# US 2014/0193050 A1 · Multispectral Imaging Systems and Methods > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 2014/0193050 A1 - Document ID: US20140193050A1 - Status: Published application - Kind: A1 - Application number: 14/149,015 - Filed: 2014-01-07 - Published: 2014-07-10 - Earliest claimed: 2013-01-10 - Assignees: Akoya Biosciences, Inc. - Applicants: Caliper Life Sciences, Inc. - Inventors: Peter J. Miller - Web page: https://patentgrep.com/patent/US20140193050A1 ## CPC - G06T 5/50: using two or more images, e.g. averaging or subtraction - G01J 3/2823: {Imaging spectrometer} - G01J 3/32: Investigating bands of a spectrum in sequence by a single detector - G01J 3/501: {Colorimeters using spectrally-selective light sources, e.g. LEDs} - G01J 3/51: using colour filters - G01J 2003/2826: {Multispectral imaging, e.g. filter imaging} - G06T 2207/20221: Image fusion; Image merging - G06T: IMAGE DATA PROCESSING OR GENERATION, IN GENERAL - G01J: MEASUREMENT OF INTENSITY, VELOCITY, SPECTRAL CONTENT, POLARISATION, PHASE OR PULSE CHARACTERISTICS OF INFRARED, VISIBLE OR ULTRAVIOLET LIGHT; COLORIMETRY; RADIATION PYROMETRY (light sources F21, H01J, H01K, H05B; investigating properties of materials by optical means G01N) ## Abstract The methods and systems disclosed herein include obtaining a first image of a sample, where the first image corresponds to light transmitted through the sample in a first plurality of distinct spectral bands, obtaining a second image of the sample, where the second image corresponds to light transmitted through the sample in a second plurality of distinct spectral bands, and where at least some members of the second plurality of spectral bands are different from the members of the first plurality of spectral bands, and combining the first and second images to form a multispectral image stack, where each pixel in the image stack includes information corresponding to at least four distinct spectral bands, and where the at least four distinct spectral bands include at least one member from the first plurality of spectral bands and at least one member from the second plurality of spectral bands. ## Claims ### Claim 1 A method, comprising: obtaining a first image of a sample, wherein the first image corresponds to light transmitted through the sample in a first plurality of distinct spectral bands; obtaining a second image of the sample, wherein the second image corresponds to light transmitted through the sample in a second plurality of distinct spectral bands, and wherein at least some members of the second plurality of spectral bands are different from the members of the first plurality of spectral bands; and using an electronic processor to combine the first and second images to form a multispectral image stack, wherein each pixel in the image stack comprises information corresponding to at least four distinct spectral bands, and wherein the at least four distinct spectral bands comprise at least one member from the first plurality of spectral bands and at least one member from the second plurality of spectral bands. ### Claim 2 (depends on 1) The method of claim 1, wherein each pixel in the image stack comprises information corresponding to at least five distinct spectral bands. ### Claim 3 (depends on 1) The method of claim 1, wherein each pixel in the image stack comprises information corresponding to at least six distinct spectral bands. ### Claim 4 (depends on 1) The method of claim 1, wherein the first plurality of distinct spectral bands comprises a spectral band in a green region of the electromagnetic spectrum having a full width at half maximum of 40 nm or less. ### Claim 5 (depends on 4) The method of claim 4, wherein the spectral band in the green region has a full width at half maximum of 30 nm or less. ### Claim 6 (depends on 2) The method of claim 2, further comprising using the electronic processor to unmix the image stack to obtain one or more component images, wherein each of the component images corresponds substantially only to contributions from one of the spectral contributors in the sample. ### Claim 7 (depends on 6) The method of claim 6, further comprising using the electronic processor to convert light intensity values corresponding to each of the pixels in the image stack to optical density units prior to unmixing the image stack. ### Claim 8 (depends on 6) The method of claim 6, further comprising using the electronic processor to analyze the sample based on the one or more component images to determine locations of features within the sample. ### Claim 9 (depends on 1) The method of claim 1, further comprising using a slide scanner to obtain the first and second images. ### Claim 10 (depends on 9) The method of claim 9, wherein the first plurality of spectral bands and the second plurality of spectral bands each comprise a shared spectral band, the method further comprising using the electronic processor to align the first and second images prior to forming the image stack based on information from the first and second images corresponding to the shared spectral band. ### Claim 11 A system, comprising: a light source configured to illuminate a sample; a detector configured to obtain one or more images of the sample; and an electronic processor coupled to the detector and configured to: obtain a first image of a sample, wherein the first image corresponds to light transmitted through the sample in a first plurality of distinct spectral bands; obtain a second image of the sample, wherein the second image corresponds to light transmitted through the sample in a second plurality of distinct spectral bands, and wherein at least some members of the second plurality of spectral bands are different from the members of the first plurality of spectral bands; and combine the first and second images to form a multispectral image stack, wherein each pixel in the image stack comprises information corresponding to at least four distinct spectral bands, and wherein the at least four distinct spectral bands comprise at least one member from the first plurality of spectral bands and at least one member from the second plurality of spectral bands. ### Claim 12 (depends on 11) The system of claim 11, wherein each pixel in the image stack comprises information corresponding to at least five distinct spectral bands. ### Claim 13 (depends on 11) The system of claim 11, wherein each pixel in the image stack comprises information corresponding to at least six distinct spectral bands. ### Claim 14 (depends on 11) The system of claim 11, wherein the first plurality of distinct spectral bands comprises a spectral band in a green region of the electromagnetic spectrum having a full width at half maximum of 40 nm or less. ### Claim 15 (depends on 14) The system of claim 14, wherein the spectral band in the green region has a full width at half maximum of 30 nm or less. ### Claim 16 (depends on 12) The system of claim 12, wherein the electronic processor is further configured to unmix the image stack to obtain one or more component images, wherein each of the component images corresponds substantially only to contributions from one of the spectral contributors in the sample. ### Claim 17 (depends on 16) The system of claim 16, wherein the electronic processor is further configured to convert light intensity values corresponding to each of the pixels in the image stack to optical density units prior to unmixing the image stack. ### Claim 18 (depends on 16) The system of claim 16, wherein the electronic processor is further configured to analyze the sample based on the one or more component images to determine locations of features within the sample. ### Claim 19 (depends on 11) The system of claim 11, further comprising a slide scanner that comprises the light source and the detector. ### Claim 20 (depends on 19) The system of claim 19, wherein the first plurality of spectral bands and the second plurality of spectral bands each comprise a shared spectral band, and the electronic processor is further configured to align the first and second images prior to forming the image stack based on information from the first and second images corresponding to the shared spectral band. ### Claim 21 A method, comprising: obtaining a first image of a sample using a color camera, wherein: the first image comprises a plurality of pixels, each pixel comprising three light intensity signals; for each pixel, each of the light intensity signals corresponds to illumination light in one or more of three distinct wavelength bands that is incident on the sample; and one of the wavelength bands corresponds to a green region of the electromagnetic spectrum and has a full width at half maximum of 40 nm or less; and using an electronic processor to: transform the first image to generate a corrected image, wherein the transformation comprises, for each one of the pixels, adjusting each of the three light intensity signals to reduce contributions thereto from illumination light in all but one of the three distinct wavelength bands; and convert the light intensity values in the corrected image to optical density units. ### Claim 22 (depends on 21) The method of claim 21, further comprising using the electronic processor to unmix the corrected image to obtain one or more component images. ### Claim 23 (depends on 22) The method of claim 22, further comprising using the electronic processor to analyze the sample based on the one or more component images to determine locations of features within the sample. ### Claim 24 A method, comprising: obtaining a first image of a sample using a color camera, wherein: the first image comprises a plurality of pixels, each pixel comprising three light intensity signals; for each pixel, each of the light intensity signals corresponds to light emitted from the sample in one or more of three distinct wavelength bands; and one of the wavelength bands corresponds to a green region of the electromagnetic spectrum and has a full width at half maximum of 40 nm or less; and using an electronic processor to: transform the first image to generate a corrected image, wherein the transformation comprises, for each one of the pixels, adjusting each of the three light intensity signals to reduce contributions thereto from the light emitted from the sample in all but one of the three distinct wavelength bands; and convert the light intensity values in the corrected image to optical density units. ### Claim 25 (depends on 24) The method of claim 24, further comprising using the electronic processor to unmix the corrected image to obtain one or more component images. ### Claim 26 (depends on 25) The method of claim 25, further comprising using the electronic processor to analyze the sample based on the one or more component images to determine locations of features within the sample.