WO2015020884A1 - Système de diagnostic portable à porte-échantillon jetable et microscope sur puce - Google Patents

Système de diagnostic portable à porte-échantillon jetable et microscope sur puce Download PDF

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Publication number
WO2015020884A1
WO2015020884A1 PCT/US2014/049226 US2014049226W WO2015020884A1 WO 2015020884 A1 WO2015020884 A1 WO 2015020884A1 US 2014049226 W US2014049226 W US 2014049226W WO 2015020884 A1 WO2015020884 A1 WO 2015020884A1
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WO
WIPO (PCT)
Prior art keywords
sample
sample holder
analysis module
biological sample
images
Prior art date
Application number
PCT/US2014/049226
Other languages
English (en)
Inventor
Kenneth Edward Salsman
Original Assignee
Nanoscopia (Cayman), Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from US13/959,304 external-priority patent/US9347962B2/en
Priority claimed from US13/959,122 external-priority patent/US20150036131A1/en
Priority claimed from US13/959,491 external-priority patent/US20150035966A1/en
Application filed by Nanoscopia (Cayman), Inc. filed Critical Nanoscopia (Cayman), Inc.
Publication of WO2015020884A1 publication Critical patent/WO2015020884A1/fr

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Classifications

    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/34—Microscope slides, e.g. mounting specimens on microscope slides
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/0004—Microscopes specially adapted for specific applications
    • G02B21/0008—Microscopes having a simple construction, e.g. portable microscopes
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/0004—Microscopes specially adapted for specific applications
    • G02B21/0088—Inverse microscopes
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/36—Microscopes arranged for photographic purposes or projection purposes or digital imaging or video purposes including associated control and data processing arrangements
    • G02B21/365—Control or image processing arrangements for digital or video microscopes

Definitions

  • This relates generally to diagnostic systems and, more particularly, to handheld diagnostic systems with disposable sample holders and chip-scale microscopes.
  • microscopic imaging is traditionally limited to a very narrow depth of field that shrinks as the magnification increases.
  • Scanning techniques are sometimes used to build a large depth of field image by combining multiple image frames at various focal lengths or to construct detailed images by stacking frames that have a focal plane at an angle to the sample surface.
  • Scanning techniques require precise control of the motion of the sample in order to accurately position the imaging frames.
  • Typical systems achieve this level of control using step and repeat image capture and calibrated motions stages.
  • Complex sample stage mechanisms and drive systems add significant weight, size, and cost to a system and can negatively affect its reliability and power requirements.
  • Figure 1 is a diagram of an illustrative diagnostic system having a sample holder and an analysis module for capturing and analyzing magnified images of cells and other biological specimens.
  • Figure 2 is a diagram of an illustrative chip-scale microscope.
  • Figure 3 is a cross-sectional top view of an illustrative sample holder.
  • Figure 4 is a cross-sectional top view of an illustrative sample holder and analysis module having a chip-scale microscope.
  • Figure 5 is a diagram of an illustrative diagnostic system having a sample holder for containing a sample, an analysis module having a chip-scale microscope for capturing magnified images of the sample, and an electronic device for obtaining sample analysis information from the analysis module.
  • Figure 6 is a flow chart of illustrative steps involved in operating a handheld diagnostic system of the type shown in Figures 1-5.
  • Figure 7 is a diagram of an illustrative handheld diagnostic system that employs an automated image capture mechanism using a sensor and a series of reference markings on a sample holder.
  • Figure 8 is a diagram of an illustrative handheld diagnostic system that employs an automated image capture mechanism using a portion of an image sensor and a series of reference markings on a sample holder.
  • Figure 9 is a top view of an illustrative sample holder having a series of reference markings for triggering an automated image capture mechanism in a handheld diagnostic system of the type shown in Figure 7.
  • Figure 10 is a top view of an illustrative sample holder having a series of reference markings for triggering an automated image capture mechanism in a handheld diagnostic system of the type shown in Figure 8.
  • Figure 11 is a flow chart of illustrative steps involved in operating a handheld diagnostic system of the type described herein.
  • Figure 12 is a cross-sectional side view of an illustrative sample holder having reference surfaces for obtaining accurate color, opacity, and reflectivity measurements from a sample using a chip- scale microscope.
  • Figure 13 is a cross-sectional side view of an illustrative sample holder having reference surfaces for obtaining accurate color, opacity, and reflectivity measurements from a sample using a chip- scale microscope.
  • Figure 14 is a cross-sectional top view of an illustrative sample holder having reference markings outside of the sample imaging frame for determining the size of a sample using a chip-scale microscope.
  • Figure 15 is a cross-sectional top view of an illustrative sample holder having reference markings that overlap the sample imaging frame for determining the size of a sample using a chip-scale microscope.
  • Figure 16 is a cross-sectional top view of an illustrative sample holder having reference structures outside of the sample imaging frame for determining the size of a sample using a chip-scale microscope.
  • Figure 17 is a cross-sectional top view of an illustrative sample holder having reference structures that overlap the sample imaging frame for determining the size of a sample using a chip-scale microscope.
  • Figure 18 is a flow chart of illustrative steps involved in operating a handheld diagnostic system of the type described herein.
  • Systems such as diagnostic systems may be provided with a disposable sample holder and a handheld, portable analysis module having a chip-scale microscope.
  • the disposable sample holder may have internal flow control structures and mechanisms for moving fluids, samples, particles, reactants and/or reagents from one part of the system to another.
  • the sample holder may have multiple test chambers for performing multiple tests simultaneously on a single sample.
  • the sample holder may be configured to protect the sample from contamination, to protect the user from exposure to infectious agents, and to provide the ability to add reagents to the sample.
  • the analysis module may be configured to receive the sample holder.
  • the chip-scale microscope may capture magnified images of the sample using the chip-scale microscope as the sample holder is inserted into the analysis module.
  • the handheld analysis module may be configured to connect with and provide sample analysis information to an electronic device such as a cellular telephone, a laptop, a tablet computer, or other portable computing device.
  • the electronic device may display images captured by the analysis module, may perform additional image analysis, and/or may control specific functions within the analysis module.
  • the analysis module and/or the electronic device may be configured to communicate sample analysis information from the analysis module over a communications network.
  • the chip-scale microscope may include an image sensor formed from complementary metal- oxide-semiconductor (CMOS) technology or other suitable image sensor integrated circuit technology.
  • CMOS complementary metal- oxide-semiconductor
  • the chip-scale microscope may also include optics for focusing light from the sample onto the image sensor.
  • An interchangeable illumination module in the analysis module may be used to illuminate the sample with a desired light source.
  • This type of diagnostic system may be used to analyze biological materials, bio-chemical materials, chemical materials, and/or other types of materials, and may be configured to perform spectral imaging operations such as narrow band imaging, multiple discrete band imaging, and fluorescence imaging (e.g., bio-fluorescence imaging as may be used in molecular analysis of biological samples).
  • spectral imaging operations such as narrow band imaging, multiple discrete band imaging, and fluorescence imaging (e.g., bio-fluorescence imaging as may be used in molecular analysis of biological samples).
  • the diagnostic system may be capable of performing medically viable diagnostics without requiring external wet chemistry or laboratory-trained personnel, may operate at low cost-per-test, and may be capable of operation in a variety of field environments (e.g., environments in which modern medical facilities are not available or are inconvenient).
  • the chip-scale microscope may be configured to capture spatially uniform imaging frames using an automated image capture mechanism.
  • the automated image capture mechanism may be based on a sensor that detects when the next imaging frame should be captured.
  • the sample holder may include a series of uniformly spaced markings. When a user inserts the sample holder into the analysis module, the series of uniformly spaced markings may be detected by a sensor in the analysis module. Upon detecting one of the markings, a control signal may be issued to capture an imaging frame using the chip-scale microscope.
  • This type of automated triggering ensures that the chip-scale microscope captures imaging frames at a uniform spatial distribution even when the sample is moving and even when the sample holder is inserted manually into the analysis module at variable speed.
  • the sensor may be a photodiode that is separate from the image sensor in the chip-scale microscope or may formed from an edge of the image sensor itself in the chip-scale microscope.
  • the sample holder may have built-in reference features for obtaining accurate colorimetric, opacity, and reflectivity measurements from a sample.
  • the built-in reference features may include reference surfaces having a predetermined color, transmissivity, and/or reflectivity.
  • the reference surfaces may be imaged by the chip-scale microscope and compared with images of the sample to determine the color, opacity, and/or reflectivity of the sample.
  • the sample holder may also include reference features such as reference markings having a known size and spacing or reference objects having a known size and spacing.
  • Images of reference markings or objects may be compared with images of the sample and may be used to determine the magnification of the chip-scale microscope, the size of a sample, the volume of a sample, the size or volume of cells within a sample, and/or other parameter values.
  • system 10 may include a sample holder such as sample holder 12 and an analysis module such as analysis module 14. As indicated by arrow 36, analysis module 14 may be configured to receive sample holder 12. Analysis module 14 may be configured to image and analyze samples in different types of disposable sample holders such as sample holder 12.
  • Sample holder 12 and analysis module 14 may be relatively small in size.
  • sample holder 12 may have a maximum lateral width of less than one inch, less than half of one inch, less than one quarter of one inch, less than four inches, or less than ten inches.
  • Analysis module 14 may have a maximum lateral length of less than three inches, less than two inches, less than one inch, less than four inches, or less than ten inches. Sample holder 12 and analysis module 14 may each be small enough to fit in a user's hand, if desired.
  • Sample holder 12 may have a sample chamber such as sample chamber 16, one or more reagent packs such as reagent pack 18, flow control components such as flow control components 20, and one or more test chambers such as test chambers 22.
  • Sample chamber 16 may be configured to receive a sample from a user of system 10. For example, a user may place a swab on which a sample has been collected into sample chamber 16, or a user may place a sample on its own (e.g., a blood sample that has been collected with a lancet) into sample chamber 16.
  • the sample may be a biological sample including cells or other biological elements.
  • system 10 may be used to analyze and capture high-magnification images of other types of samples (e.g., other biological specimen or other particles or materials). Arrangements in which system 10 is used to image cells are sometimes described herein as an example.
  • Reagent pack 18 may be used to contain reagents until they are introduced to the sample in sample chamber 16. If desired, there may be one, two, or more than two reagent packs within a single sample holder.
  • Flow control components 20 may be used to control the flow of a sample within sample holder 12 without requiring electrical power.
  • Flow control components 20 may, for example, include one or more compartments of chemicals configured to react with each other and produce gas which then forces the sample through a channel in the sample holder and distributes portions of the sample into respective test chambers 22 in sample holder 12.
  • flow control components 20 may include a pack or compartment of acetic acid (vinegar) and a pack or compartment of sodium bicarbonate (baking soda). When combined, the sodium bicarbonate and acetic acid may produce carbon dioxide gas which then pushes the sample through the channel in a smooth, continuous, and predictable manner.
  • This type of configuration is advantageous in that it does not require electrical power and also avoids the abrupt jerking of the sample which occurs when a pump is used to control the flow of a sample.
  • other types of flow control structures such as one or more pumps may be used to move the sample from one location in sample holder 12 to another location in sample holder 12.
  • Test chambers 22 may each be configured to receive a portion of the sample from sample chamber 16. Each test chamber 22 may, for example, contain a different marker such as marker 98 configured to tag a specific chain of DNA, RNA, or protein.
  • markers 98 in test chambers 22 may be configured to locate and mark specific nucleic acids or proteins (e.g., nucleic acids or proteins associated with a bacterium, virus, poison, fungus, parasite, etc.) in the sample with specific colors (e.g., using stains, dyes, and/or fluorescent tagging).
  • Each marker 98 in each test chamber 22 may be used to identify a different bacteria, virus, poison, fungus, or parasite in a single sample, thereby providing system 10 with the ability to perform multiple tests on a single sample simultaneously.
  • Illustrative examples of substances or structures that may be identified using system 10 include S. aureus, Coagulase-negative staphylococci (CNS), E. faecalis, E. faecium and other Enterococci, E. coli, K. pneumoniae , P.
  • aeruginosa C. albicans, C. parapsilosis, C. tropicalis, C. glabrata, C. krusei, Listeria, foot-and-mouth disease virus, Methicillin-resistant Staphylococcus aureus (MRSA), and malaria parasites such as P. falciparum and other malaria parasites.
  • MRSA Methicillin-resistant Staphylococcus aureus
  • markers 98 may be configured to tag structures within the sample using a process referred to as immunolabeling.
  • markers 98 may include tagged conjugate antibodies that are configured to attach themselves to locations where the corresponding target antigen is found.
  • the conjugate antibodies may be tagged with a fluorescent compound, gold beads, an epitope tag, or an enzyme that produces a colored compound.
  • markers 98 may be configured to attach fluorophores to olignoucleotides complementary to the target RNA molecules (as an example).
  • Reagents and markers in sample holder 12 can be stored in active or in freeze-dried form. Substances stored in freeze-dried form may be activated with the addition of water and/or other reagents.
  • Sample holder 12 allows the chemistry required for sample processing and the sample itself to be sealed and safely contained once acquired and allows for the processing to be automated within a low-cost structure. If desired, sample holder 12 may be disposed with the sample when the sample analysis is complete or may be used to keep the sample in a safe, contained enclosure until further analysis can be performed in a fully-equipped laboratory. The chemistry, sample processing, and internal structure of a given sample holder may be customized depending on the type of test(s) or analysis being performed. Sample holders 12 may be provided with a common external mechanical structure so that analysis modules 14 are compatible with many different types of sample holders 12, each of which is designed for performing a specific set of tests. Sample holder 12 may be produced inexpensively in high volume and may be disposed of after a single use (if desired).
  • Analysis module 14 may include chip-scale microscope 24, illumination module 26, sample holder receiving structures 28, storage and processing circuitry 30, input-output components 32, and output ports 34.
  • Chip-scale microscope 24 may include an image sensor for imaging samples within sample holder 12 and optics such as one or more lenses and/or mirrors for focusing light from the sample onto the image sensor.
  • Illumination module 26 may include one or more light sources (e.g., one or more light-emitting diodes, arc lamps, lasers, or other suitable type of light source) for illuminating the sample in sample holder 12. Illumination module 26 may also include one or more optical structures such as mirrors, gratings, and/or condenser lenses for focusing light from the light source onto the sample.
  • light sources e.g., one or more light-emitting diodes, arc lamps, lasers, or other suitable type of light source
  • Illumination module 26 may also include one or more optical structures such as mirrors, gratings, and/or condenser lenses for focusing light from the light source onto the sample.
  • Analysis module 14 may include a housing having sample holding receiving structures 28 for receiving sample holder 12.
  • Sample holder receiving structures 28 may include an opening into which sample holder 12 is inserted. The opening may be provided with guide rails or other alignment structures to facilitate insertion of sample holder 12 into analysis module 14.
  • sample holder receiving structures 28 may include structures for controlling the rate of insertion of sample holder 12 into analysis module 14.
  • the opening into which sample holder 12 is inserted may include a pattern of gears or other structures configured to mate with a corresponding pattern of gears on an external surface of sample holder 12. Such structures may be used to ensure that the rate at which sample holder 12 is guided into analysis module 14 is kept constant or within a given range (if desired).
  • Chip-sale microscope 24 may capture images of the sample as sample holder 12 is being inserted into analysis module 14.
  • Storage and processing circuitry 30 may include volatile memory (e.g., static or dynamic random-access memory), non-volatile memory (e.g., flash memory), microprocessors, integrated circuits, printed circuit boards, or other circuitry. Storage and processing circuitry 30 may be used for storing, processing, and analyzing image data captured using chip-scale microscope 24, and/or for operating components such as illumination module 26 and input-output components 32.
  • volatile memory e.g., static or dynamic random-access memory
  • non-volatile memory e.g., flash memory
  • microprocessors integrated circuits
  • printed circuit boards or other circuitry.
  • Storage and processing circuitry 30 may be used for storing, processing, and analyzing image data captured using chip-scale microscope 24, and/or for operating components such as illumination module 26 and input-output components 32.
  • Storage and processing circuitry 30 may include communications circuitry such as circuitry coupled to output ports 34.
  • Storage and processing circuitry 30 may include wireless communications circuitry for conveying data such as image data, sample analysis information, diagnosis information, etc. to external equipment such as a computer, a handheld electronic device, a cellular telephone, a network router, a network antenna, etc.
  • wireless communications circuitry associated with circuitry 30 may be configured to transmit and/or receive data at WiFi® frequencies (e.g., 2.4 GHz and 5 GHz), Bluetooth® frequencies (e.g., 2.4 GHz), cellular telephone frequencies (e.g., 85- MHz, 900 MHz, 1800 MHz, 1900 MHz, and 2100 MHz), or other frequencies.
  • Output ports 34 may include one or more universal serial bus (USB) ports, serial ports, audio ports, video ports, etc. coupled to storage and processing circuitry 30.
  • USB universal serial bus
  • Data that may be transmitted using ports 34 or wireless communications circuitry associated with circuitry 30 may include identification data associated with a particular analysis module, identification data associated with a particular sample holder, identification data associated with a sample, geographic location data associated with the location of the analysis module, sample analysis information resulting from analysis of a sample within sample holder 12, raw and/or processed imaging data obtained using chip-scale microscope 24, and/or other information.
  • Sample analysis information may, for example, include a medical diagnosis or an identification of which substances or structures were found to be present or absent in the sample.
  • Illustrative examples of procedures that may be performed using system 10 include whole blood cell analysis, cell counting, Complete Blood Count (CBC), nucleic acid amplification, PNA-FISH® bacterial testing, antigen and antibody infectious disease detection, and other tests. Because system 10 is handheld and portable, such tests may be performed in locations where laboratory facilities are unavailable or inconvenient for a user.
  • CBC Complete Blood Count
  • PNA-FISH® bacterial testing antigen and antibody infectious disease detection
  • System 10 may provide a user with the ability to interact with analysis module 14.
  • User interactions may include inputting identification information (e.g., information identifying a sample, a sample donor, a geographic location, etc.) and obtaining output information (e.g., reading the result of an analysis performed using chip-scale microscope 24).
  • analysis module 14 may have input-output components 32 such as keypads, virtual keypads, buttons, displays, or other suitable input-output components.
  • Input-output components 32 may include circuitry coupled to one or more output ports such as output port 34 mounted in a housing structure.
  • chip-scale microscope 24 An illustrative configuration for chip-scale microscope 24 is shown in Figure 2. As shown in Figure 2, chip-scale microscope 24 may include optics such as optics 38 and an image sensor
  • Image sensor 40 may include an array of image pixels such as pixel array 42 and image sensor circuitry such as image sensor circuitry 44.
  • Image sensor circuitry 44 may include row control circuitry, column readout circuitry, analog-to-digital conversion circuitry, and other circuitry associated with capturing raw data using image pixel array 42 of image sensor 40.
  • Circuitry 30 of Figure 1 may, for example, be used to control imaging functions performed using chip-scale microscope 24.
  • Optics 38 may include optical elements for gathering light from the sample in sample holder 12 and focusing the light onto pixel array 42 of image sensor 40.
  • Optics 38 may include one or more objective lenses, one or more mirrors, one or more layers of glass, and/or other optical structures for focusing light from the sample onto image sensor 40.
  • Optics 38 may, for example, be interposed between the sample (when sample holder 12 is inserted into analysis module 14) and image sensor 40.
  • Optics 38 may be characterized by a magnification of 10OOx, 400x, 200x, or other suitable magnification; may be characterized by a numerical aperture of less than 0.5, less than 1.0, less than 1.5, or greater than 1.5; and may be characterized by a working distance of 5 mm, greater than 5 mm, less than 5 mm, less than 10 mm, or greater than 10 mm.
  • Chip-scale microscope 24 may be configured to achieve a depth of field of about 125 microns, about 130 microns, about 120 microns, about 100 microns, less than 100 microns, greater than 100 microns, or greater than 120 microns.
  • Microscope objective 38 may, if desired, operate with an air medium, thereby eliminating the need for an immersion liquid between the front lens element and the sample.
  • Chip-scale microscope 24 may be equipped to obtain volumetric data using the automatic focus functionality of image sensor 40 without requiring an automated stage.
  • sample holder 12 may include a first portion such as sample-receiving portion 62, and a second portion such as sample imaging portion 64.
  • Sample-receiving portion 62 may include reagent pack 18, flow control components 20, and sample chamber 16. As described in connection with Figure 1 , reagent pack 18 may be used to contain reagents until they are introduced to the sample in sample chamber 16. Initially, reagent pack 18 may be sealed from sample chamber 16. Upon breaking the seal, reagents in reagent pack 18 may be allowed to interact with a sample such as sample 80 in sample chamber 16 via path 66.
  • Flow control components 20 may provide a sample distribution mechanism for distributing portions of sample 80 in sample chamber 16 to respective test chambers 22.
  • Flow control components 20 may be implemented as a gas generating component having two adjacent chambers 48 and 50.
  • Chamber 48 may contain a first reactant such as liquid reactant 48A (e.g., acetic acid).
  • Chamber 50 may contain a second reactant such as solid or powder reactant 50A (e.g., sodium bicarbonate).
  • First and second reactants 48A and 50A may be selected to be stable chemicals (e.g., acetic acid (vinegar) and sodium bicarbonate (baking soda), respectively) that generate a gas such as carbon dioxide when mixed.
  • Chambers 48 and 50 may initially be separated by structural member 70 (e.g., a plastic seal).
  • structural member 70 e.g., a plastic seal
  • chemical reactants 48A and 50A may be allowed to interact and a chemical reaction may occur, leading to the release of a significant volume of gas (e.g., carbon dioxide).
  • the gas produced may provide pressure to chamber 16 via path 68, which may in turn move sample 80 in sample chamber 16 through channel 52 in direction 82. Portions of sample 80 may be distributed to respective test chambers 22 in sample imaging portion 64.
  • a particle filter such as particle filter 54 may be configured to filter sample 80 to prevent certain substances or structures from passing through channel 52 to sample imaging portion 64.
  • Vent line 56 may allow air to escape via exit port 58 and may be used in regulating the flow of air and the movement of sample 80, if desired.
  • sample distribution mechanisms may be employed to distribute sample 80 in sample chamber 16 to test chambers 22.
  • the use of sodium bicarbonate and acetic acid is merely as an example.
  • Sample-receiving portion 62 may have a clamshell shape with first and second portions 62A and 62B connected by a bendable joint such as bendable joint 60. With this type of configuration, sample- receiving portion 62 of sample holder 12 may be configurable in open and closed positions. In the open configuration (as shown in Figure 3), compartments within sample-receiving portion 62 may be sealed. For example, reagent pack 18 may be sealed and compartments 48 and 50 may be sealed and separated from each other. While sample-receiving portion 62 is open, a user may place a sample into sample chamber 16 and may then close sample-receiving portion 62 by bending sample-receiving portion 62 at bendable portion 60.
  • a protrusion such as protrusion 46 (e.g., a structure having one or more sharp edges) within portion 62 may puncture reagent pack 18 and seal 70, thereby allowing reagents in reagent pack 18 to interact with the sample in sample chamber 16 while also allowing reactants 48A and 50A in compartments 48 and 50 to interact with each other.
  • Sample 80 is mixed with reagents in reagent pack 18 and is moved through channel 52 to test chambers 22. With this type of configuration, the appropriate chemistry and sample processing may automatically occur within sample holder 12 by merely closing sample-receiving portion 62 after placing sample 80 in sample chamber 16.
  • sample chamber 16 may include a permeable or semi-permeable cover such as a neoprene membrane through which a needle may be inserted (as an example).
  • a permeable or semi-permeable cover such as a neoprene membrane through which a needle may be inserted (as an example).
  • each test chamber 22 in sample holder 12 may contain a different marker for tagging a specific substance (e.g., via staining, dying, fluorescent tagging, etc.).
  • one test chamber 22 may contain a marker for tagging foot-and-mouth disease virus, while another test chamber 22 may contain a marker for tagging Methicillin-resistant Staphylococcus aureus (MRSA).
  • MRSA Methicillin-resistant Staphylococcus aureus
  • test tagging different portions of a single sample in sample holder 12 with different markers may be performed simultaneously on a single sample.
  • different types of tests e.g., tests for different types of bacteria, viruses, fungi, parasites, etc.
  • Sample holder 12 may be formed from plastic, glass, metal, carbon fiber and/or other fiber composites, ceramic, glass, wood, other materials, or combinations of any two or more of these materials.
  • Sample imaging portion 64 may be designed for microscopic imaging (e.g., may be partially or fully transparent so that sample 80 in test chambers 22 may be illuminated for microscopic imaging).
  • FIG 4 is a cross-sectional top view of system 10 in which sample holder 12 has been inserted into analysis module 14.
  • analysis module 14 may include a housing such as housing 84 having an opening such as opening 86. Opening 86 may be have a shape that corresponds to the shape of sample imaging portion 64 of sample holder 12 so that sample imaging portion 64 of sample holder 12 may be inserted into analysis module 14.
  • Sample holder 12 may be engaged with analysis module 14 by inserting sample imaging portion 64 of sample holder 12 into opening 86 in direction 88.
  • output port 34 may be implemented as a USB connector for coupling module 14 to external equipment such as a computer, cell phone, laptop computer, tablet computer, etc.
  • output port 34 may also be configured to provide power to components within analysis module 14.
  • port 34 may include a power supply for providing power to illumination module 26, image sensor 40, and storage and processing circuitry 30. This is, however, merely illustrative. If desired, electrical components in analysis module 14 may receive power from an external power source.
  • Storage and processing circuitry 30 may be implemented using a printed circuit substrate such as printed circuit substrate 76, integrated circuits or other electrical components such as electrical components 78, and/or other circuitry in analysis module 14.
  • Image sensor 40 may be coupled to printed circuit board 76 using an array of solder balls (e.g., a ball grid array) or may be coupled to printed circuit board 76 using other mounting techniques.
  • Printed circuit board 76 may include metal traces 90 for electrically coupling image sensor 40 to other circuitry such as integrated circuit 78.
  • Lighting components 26 may be mounted in analysis module 14 so that light from lighting sources 74 passes through test chambers 22 of sample holder 12 during sample analysis operations.
  • illumination module 26 may include one or more light sources such as light sources 74 (e.g., one or more light-emitting diodes, arc lamps, lasers, or other suitable type of light source) for illuminating sample 80 in sample holder 12.
  • Light sources 74 may be white light sources or may be configured to emit different colors of light.
  • light source 74 may be white light sources that are provided with different colored filters.
  • Illumination module 26 may include one or more optical structures such as lenses 92L mirror 92M for focusing light 94 from light source 74 onto sample 80.
  • light sources 74 may produce light 94 of a desired color and intensity.
  • Light 94 may be directed through sample holder 12 (when sample holder 12 is inserted into analysis module 14) towards image sensor 40.
  • Illumination module 26 may be interchangeable so that different types of microscopy may be performed.
  • a first illumination module may be used to perform fluorescence microscopy using chip-scale microscope 24, and a second illumination module may be used to perform bright field microscopy using chip-scale microscope 24.
  • the first illumination module may be removed from analysis module 14 and the second illumination module may be installed in its place (or vice versa).
  • Light 94 may pass through sample 80 and may be focused onto image sensor 40 using optics 38.
  • optics 38 may include one or more objective lenses, one or more mirrors, one or more layers of glass, and/or other optical structures for focusing light from sample 80 onto image sensor 40.
  • one or more optical filters such as optical filter 96 may be interposed between optics 38 and image sensor 40.
  • optical filters in analysis module 14 such as optical filter 96 may be interchangeable so that different types of microscopy may be performed.
  • Illustrative types of filters that may be used in analysis module 14 include longpass filters, colored and/or neutral density filters, absorptive filters, interference filters, dichroic filters, polarization filters, other suitable types of filters, or a combination of any two or more of these types of filters.
  • flow control components 20 may automatically be activated to distribute portions of sample 80 into respective test chambers 22 (as shown in Figure 4).
  • the user may then insert sample holder 12 into analysis module 14 by sliding sample imaging portion 64 of sample holder 12 into opening 86 of analysis module 14 in direction 88.
  • sample imaging portion 64 of sample holder 12 moves in direction 88 within cavity 86, each test chamber 22 may pass through light 94 and over image sensor 40.
  • sample 80 in the rightmost chamber 22 will be the first to pass through light 94 over image sensor 40 and will therefore be the first specimen to be imaged with image sensor 40.
  • sample 80 in the second chamber 22 from the right will pass through light 94 over image sensor 40 and will therefore be the second specimen to be imaged with image sensor 40.
  • light 94 may successively illuminate sample 80 in each test chamber 22, and images may be successively captured of sample 80 in each chamber 22 as each chamber 22 is moved across the field of view of chip-scale microscope 24.
  • Image sensor 40 may include circuitry for automatically triggering each image capture operation as test chambers 22 move across image sensor 40.
  • FIG. 5 is a diagram showing how a handheld diagnostic system such as system 10 may be configured to communicate with computing equipment such as computing equipment 102.
  • Computing equipment 102 may be a portable electronic device (e.g., a mobile phone, a personal digital assistant, a laptop computer, a tablet computer, or other computing equipment).
  • Computing equipment 102 may include a display such as display 104 for presenting visual information to a user based on data received from system 10.
  • display 104 may be used in displaying images of samples acquired by system 10 (sometimes referred to as sample image data) and/or may be used in displaying sample analysis information (e.g., may present a list of bacteria, viruses, poisons, fungi, or parasites which were found present in the sample).
  • Computing equipment 102 may have a user input interface for gathering input from a user and for supplying output to a user.
  • the user input interface may include user input devices such as keyboard, keypads, mice, trackballs, track pads, etc.
  • display 104 may be touch-sensitive (i.e., display 104 may be a touch screen) and may be used to gather user input from a user.
  • Computing equipment 102 may also include equipment for supplying output such as speakers for providing audio output, status indicator lights for providing visible output, etc.
  • Computing equipment 102 may include a data port such as data port 110.
  • Data port 110 may be connected to analysis module 14 using a cable such as cable 1 12.
  • cable 112 may have a connector such as connector 1 14 configured to mate with output port 34 of analysis module 14 ( Figure 4).
  • cable 1 12 may have a connector such as connector 116 configured to mate with data port 110 of computing equipment 102.
  • Sample image data and/or sample analysis information may be conveyed from analysis module 14 to computing equipment 102 via cable 1 12. This is, however, merely illustrative. If desired, information may be conveyed from sample analysis module 14 to computing equipment 102 over a wireless network.
  • data port 1 10 may be a Universal Serial Bus (USB) port and may be configured to receive output port 34 of analysis module 14 directly (without requiring cable 1 12).
  • USB Universal Serial Bus
  • Computing equipment 102 may be used to analyze sample image data and/or sample analysis information (e.g., to produce images of the sample from raw image data, to produce enhanced images of the sample, to analyze images of the sample to produce sample evaluation information or diagnosis information, etc.). Computing equipment 102 may, if desired, transmit data from system 10 to computing and data processing equipment 118 via communications network 106.
  • Communications network 106 may include wired and wireless local area networks and wide area networks (e.g., the internet).
  • Computing equipment 102 may be connected to network 106 using a link such as link 108 (e.g., a wired link that uses a modem or wireless link such as a local wireless link), and computing and data processing equipment 1 18 may be connected to network106 using a link such as link 120 (e.g., a wired link that uses a modem or wireless link such as a local wireless link).
  • link 108 e.g., a wired link that uses a modem or wireless link such as a local wireless link
  • link 120 e.g., a wired link that uses a modem or wireless link such as a local wireless link
  • Computing and data processing equipment 1 18 may be a remote mainframe computer, may be a cloud computing network (i.e., a network of computers on which software can be run from computing equipment 102) or other computing equipment. If desired, computing and data processing equipment 1 18 may be used to perform advanced analysis on sample image data and/or sample analysis information from system 10 (e.g., advanced analysis that requires more
  • Figure 6 is a flow chart of illustrative steps involved in using a system such as system 10 of Figures 1 -5 in acquiring images of cells or other samples.
  • a sample may be injected into a sample chamber in a sample holder such as sample chamber 16 in sample holder 12.
  • the sample holder may be closed to automatically activate the sample distribution mechanism and thereby distribute portions of the sample from the sample chamber to respective test chambers in the sample holder.
  • the sample distribution mechanism may be controlled by flow control components such as flow control components 20 of Figure 3.
  • a user may insert the sample holder into an analysis module such as analysis module 14 of Figure 4 to trigger image capture operations and to thereby capture images of the sample using a chip-scale microscope in the analysis module.
  • Circuitry in analysis module 14 may be configured to automatically trigger image capture operations as sample holder 12 is inserted into analysis module 14. During insertion, images may be captured of the sample in each test chamber as each test chamber passes through the field of view of the chip-scale microscope in analysis module 14.
  • raw image data, processed image data, and/or sample analysis information may be transmitted to external computing equipment such as a portable electronic device (e.g., a cellular telephone, a tablet computer, a laptop computer, or other portable electronic device).
  • external computing equipment such as a portable electronic device (e.g., a cellular telephone, a tablet computer, a laptop computer, or other portable electronic device).
  • images of the sample and/or sample analysis information e.g., diagnosis information, test results, or other sample evaluation data
  • the pixel data, processed pixel data, and/or sample analysis information may be provided to the external computing equipment through a USB port in the analysis module or using wireless communications circuitry in the analysis module.
  • Sample imaging portion 64 of sample holder 12 may have uniformly spaced reference markings distributed along the length of sample imaging portion 64 (i.e., along the portion of sample holder 12 that is inserted into analysis module 14). Reference markings in sample holder 12 may be detected by a sensor in analysis module 14 and may be configured to trigger an automated image capture mechanism whereby chip-scale microscope 24 captures imaging frames at a uniform spatial distribution.
  • Figure 7 is a diagram of a portion of system 10 showing how system 10 may include a sensor for detecting reference markings on sample holder 12 for automatically triggering image capture operations as sample holder 12 is inserted into analysis module 14.
  • sample holder 12 may have reference markings such as reference markings 122. There may be five, ten, fifteen, twenty, more than twenty, or less than twenty reference markings 122 on sample holder 12. Reference markings 122 may be separated from each other by a distance D.
  • Analysis module 14 may include a sensor such as sensor 126 and a light source such as light source 124.
  • Light source 124 e.g., a light-emitting diode light source or other type of light source
  • Sensor 126 and light source 124 may be aligned such that sensor 126 is configured to receive light 128 emitted by light source 124.
  • Sensor 126 may include one or more photodiodes or other suitable type of light sensor.
  • Sensor 126 may be coupled to a trigger generator such as trigger generator 130 and control circuitry such as control circuitry 132.
  • Control circuitry 132 and trigger generator 130 may, for example, form part of storage and processing circuitry 30 ( Figure 4).
  • Control circuitry 132 may be coupled to image sensor 40 and may be configured to issue control signals to image sensor 40 based on signals received from sensor 126 via trigger generator 130.
  • sensor 126 may be configured to detect when reference markings 122 pass through light 128.
  • trigger generator 130 may generate a trigger signal for control circuitry 132, which may in turn issue control signals to image sensor 40 to capture an imaging frame.
  • image sensor 40 may capture an image of sample 80 in sample holder 12. This automated image capture mechanism ensures that imaging frames are captured at a uniform spatial distribution even when the sample is moving and even when the sample holder is inserted manually into the analysis module at variable speeds.
  • the distance D between reference markings 122 may be any suitable distance (e.g., 1 mm, 2 mm, 3 mm, 5 mm, less than 5 mm, or more than 5 mm).
  • multiple imaging frames may be captured of each portion of sample 80 in each respective test chamber 22. Capturing multiple imaging frames of sample 80 at uniform spatial distribution may allow processing circuitry (e.g., processing circuitry 30) to build a large depth of field image of sample 80 by combining multiple imaging frames at different focal lengths; to construct a detailed image of sample 80 by stacking frames that have a focal plane at an angle to the sample surface, thereby providing a focal region that is larger than a single frame focal region; and to build images of large samples by stitching together multiple imaging frames that have a uniform spatial distribution .
  • processing circuitry e.g., processing circuitry 30
  • Sensor 126 need not be separate from image sensor 40. If desired, a portion of pixel array 42 ( Figure 2) of image sensor 40 may be used to detect reference markings 122. A diagram illustrating how sensor 126 may be formed from a portion of image sensor 40 is shown in Figure 8. As shown in Figure 8, sensor 126 may be located on an edge of image sensor 40 and may be formed from a portion of pixel array 42 (e.g., one or more rows or columns of pixels in pixel array 42, one or more individual pixels or groups of pixels in pixel array 42, etc.).
  • Sensor 126 may be configured to detect when reference markings 122 pass through light 94 emitted by illumination module 26. Upon detecting one of reference markings 122, trigger generator 130 may generate a trigger signal for control circuitry 132, which may in turn issue control signals to image sensor 40 to capture an imaging frame. Thus, each time a reference marking 122 in sample holder 12 passes over sensor 126 at the edge of image sensor 40 in analysis module 14, image sensor 40 may capture an image of sample 80 in sample holder 12. This automated image capture mechanism ensures that imaging frames are captured at a uniform spatial distribution even when the sample is moving and even when the sample holder is inserted manually into the analysis module at variable speeds.
  • FIG. 9 A top view of the arrangement of Figure 5 is shown in Figure 9.
  • reference markings 122 may pass through field of view 126' of sensor 126 as sample holder 12 is moved in direction 88.
  • control circuitry 132 may issue a control signal to image sensor 40 to capture an imaging frame.
  • Region 40' indicates the field of view of image sensor 40 during an image capture.
  • tick marks 122T will each successively pass over sensor 126, and chip-scale microscope 24 will capture a corresponding series of imaging frames as test chambers 22 move across imaging frame region 40' from edge 134 to edge 136.
  • each imaging frame region 40' may be separated from the rightmost edge of the adjacent imaging frame region by a distance D (i.e., a distance corresponding to the separation between tick marks 122T).
  • tick marks 122T may be spaced such that a region of overlap exists between adjacent imaging frames so that the images can be integrated as tiles to generate a larger field of view than chip-scale microscope 24 can achieve in a single imaging frame.
  • a top view of the arrangement of Figure 8 is shown in Figure 10.
  • sensor 126 is formed from a portion of pixel array 42 ( Figure 2). In other words, a first portion of pixel array 42 is used to detect reference markings 122, while a second portion of pixel array 42 is used to capture images of sample 80 in sample holder 12.
  • Region 126' indicates the field of view of sensor 126 at the edge of image sensor 40 and region 40' indicates the field of view of the portion of image sensor 40' that is used to capture images of sample 80.
  • reference markings 122 may pass through field of view 126' of sensor 126 as sample holder 12 is moved in direction 88.
  • control circuitry 132 may issue a control signal to image sensor 40 to capture an imaging frame.
  • tick marks 122T will each successively pass over sensor 126, and chip-scale microscope 24 will capture a corresponding series of imaging frames as test chambers 22 move across imaging frame region 40' from edge 134 to edge 136.
  • each imaging frame region 40' may be separated from the rightmost edge of the adjacent imaging frame region by a distance D (i.e., a distance corresponding to the separation between tick marks 122T).
  • tick marks 122T may be spaced such that a region of overlap exists between adjacent imaging frames so that the images can be integrated as tiles to generate a larger field of view than chip-scale microscope 24 can achieve in a single imaging frame.
  • tick marks 122T are formed along a line that is parallel to the length of sample imaging portion 64 of sample holder 12 (i.e., along the x-axis as shown in Figure 9). This arrangement is merely illustrative. If desired, tick marks 122T may be formed along multiple axes such as both the x-axis and y-axis. Using a multiple-axis tracking system may be used to compensate for sample motion in multiple directions. Motion along each axis may be sensed independently by sensor 126 by using a unique color for each axial marking or by using other suitable identifying characteristic.
  • Figure 1 1 is a flow chart of illustrative steps involved in using a system such as system 10 of Figures 1 -10 in acquiring images of cells or other samples.
  • a sample may be injected into a sample chamber in a sample holder such as sample chamber 16 in sample holder 12.
  • the sample holder may be closed to automatically activate the sample distribution mechanism and thereby distribute portions of the sample from the sample chamber to respective test chambers in the sample holder.
  • the sample distribution mechanism may be controlled by flow control components such as flow control components 20 of Figure 3.
  • a user may insert the sample holder into an analysis module such as analysis module 14 of Figure 4 by inserting sample imaging portion 64 of sample holder 12 into opening 86 of analysis module 14.
  • sensor 126 may detect reference markings 122 as they pass through its field of view during insertion of sample holder 12 into analysis module 14.
  • a light source such as light source 124 may emit light towards sensor 126.
  • Sensor 126 may detect each tick mark 122T by detecting a change in received light as the tick mark passes through the light emitted by light source 124.
  • sensor 126 may detect each tick mark 122T by detecting a change in received light as the tick mark passes through the light emitted by illumination module 26.
  • trigger generator 130 may generate trigger signals for control circuitry 132 in response to sensor 126 detecting tick marks 122T.
  • control circuitry 132 may issue control signals to chip-scale microscope 24 to capture an imaging frame.
  • Multiple imaging frames may be captured of sample 80. Because imaging frame capture operations are triggered based on the detected reference markings, the imaging frames may have a uniform spatial distribution regardless of whether or not the user inserts sample holder 12 into analysis module 14 at a uniform speed. If desired, adjacent imaging frames may have some overlap with each other so that the imaging frames may be integrated as tiles to generate a large field of view image.
  • Sample imaging portion 64 of sample holder 12 may have built-in reference features for obtaining accurate color, opacity, and reflectivity measurements from a sample.
  • Figure 12 is a cross- sectional view of part of system 10 showing how sample holder 12 may have built-in reference features for obtaining accurate color, opacity, and reflectivity measurements from a sample such as sample 80.
  • sample holder 12 may have reference chambers such as reference chambers 222. There may be one, two, three, four, or more than four reference chambers 222 in sample holder 12. Each reference chamber 222 may be interposed between an associated pair of sample chambers 22, or reference chambers 222 may be grouped separately from sample chambers 22.
  • Each reference chamber 222 may include a reference surface such as reference surface 224.
  • Reference surface 224 may be a surface of sample holder 12 itself or may be a surface of an object or material in reference chamber 222.
  • Reference surface 224 may have known properties that may be compared with measurements obtained from sample 80 in test chambers 22 to determine corresponding properties of sample 80.
  • Reference surfaces 224 may, for example, have known properties such as known color, known transmissivity, known reflectivity, known absorbance, and/or other known properties. Reference surfaces 224 may be fully transparent or fully opaque, or may be configured to transmit light within a range of wavelengths. Reference surfaces 224 may include white surfaces, red surfaces, blue surfaces, green surfaces, or surfaces of other colors. Light sources in illumination module 26 (e.g., light sources 74 of Figure 4) may also have known properties and may be calibrated to increase the accuracy of colorimetric measurements, opacity measurements, and reflectivity measurements.
  • Image sensor 40 may gather reference information from reference surfaces 224 and may gather sample imaging data from sample 80. The reference information may be compared with the sample imaging data to determine information about sample 80. For example, image sensor 40 may compare the reference information with sample imaging data to determine the color of sample 80 based on the known information about reference surface 224 (e.g., based on the known color of reference surface 224). As another example, image sensor 40 may compare the reference information with sample imaging data to determine the transmissivity of sample 80 (e.g., based on the known transmissivity of reference surface 224).
  • one reference chamber 222 may contain an optically transparent reference surface 224 and another reference chamber 222 may contain a white reference surface 224.
  • Image sensor 40 may capture images of both the white and the transparent reference surfaces while the reference surfaces are illuminated by illumination module 26.
  • the two reference surfaces may be imaged in the same imaging frame or may be imaged in separate imaging frames.
  • Reference information gathered from the white reference surface and reference information gathered from the transparent reference surface may be compared with each other to determine the white balance of image sensor 40. Knowing the white balance of image sensor 40 may be useful in obtaining accurate color measurements from sample 80 in test chambers 22.
  • image sensor 40 may gather additional reference information from one or more additional reference surfaces 224.
  • the additional reference surfaces may have known colors.
  • the reference information gathered from colored reference surfaces 224 in reference chambers 222 may be compared with sample imaging data gathered from sample 80 in test chambers 22.
  • image sensor 40 may gather reference data from reference surfaces 224 and sample imaging data from sample 80 in the same imaging frame or in separate imaging frames. Based on the known white balance of image sensor 40 and based on the color variation between images of sample 80 and images of colored reference surfaces 224, the color of sample 80 may be accurately determined.
  • reference surfaces 224 need not exactly match the color of sample 80.
  • red and blue reference surfaces may be used to measure the color of a purple or violet sample.
  • Reference surfaces 224 may also be used in determining the opacity or transmissivity of a sample. For example, the intensity of light 94 received through reference surface 224 may be compared with the intensity of light 94 received through sample 80 and, based on the known transmissivity of reference surface 224 and based on the known intensity of light 94 emitted by illumination module 26, may be used to determine the opacity and/or transmissivity of sample 80.
  • illumination module 26 may be located on the same side of sample 80 as image sensor 40.
  • image sensor 40 may be configured to gather light that is reflected off of sample 80 and off of reference surfaces 224.
  • Figure 13 is a cross-sectional view of part of system 10 showing how sample holder 12 may have built-in reference features and showing how an illumination module may be located on the same side of sample holder 12 as image sensor 40.
  • illumination from illumination module 26' and detection by image sensor 40 occur on the same side of sample 80.
  • This type of configuration is sometimes referred to as epi- illumination and can be useful for fluorescence microscopy and for imaging opaque specimens.
  • Illumination module 26' may lie horizontally at a 90 degree angle to the optical axis of chip-scale microscope 24.
  • Light 94 emitted by illumination module 26' may strike optical member 226 and may be reflected upwards towards sample 80 in sample holder 12.
  • Optical member 226 may be a plane glass member that is partially reflective and partially transmissive.
  • Optical member 226 may, for example, be partially coated with a reflective material such silver paint. If desired, the transmissive portion of optical member 226 may be coated with an anti-reflection coating.
  • Optical member 226 may be tilted at a 45 degree angle to the path of light emitted by illumination module 26'.
  • Some or all of light 94 that strikes sample 80 may be reflected back towards image sensor 40 and may pass through the transmissive portion of optical member 226.
  • objective 38 may gather light 94 that is reflected off of sample 80 and focus the light onto image sensor 40.
  • reference surfaces 224 may have known properties (e.g., known color, known transmissivity, known reflectivity, known absorbance, and/or other known properties). Reference surfaces 224 may be fully transparent or fully opaque, or may be configured to transmit light within a range of wavelengths. Reference surfaces 224 may include white surfaces, red surfaces, blue surfaces, green surfaces, or surfaces of other colors. Light sources in illumination module 26' may also have known properties and may be calibrated to increase the accuracy of colorimetric measurements, opacity measurements, and reflectivity measurements.
  • Color may be measured by comparing the color of light reflected from one or more of reference surfaces 224 with that reflected from sample 80.
  • Opacity and reflectivity may be measured by comparing the intensity of light reflected from one or more of reference surfaces 224 with that reflected from sample 80.
  • Providing sample holder 12 with built-in reference features such as reference surfaces 224 may allow accurate color, opacity, and reflectivity measurements to be obtained from sample 80 using a single handheld diagnostic system such as system 10.
  • sample holder 12 may have built-in reference features such as built-in reference markings. The reference markings may be used to determine the number of pixels between reference points or across reference objects in sample holder 12.
  • Figure 14 is a cross-sectional top view of sample holder 12 showing how reference markings such as reference markings 228 may be formed on a surface of sample holder 12 (e.g., in the transparent portion of sample holder 12). There may be two, three, five, ten, less than ten, or more than ten reference markings on sample holder 12. Reference markings 228 may be spaced from each other at regular intervals and may have a known line spacing and line width.
  • reference markings 228 are separate from test chamber 22 and may therefore be imaged in a separate imaging frame from sample 80.
  • image sensor 40 may capture one or more images of reference markings 228 prior to or after capturing images of sample 80. The same level of magnification may be used during reference imaging of reference markings 228 as that used during sample imaging of sample 80.
  • Image sensor 40 may be configured to determine the number of pixels between reference markings 228 and analysis module 14 may use this information to determine the level magnification of chip-scale microscope 24 during that particular series of image capture operations. The level of magnification of chip-scale microscope 24 may in turn be used to determine the size of cells in sample 80 and/or to determine the concentration of cells in sample 80 (i.e., the number of cells within a given volume of sample 80).
  • reference markings may be incorporated into the test chamber in which the sample is imaged.
  • Figure 15 is a cross-sectional top view of sample holder 12 showing how reference features such as reference markings 228 may be located within test chamber 22 of sample holder 12. With this type of configuration, reference markings 228 may be imaged within the sample field of view. Reference markings 228 may overlap sample 80 while being located in a different plane of focus from sample 80 or may be minimally obscurant markings that are visible with sample 80.
  • Analysis module 14 may determine the level of magnification used during an image capture operation based on the number of pixels between reference markings 228.
  • sample holder 12 may include reference objects for determining the size of cells in sample 80 and/or for determining the level of magnification of chip-scale microscope 24.
  • Figure 16 is a cross-sectional top view of sample holder 12 illustrating how sample holder 12 may include reference features such as reference structure 230.
  • Reference structure 230 may be a sphere, cube, or other structure having a known size. There may be one, two, three, or more than three reference structures 230 in sample holder 12.
  • reference structures 230 are separate from test chamber 22 and may therefore be imaged in a separate imaging frame from sample 80.
  • image sensor 40 may capture one or more images of reference structures 230 prior to or after capturing images of sample 80. The same level of magnification may be used during reference imaging of reference structures 230 as that used during sample imaging of sample 80.
  • Image sensor 40 may be configured to determine the number of pixels across reference structures 230 and analysis module 14 may use this information to determine the level magnification of chip-scale microscope 24 during that particular series of image capture operations. This information may in turn be used to determine the size of cells in sample 80 and/or to determine the concentration of cells in sample 80 (i.e., the number of cells within a given volume of sample 80).
  • reference structures may be incorporated into the test chamber in which the sample is imaged.
  • Figure 17 is a cross-sectional top view of sample holder 12 showing how reference features such as reference structures 230 may be located within test chamber 22 of sample holder 12. With this type of configuration, reference structures 230 may be imaged within the sample field of view. Reference structures 230 may, for example, be located in a different plane of focus from sample 80 or may be minimally obscurant structures that are visible with sample 80.
  • Analysis module 14 may determine the level of magnification used during an image capture operation and may determine the size or concentration of cells in sample 80 based on the number of pixels across reference structures 230.
  • Figure 18 is a flow chart of illustrative steps involved in using a handheld diagnostic system with a chip-scale microscope and disposable sample holder having built in reference features such as reference surfaces 224 ( Figures 12 and 13), reference markings 228 ( Figures 14 and 15), and/or reference structures 230 ( Figures 16 and 17).
  • reference features such as reference surfaces 224 ( Figures 12 and 13), reference markings 228 ( Figures 14 and 15), and/or reference structures 230 ( Figures 16 and 17).
  • a sample may be injected into a sample chamber in a sample holder such as sample chamber 16 in sample holder 12. Portions of the sample may automatically be distributed from the sample chamber to respective test chambers in the sample holder.
  • a user may insert the sample holder into an analysis module such as analysis module 14 of Figure 4.
  • Circuitry in analysis module 14 may be configured to automatically trigger image capture operations at predetermined spatial or temporal intervals as sample holder 12 is inserted into analysis module 14.
  • built-in reference features in sample holder 12 may pass through the field of view of chip-scale microscope 24 in analysis module 14.
  • Sample 80 may also pass through the field of view of chip-scale microscope 24 during sample holder insertion.
  • image sensor 40 of chip- scale microscope 24 may capture images of the sample and of the built-in reference features (e.g., reference surfaces having known color, opacity, and reflectivity, reference markings having a known line spacing and line width, reference structures having a known size, and/or other reference features) as the sample holder is inserted into the analysis module.
  • the built-in reference features e.g., reference surfaces having known color, opacity, and reflectivity, reference markings having a known line spacing and line width, reference structures having a known size, and/or other reference features
  • Reference data e.g., images of reference features
  • Reference data may be gathered in the same imaging frame that sample data is gathered, or reference data may be gathered in a separate imaging frame from sample data (e.g., by capturing images of reference features prior to or after capturing images of the sample).
  • storage and processing circuitry in analysis module 14 may analyze images of the sample based on images of the built-in reference features. This may include, for example, determining a color, opacity, and/or reflectivity associated with the sample using reference surfaces 224 ( Figures 12 and 13), determining the size and/or concentration of cells or other substances in the sample using reference markings 228 ( Figures 14 and 15) or reference structures 230 ( Figures 16 and 17), and/or determining other parameter values based on images of reference features that are built into sample holder 12.
  • the handheld diagnostic system may include a disposable sample holder for collecting a sample, safely containing the sample, and for presenting the sample to an analysis module having a chip-scale microscope.
  • the sample holder may include fluid control components for automatically distributing portions of the sample to respective test chambers in the sample holder for imaging.
  • the test chambers may include markers (e.g., dyes, stains, fluorescence markers, etc.) configured to mark or otherwise identify specific nucleic acids or proteins in the sample if present in the sample.
  • the test chambers may be located in a transparent portion of the sample holder.
  • the analysis module may have a housing with an opening.
  • the opening may be configured to receive the transparent portion of the sample holder. While a user inserts the transparent portion of the sample holder into the opening of the analysis module, the chip-scale microscope may capture images of the sample in each test chamber as each test chamber passes through the field of view of the chip-scale microscope.
  • the analysis module may include an interchangeable illumination module for illuminating the sample and a chip-scale microscope for capturing images of the sample.
  • the chip-scale microscope may include an image sensor having an array of image pixels configured to gather pixel data from the sample.
  • the chip-scale microscope may also include optics such as one or more objective lenses for gathering light from the sample and focusing the light onto the image sensor.
  • the analysis module may include storage and processing circuitry for processing pixel data and, if desired, analyzing the processed pixel data to produce sample analysis information.
  • the pixel data and/or the sample analysis information may be transmitted to external computing equipment such as a portable electronic device for further analysis and/or for displaying sample analysis information for a user based on the sample images acquired using the chip-scale microscope.
  • the chip-scale microscope may be configured to capture spatially uniform imaging frames using an automated image capture mechanism.
  • the automated image capture mechanism may be based on a sensor that detects when the next imaging frame should be captured.
  • the sample holder may include a series of uniformly spaced markings. When a user inserts the sample holder into the analysis module, the series of uniformly spaced markings may be detected by a sensor in the analysis module. Upon detecting one of the markings, a control signal may be issued to capture an imaging frame using the chip-scale microscope.
  • This type of automated triggering ensures that the chip-scale microscope captures imaging frames at uniform spatial distribution even when the sample is moving and even when the sample holder is inserted manually into the analysis module at variable speed.
  • the sensor may be a photodiode that is separate from the image sensor in the chip-scale microscope or may formed from one or more image pixels at an edge of the image sensor in the chip-scale microscope.
  • the sample holder may have built-in reference features that are configured to be imaged by the chip-scale microscope.
  • the reference features may be imaged with the sample or may be imaged before or after the sample in the sample holder is imaged.
  • the reference features may include reference surfaces for determining the color, opacity, and reflectivity of a sample and/or the reference features may include reference markings or structures for determining the size and concentration of cells or other substances in the sample.

Abstract

Un système de diagnostic portable peut comporter un porte-échantillon jetable conçu pour recevoir et contenir un échantillon biologique, ainsi qu'un module d'analyse pourvu d'un microscope sur puce. Le porte-échantillon peut présenter une partie transparente qui est pourvue de chambres de test destinées à contenir des parties respectives d'un échantillon biologique et qui peut être insérée dans le module d'analyse. Le porte-échantillon peut comporter une pluralité de marques de pointage espacées régulièrement. Le module d'analyse peut être équipé d'un logement pourvu d'une ouverture conçue pour recevoir la partie transparente du porte-échantillon. Le module d'analyse peut en outre être équipé d'un capteur conçu pour détecter les marques de pointage à mesure que le porte-échantillon est inséré dans le module d'analyse. La partie transparente peut également présenter des caractéristiques de référence intégrées telles que des surfaces de référence, des marquages de référence et/ou des structures de référence. Le microscope sur puce peut être équipé d'un capteur d'images destiné à capturer des images de l'échantillon biologique et des caractéristiques de référence à mesure la partie transparente du porte-échantillon est insérée dans l'ouverture du module d'analyse. A chaque fois que le capteur détecte une marque de pointage, des circuits de commande peuvent transmettre un signal de commande au capteur d'images afin de capturer une image de l'échantillon biologique. Ce type de mécanisme de capture d'images automatisé garantit que les images sont capturées avec une répartition spatiale uniforme, même quand le porte-échantillon est inséré dans le module d'analyse à une vitesse variable. Le module d'analyse peut comprendre une source de lumière destinée à éclairer l'échantillon pendant les opérations de capture d'images, ainsi que des composants optiques destinés à collecter la lumière provenant de l'échantillon et à focaliser la lumière sur le capteur d'images. Des images des caractéristiques de référence peuvent être comparées à des images de l'échantillon. De plus, elles peuvent être utilisées pour déterminer la couleur, l'opacité, la réflectivité, la taille des cellules et/ou la concentration des cellules associées à l'échantillon. Le module d'analyse peut transmettre les informations d'imagerie de l'échantillon à un dispositif électronique portable qui peut, à son tour, afficher des informations d'analyse de l'échantillon correspondantes à l'attention d'un utilisateur.
PCT/US2014/049226 2013-08-05 2014-07-31 Système de diagnostic portable à porte-échantillon jetable et microscope sur puce WO2015020884A1 (fr)

Applications Claiming Priority (6)

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US13/959,304 2013-08-05
US13/959,304 US9347962B2 (en) 2013-08-05 2013-08-05 Handheld diagnostic system with chip-scale microscope and automated image capture mechanism
US13/959,122 2013-08-05
US13/959,491 2013-08-05
US13/959,122 US20150036131A1 (en) 2013-08-05 2013-08-05 Handheld diagnostic system with chip-scale microscope and disposable sample holder having built-in reference features
US13/959,491 US20150035966A1 (en) 2013-08-05 2013-08-05 Handheld diagnostic system with disposable sample holder and chip-scale microscope

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