provides real-time viewing of the reflected visible light and fluorescent images.

130. The method of any one of claims 114-129, further comprising directing the emitted light and the reflected light to a beam splitter before directing the emitted light and the reflected light to the liquid crystal assembly.

131. A method as claimed in claim 130, wherein the beam splitter has a front surface and a rear surface.

132. The method of claim 131, wherein the front surface has a front surface coating and the rear surface has a rear surface coating.

133. The method of claim 132, wherein the front surface coating has a higher P-polarization reflectance than the rear surface coating.

134. A method as claimed in claim 132 or 133, wherein the front surface coating and the rear surface coating produce a front surface to rear surface reflectivity ratio of P-polarized light of at least 2:1, at least 4:1, at least 6:1, at least 8:1, at least 10:1, at least 12:1, at least 14:1, at least 16:1, at least 18:1 or at least 20:

1.

135. The method of any one of claims 132-134, wherein S-polarized reflections of the front surface and the rear surface of the beam splitter are minimized by the front surface coating and the rear surface coating.

136. A method as described in any of claims 114-135, wherein the first polarizer is configured to block or attenuate ghost images or secondary reflections from a beam splitter.

137. A method as described in any of claims 130-136, wherein the first polarizer is configured to block S-polarized light from the beam splitter.

138. A method as described in any of claims 130-137, wherein the first polarizer and the second polarizer are configured to allow P-polarized light to pass through the liquid crystal assembly and reach the imaging lens and / or camera when the liquid crystal is in the open state.

139. A method as described in any of claims 130-138, wherein the first polarizer allows a front-to-back reflectivity ratio of the beam splitter to be at least 2:1, 4:1, 6:1, 8:1, 10:1, 12:1, 15:1, 17:1 or 20:1 to pass therethrough and reach the imaging lens and camera.

140. The method of any one of claims 130-139, wherein the total emissivity of the beam splitter is about 11% or less, about 10% or less, about 9% or less, about 8% or less, about 7% or less, about 6% or less, about 5% or less, about 4% or less, or about 3% or less.

141. A method of imaging abnormal tissue, cancer, tumor, vasculature, or structure in a sample from a subject, the method comprising generating an image of the vasculature or structure by fluorescence imaging using an imaging system as described in any of claims 1-42 or 79-106.

142. A method for imaging abnormal tissue, cancer, tumor, vasculature or structure in a sample from a subject according to any of the methods of claims 43-78 or 107-140, the method comprising generating an image of the abnormal tissue, cancer, tumor, vasculature or structure by fluorescence imaging using an imaging system comprising a liquid crystal assembly.

143. The method of claim 141 or 142, wherein the fluorescence imaged is autofluorescence, a contrast or imaging agent, a chemical agent, a radiolabeled agent, a radiosensitizer, a photosensitizer, a fluorophore, a therapeutic agent, an imaging agent, a diagnostic agent, a protein, a peptide, a nanoparticle or a small molecule or any combination thereof or any combination thereof.

144. The method of any one of claims 141-143, wherein the fluorescence imaged is autofluorescence, a contrast or imaging agent, a chemical agent, a radiolabeled agent, a radiosensitizer, a photosensitizer, a fluorophore, a therapeutic agent, an imaging agent, a diagnostic agent, a protein, a peptide, a nanoparticle or a small molecule, or any combination thereof.

145. The method of any one of claims 141-144, wherein the method further comprises administering a contrast or imaging agent to the subject.

146. A method of imaging abnormal tissue, cancer, tumor, vasculature, or structure in a fluorophore from a subject using the imaging system of any one of claims 1-42 or 79-106; the method comprising: (a) administering a contrast or imaging agent to the subject; as well as (b) generating an image of the abnormal tissue, cancer, tumor, vasculature or structure by imaging the contrast or imaging agent using an imaging system.

147. A method of imaging abnormal tissue, cancer, tumor, vasculature or structure in a fluorophore from a subject according to any of claims 43-78 or 107-140, the method further comprising: (a) administering a contrast or imaging agent to the subject; as well as (b) generating an image of the abnormal tissue, cancer, tumor, vasculature or structure by imaging the contrast or imaging agent using an imaging system.

148. The method of claim 146 or 147, wherein the contrast or imaging agent comprises a dye, a fluorophore, a fluorescent biotin compound, a luminescent compound, a chemiluminescent compound, or any combination thereof.

149. The method of claims 146-148, wherein the contrast or imaging agent further comprises a protein, a peptide, an amino acid, a nucleotide, a polynucleotide, or any combination thereof.

150. The method of claims 146-149, wherein the contrast or imaging agent further comprises tocilizumab.

151. The method of any one of claims 146-150, wherein the contrast or imaging agent absorbs wavelengths between about 200 nm and about 900 nm.

152. The method of any one of claims 146-151, wherein the contrast or imaging agent comprises: DyLight-680, DyLight-750, VivoTag-750, DyLight-800, IRDye-800, VivoTag-680, Cy5.5 or indocyanine green (ICG) and any derivatives thereof; fluorescein and fluorescent dyes (e.g., fluorescein isothiocyanate or FITC, naphthyl fluorescein, 4′,5′-dichloro-2′,7′-dimethoxyfluorescein, 6-carboxyfluorescein or FAM, etc.), carbocyanines, merocyanines, styryl dyes, oxonol dyes, phycoerythrin, erythrosine, eosin, rhodamine dyes (e.g., tetramethylcarboxy-rhodamine Rhodamine or TAMRA, carboxyrhodamine 6G, carboxy-X-rhodamine (ROX), lissamine rhodamine B, rhodamine 6G, rhodamine green, rhodamine red, tetramethylrhodamine (TMR), etc.), coumarin, coumarin dyes (e.g., methoxycoumarin, dialkylaminocoumarin, hydroxycoumarin, aminomethylcoumarin (AMCA), etc.), Oregon green dyes (e.g., Oregon Green 488, Oregon Green 500, Oregon Green 514, etc.), Texas Red, Texas Red-X, Spectrum Red, Spectrum Green, cyanine dyes (e.g., CY-3, Cy-5, CY-3.5, CY-5.5, etc.), ALEXA FLUOR dyes (e.g., ALEXA FLUOR 350, ALEXA FLUOR 488, ALEXAFLUOR 532, ALEXA FLUOR 546, ALEXA FLUOR 568, ALEXA FLUOR 594, ALEXA FLUOR 633, ALEXA FLUO R660, ALEXA FLUOR 680, etc.), BODIPY dyes (e.g., BODIPY FL, BODIPY R6G, BODIPY TMR, BODIPY TR, BODIPY 530 / 550, BODIPY 558 / 568, BODIPY 564 / 570, BODIPY 576 / 589, BODIPY 581 / 591, BODIPY 630 / 650, BODIPY 650 / 665, etc.), IRD dyes (e.g., IRD40, IRD700, IRD 800, etc.), 7-aminocoumarin, dialkylaminocoumarin reactive dyes, 6,8-difluoro-7-hydroxycoumarin fluorophores, hydroxycoumarin derivatives, alkoxycoumarin derivatives, succinimide esters, pyrene succinimide esters, pyridyl oxazole derivatives, aminonaphthyl dyes, dansyl chloride, dapoxyl dyes, Dapoxyl sulfonyl chloride, amine-reactive Dapoxyl succinimide esters, carboxylic acid-reactive Dapoxyl (2-aminoethyl) sulfonamide), bimane dyes, bimane thioglycolic acid, NBD dyes, QsY 35, or any combination thereof.

153. The method of any one of claims 146-152, wherein the administration comprises intravenous administration, intramuscular administration, subcutaneous administration, intraocular administration, intraarterial administration, peritoneal administration, intratumoral administration, intradermal administration, or any combination thereof.

154. The method of any one of claims 146-153, wherein the imaging comprises tissue imaging, ex vivo imaging, intraoperative imaging, or any combination thereof.

155. The method of any one of claims 146-154, wherein the sample is an in vivo sample, an in situ sample, an ex vivo sample, or an intraoperative sample.

156. The method of any one of claims 146-155, wherein the sample is an organ, an organ substructure, a tissue or a cell.

157. The method of any one of claims 146-156, wherein the sample autofluoresces.

158. The method of claim 157, wherein the autofluorescence of the sample comprises an ocular fluorophore, tryptophan, or protein present in a tumor or malignancy.

159. The method of any one of claims 146-158, wherein the method is used to visualize vascular flow or vascular patency.

160. The method of any one of claims 146-159, wherein the abnormal tissue, cancer, tumor, vasculature, or structure comprises a blood vessel, lymphatic vasculature, neuronal vasculature, or a CNS structure.

161. The method of any one of claims 146-160, wherein the imaging is angiography, arteriography, lymphangiography, or cholangiography.

162. The method of any one of claims 146-161, wherein the imaging comprises detecting a vascular abnormality, a vascular malformation, a vascular lesion, an organ or organ substructure, a cancer or diseased area, a tissue, a structure, or a cell.

163. The method of claim 162, wherein the vascular abnormality, vascular malformation or vascular lesion is an aneurysm, an arteriovenous malformation, a cavernous malformation, a venous malformation, a lymphatic malformation, a telangiectasia, a mixed vascular malformation, a spinal dural arteriovenous fistula or a combination thereof.

164. The method of any one of claims 146-163, wherein the organ or organ substructure is brain, heart, lung, kidney, liver, or pancreas.

165. The method of any one of claims 146-164, further comprising performing surgery on the subject.

166. The method of claims 146-165, wherein the surgery comprises angioplasty, cardiovascular surgery, aneurysm repair, valve replacement, aneurysm surgery, arteriovenous malformation or cavernous malformation surgery, venous malformation surgery, lymphatic malformation surgery, telangiectasia surgery, mixed vascular malformation surgery or dural arteriovenous fistula surgery, repair or bypass, arterial bypass, organ transplantation, plastic surgery, ophthalmic surgery, reproductive system surgery, stent insertion or replacement, plaque ablation, removal of the cancer or diseased area, tissue, structure or cells of the subject, or any combination thereof.

167. The method of any one of claims 146-166, wherein the imaging comprises imaging a vascular abnormality, cancer or diseased area, tissue, structure or cell of the subject following surgery.

168. The method of any one of claims 146-167, further comprising treating cancer in the subject.

169. A method as described in any of claims 146-168, further comprising repairing intracranial CNS vascular defects, spinal CNS vascular defects; peripheral vascular defects; removal of abnormal vascularized tissue; eye imaging and repair; anastomosis; reconstruction or plastic surgery; ablation or treatment of atherosclerotic plaques or restenosis; repair or resection (including selective resection), preservation (including selective preservation) of important organs or structures such as nerves, kidneys, thyroid, parathyroid, liver segments or ureters; identification and management during surgery (sometimes preservation, sometimes selective resection); diagnosis and treatment of limb ischemia; or treatment of chronic wounds.

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