# US 10,768,078 B2 · Sample processing improvements for quantitative microscopy > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 10,768,078 B2 - Document ID: US10768078B2 - Status: Granted - Kind: B2 - Application number: 16/367,791 - Filed: 2019-03-28 - Published: 2019-08-08 - Granted: 2020-09-08 - Earliest claimed: 2013-02-06 - Assignees: Alentic Microscience Inc. - Applicants: Alentic Microscience Inc. - Inventors: Alan Marc Fine; Hershel Macaulay - Web page: https://patentgrep.com/patent/US10768078B2 ## CPC - G01N 1/2813: {Producing thin layers of samples on a substrate, e.g. smearing, spinning-on (G01N1/30 takes precedence)} - G01N 1/38: Diluting, dispersing or mixing samples - G01N 21/31: Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry {(G01N21/72 takes precedence)} - G01N 33/49: Blood {(chemical methods for determining blood cell populations G01N33/5094; chemical analysis of blood groups or blood types G01N33/80)} - G02B 21/0008: {Microscopes having a simple construction, e.g. portable microscopes} - G02B 21/365: {Control or image processing arrangements for digital or video microscopes (G02B21/361, G02B21/362 take precedence)} - G01N 2001/386: {Other diluting or mixing processes} - G01N 2201/068: Optics, miscellaneous - G01N 2201/12: Circuits of general importance; Signal processing - G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q) - G02B: OPTICAL ELEMENTS, SYSTEMS OR APPARATUS ## Abstract Among other things, a diluted sample is generated based on mixing a small sample of blood with a one or more diluents. A thin film of the diluted sample is formed on the surface of a contact optical microscopy sensor. Red blood cells within a portion of the thin film of the diluted sample are illuminated using light of a predetermined wavelength. One or more images of the diluted sample are acquired based on illuminating the red blood cells within the portion of the thin film of the diluted sample. The acquired one or more images of the diluted sample are then processed. The mean corpuscular hemoglobin in the red blood cells within the portion of the thin film of the diluted sample is determined based on processing the acquired images of the diluted sample. ## Claims ### Claim 1 A method comprising: placing a fluid sample on a surface of a contact optical microscopy sensor, the fluid sample comprising particulate matter that includes an analyte to be measured, the analyte having a distinctive absorption spectrum; forming a film of the fluid sample between a transparent chamber lid and the surface of the contact optical microscopy sensor by moving the transparent chamber lid to a predetermined distance from the surface of the contact optical microscopy sensor, the distance determined by a spacer, the moving of the transparent lid to the predetermined distance causing the particulate matter of the fluid sample to be compressed into a shape to provide a uniform path length for light passing through the particulate matter; acquiring one or more images of the fluid sample based on illuminating at least a portion of the film of the fluid sample using light of a predetermined wavelength, processing the acquired one or more images of the fluid sample; and determining a mean amount of the analyte based on the processing of the one or more images of the fluid sample by a processor. ### Claim 2 (depends on 1) The method of claim 1, in which the particulate matter of the fluid sample is compressed into a cylindrical shape. ### Claim 3 (depends on 1) The method of claim 1, wherein a magnitude of the predetermined distance is such that lowering of the transparent chamber lid onto the surface of the contact optical microscopy sensor (i) constrains the particulate matter of the fluid sample to lie parallel to the surface of the contact optical microscopy sensor, and (ii) does not result in structural damage to the particulate matter of the fluid sample. ### Claim 4 (depends on 1) The method of claim 1, wherein processing the one or more acquired images comprises: segmenting one or more regions within the respective acquired images that each contain exactly one particle; and calculating the mean amount of the analyte based on the segmented one or more regions within the respective acquired images. ### Claim 5 (depends on 1) The method of claim 1, wherein forming the film comprises: placing the fluid sample on the surface of the contact optical microscopy sensor; and enabling the fluid sample placed on the surface of a contact optical microscopy sensor to settle. ### Claim 6 (depends on 1) The method of claim 1, wherein the acquired one or more images include at least a statistically significant number of particles in the fluid sample. ### Claim 7 (depends on 1) The method of claim 1, wherein the predetermined wavelength is based on extinction coefficients of wavelengths within an absorbance band of the analyte. ### Claim 8 (depends on 1) The method of claim 1, wherein the analyte comprises hemoglobin. ### Claim 9 (depends on 8) The method of claim 8, wherein the fluid sample comprises oxygenated hemoglobin and deoxygenated hemoglobin, and the method further comprises converting both types of hemoglobin to a third type of hemoglobin. ### Claim 10 (depends on 1) The method of claim 1, comprising mixing the fluid sample with a diluent to generate a diluted sample. ### Claim 11 (depends on 10) The method of claim 10, wherein the diluent comprises a nitrite. ### Claim 12 (depends on 10) The method of claim 10, wherein the fluid sample comprises blood, and wherein generating a diluted sample comprises mixing the sample of blood with a diluent such that the mixing results in sphering of red bloods cells within the diluted sample. ### Claim 13 (depends on 10) The method of claim 10, wherein the fluid sample comprises blood, and wherein the generated diluted sample has at least two of the following properties: has an isotonicity that is substantially equal to an isotonicity of red blood cells; has coagulation properties such that the generated diluted sample is less likely to coagulate compared to coagulation properties of red blood cells; and maintains a predetermined pH level of the generated diluted sample. ### Claim 14 (depends on 1) The method of claim 1, wherein the mean amount of the analyte is an average mass of hemoglobin per red blood cell in the fluid sample. ### Claim 15 (depends on 1) The method of claim 1, comprising determining the mean amount of the analyte with a coefficient of variation of less than 1 percent. ### Claim 16 (depends on 1) The method of claim 1, in which processing the acquired one or more images of the fluid sample comprises performing a calculation based on a measured intensity of the light of the predetermined wavelength at the surface of the contact optical microscopy sensor, the calculation including an assumption that the path length for light passing through the particulate matter is uniform within each pixel of the contact optical microscopy sensor.