WO2017132727A1 - Apparatus, method and system for detecting a chemical driving property using a fluorophore - Google Patents
Apparatus, method and system for detecting a chemical driving property using a fluorophore Download PDFInfo
- Publication number
- WO2017132727A1 WO2017132727A1 PCT/AU2017/050082 AU2017050082W WO2017132727A1 WO 2017132727 A1 WO2017132727 A1 WO 2017132727A1 AU 2017050082 W AU2017050082 W AU 2017050082W WO 2017132727 A1 WO2017132727 A1 WO 2017132727A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- fluorophore
- optical
- sample
- driving property
- property
- Prior art date
Links
- 238000000034 method Methods 0.000 title claims abstract description 65
- 239000000126 substance Substances 0.000 title claims abstract description 11
- 230000003287 optical effect Effects 0.000 claims abstract description 106
- 239000000523 sample Substances 0.000 claims description 88
- 239000011159 matrix material Substances 0.000 claims description 15
- 239000013307 optical fiber Substances 0.000 claims description 15
- 229920000642 polymer Polymers 0.000 claims description 12
- 238000000295 emission spectrum Methods 0.000 claims description 6
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 claims description 4
- MWUXSHHQAYIFBG-UHFFFAOYSA-N Nitric oxide Chemical compound O=[N] MWUXSHHQAYIFBG-UHFFFAOYSA-N 0.000 claims description 4
- 238000012544 monitoring process Methods 0.000 claims description 4
- 238000001228 spectrum Methods 0.000 claims description 4
- 229920002401 polyacrylamide Polymers 0.000 claims description 3
- 230000001681 protective effect Effects 0.000 claims description 3
- 238000005259 measurement Methods 0.000 description 20
- 206010028980 Neoplasm Diseases 0.000 description 19
- 239000000835 fiber Substances 0.000 description 18
- 230000005284 excitation Effects 0.000 description 15
- 238000001514 detection method Methods 0.000 description 13
- 201000011510 cancer Diseases 0.000 description 12
- 230000008569 process Effects 0.000 description 11
- 238000012545 processing Methods 0.000 description 7
- 238000001356 surgical procedure Methods 0.000 description 7
- 238000000576 coating method Methods 0.000 description 6
- 230000002378 acidificating effect Effects 0.000 description 5
- 239000011248 coating agent Substances 0.000 description 5
- 238000010586 diagram Methods 0.000 description 5
- 238000001139 pH measurement Methods 0.000 description 5
- 230000004044 response Effects 0.000 description 5
- 239000000243 solution Substances 0.000 description 5
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 4
- 230000004071 biological effect Effects 0.000 description 4
- 230000035945 sensitivity Effects 0.000 description 4
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 description 3
- 239000002253 acid Substances 0.000 description 3
- 239000000872 buffer Substances 0.000 description 3
- 230000008859 change Effects 0.000 description 3
- 239000000975 dye Substances 0.000 description 3
- 230000007613 environmental effect Effects 0.000 description 3
- 238000003384 imaging method Methods 0.000 description 3
- 238000004519 manufacturing process Methods 0.000 description 3
- 206010006187 Breast cancer Diseases 0.000 description 2
- 208000026310 Breast neoplasm Diseases 0.000 description 2
- 206010016654 Fibrosis Diseases 0.000 description 2
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- 239000007853 buffer solution Substances 0.000 description 2
- 230000008878 coupling Effects 0.000 description 2
- 238000010168 coupling process Methods 0.000 description 2
- 238000005859 coupling reaction Methods 0.000 description 2
- 238000012938 design process Methods 0.000 description 2
- 238000007598 dipping method Methods 0.000 description 2
- -1 e.g. Substances 0.000 description 2
- 230000004761 fibrosis Effects 0.000 description 2
- 150000002596 lactones Chemical group 0.000 description 2
- 230000007246 mechanism Effects 0.000 description 2
- 230000001338 necrotic effect Effects 0.000 description 2
- 230000007170 pathology Effects 0.000 description 2
- 229920002959 polymer blend Polymers 0.000 description 2
- DYPYMMHZGRPOCK-UHFFFAOYSA-N seminaphtharhodafluor Chemical compound O1C(=O)C2=CC=CC=C2C21C(C=CC=1C3=CC=C(O)C=1)=C3OC1=CC(N)=CC=C21 DYPYMMHZGRPOCK-UHFFFAOYSA-N 0.000 description 2
- 239000000377 silicon dioxide Substances 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- LWIHDJKSTIGBAC-UHFFFAOYSA-K tripotassium phosphate Chemical compound [K+].[K+].[K+].[O-]P([O-])([O-])=O LWIHDJKSTIGBAC-UHFFFAOYSA-K 0.000 description 2
- VXDCGBVAZDYTFK-UHFFFAOYSA-N (4-acetyloxy-2,3-dicyanophenyl) acetate Chemical compound CC(=O)OC1=CC=C(OC(C)=O)C(C#N)=C1C#N VXDCGBVAZDYTFK-UHFFFAOYSA-N 0.000 description 1
- VQVUBYASAICPFU-UHFFFAOYSA-N (6'-acetyloxy-2',7'-dichloro-3-oxospiro[2-benzofuran-1,9'-xanthene]-3'-yl) acetate Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC(Cl)=C(OC(C)=O)C=C1OC1=C2C=C(Cl)C(OC(=O)C)=C1 VQVUBYASAICPFU-UHFFFAOYSA-N 0.000 description 1
- XDLMVUHYZWKMMD-UHFFFAOYSA-N 3-trimethoxysilylpropyl 2-methylprop-2-enoate Chemical compound CO[Si](OC)(OC)CCCOC(=O)C(C)=C XDLMVUHYZWKMMD-UHFFFAOYSA-N 0.000 description 1
- BZTDTCNHAFUJOG-UHFFFAOYSA-N 6-carboxyfluorescein Chemical compound C12=CC=C(O)C=C2OC2=CC(O)=CC=C2C11OC(=O)C2=CC=C(C(=O)O)C=C21 BZTDTCNHAFUJOG-UHFFFAOYSA-N 0.000 description 1
- IHHSSHCBRVYGJX-UHFFFAOYSA-N 6-chloro-2-methoxyacridin-9-amine Chemical compound C1=C(Cl)C=CC2=C(N)C3=CC(OC)=CC=C3N=C21 IHHSSHCBRVYGJX-UHFFFAOYSA-N 0.000 description 1
- HRPVXLWXLXDGHG-UHFFFAOYSA-N Acrylamide Chemical compound NC(=O)C=C HRPVXLWXLXDGHG-UHFFFAOYSA-N 0.000 description 1
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 1
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 1
- 102000008186 Collagen Human genes 0.000 description 1
- 108010035532 Collagen Proteins 0.000 description 1
- 102000016942 Elastin Human genes 0.000 description 1
- 108010014258 Elastin Proteins 0.000 description 1
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 1
- FYYHWMGAXLPEAU-UHFFFAOYSA-N Magnesium Chemical compound [Mg] FYYHWMGAXLPEAU-UHFFFAOYSA-N 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 1
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 description 1
- OREJEGKBQBIJSJ-UHFFFAOYSA-N acridin-9-ylazanium;chloride;hydrate Chemical compound O.Cl.C1=CC=C2C(N)=C(C=CC=C3)C3=NC2=C1 OREJEGKBQBIJSJ-UHFFFAOYSA-N 0.000 description 1
- 239000004411 aluminium Substances 0.000 description 1
- 229910052782 aluminium Inorganic materials 0.000 description 1
- XAGFODPZIPBFFR-UHFFFAOYSA-N aluminium Chemical compound [Al] XAGFODPZIPBFFR-UHFFFAOYSA-N 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 238000001574 biopsy Methods 0.000 description 1
- 238000004061 bleaching Methods 0.000 description 1
- 239000011575 calcium Substances 0.000 description 1
- 229910052791 calcium Inorganic materials 0.000 description 1
- 238000011088 calibration curve Methods 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 229920001436 collagen Polymers 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 238000000151 deposition Methods 0.000 description 1
- 238000002059 diagnostic imaging Methods 0.000 description 1
- 238000009792 diffusion process Methods 0.000 description 1
- 229910001873 dinitrogen Inorganic materials 0.000 description 1
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 1
- 229910000396 dipotassium phosphate Inorganic materials 0.000 description 1
- 235000019797 dipotassium phosphate Nutrition 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 229920002549 elastin Polymers 0.000 description 1
- 238000001704 evaporation Methods 0.000 description 1
- 230000008020 evaporation Effects 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- GNBHRKFJIUUOQI-UHFFFAOYSA-N fluorescein Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 GNBHRKFJIUUOQI-UHFFFAOYSA-N 0.000 description 1
- 238000002189 fluorescence spectrum Methods 0.000 description 1
- 230000002496 gastric effect Effects 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 238000005286 illumination Methods 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 239000011777 magnesium Substances 0.000 description 1
- 229910052749 magnesium Inorganic materials 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 201000010893 malignant breast melanoma Diseases 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 201000001441 melanoma Diseases 0.000 description 1
- 229910021645 metal ion Inorganic materials 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- ZIUHHBKFKCYYJD-UHFFFAOYSA-N n,n'-methylenebisacrylamide Chemical compound C=CC(=O)NCNC(=O)C=C ZIUHHBKFKCYYJD-UHFFFAOYSA-N 0.000 description 1
- 230000001575 pathological effect Effects 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 230000002980 postoperative effect Effects 0.000 description 1
- 239000011591 potassium Substances 0.000 description 1
- 229910052700 potassium Inorganic materials 0.000 description 1
- 229910000160 potassium phosphate Inorganic materials 0.000 description 1
- 239000008057 potassium phosphate buffer Substances 0.000 description 1
- 235000011009 potassium phosphates Nutrition 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 230000035484 reaction time Effects 0.000 description 1
- 230000035440 response to pH Effects 0.000 description 1
- 238000002444 silanisation Methods 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 230000003319 supportive effect Effects 0.000 description 1
- 238000004441 surface measurement Methods 0.000 description 1
- 230000009897 systematic effect Effects 0.000 description 1
- 239000004753 textile Substances 0.000 description 1
- HRXKRNGNAMMEHJ-UHFFFAOYSA-K trisodium citrate Chemical compound [Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O HRXKRNGNAMMEHJ-UHFFFAOYSA-K 0.000 description 1
- 238000012800 visualization Methods 0.000 description 1
- 229910052725 zinc Inorganic materials 0.000 description 1
- 239000011701 zinc Substances 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/645—Specially adapted constructive features of fluorimeters
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01J—MEASUREMENT OF INTENSITY, VELOCITY, SPECTRAL CONTENT, POLARISATION, PHASE OR PULSE CHARACTERISTICS OF INFRARED, VISIBLE OR ULTRAVIOLET LIGHT; COLORIMETRY; RADIATION PYROMETRY
- G01J3/00—Spectrometry; Spectrophotometry; Monochromators; Measuring colours
- G01J3/02—Details
- G01J3/0205—Optical elements not provided otherwise, e.g. optical manifolds, diffusers, windows
- G01J3/0218—Optical elements not provided otherwise, e.g. optical manifolds, diffusers, windows using optical fibers
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01J—MEASUREMENT OF INTENSITY, VELOCITY, SPECTRAL CONTENT, POLARISATION, PHASE OR PULSE CHARACTERISTICS OF INFRARED, VISIBLE OR ULTRAVIOLET LIGHT; COLORIMETRY; RADIATION PYROMETRY
- G01J3/00—Spectrometry; Spectrophotometry; Monochromators; Measuring colours
- G01J3/28—Investigating the spectrum
- G01J3/30—Measuring the intensity of spectral lines directly on the spectrum itself
- G01J3/36—Investigating two or more bands of a spectrum by separate detectors
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01J—MEASUREMENT OF INTENSITY, VELOCITY, SPECTRAL CONTENT, POLARISATION, PHASE OR PULSE CHARACTERISTICS OF INFRARED, VISIBLE OR ULTRAVIOLET LIGHT; COLORIMETRY; RADIATION PYROMETRY
- G01J3/00—Spectrometry; Spectrophotometry; Monochromators; Measuring colours
- G01J3/28—Investigating the spectrum
- G01J3/44—Raman spectrometry; Scattering spectrometry ; Fluorescence spectrometry
- G01J3/4406—Fluorescence spectrometry
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/7703—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator using reagent-clad optical fibres or optical waveguides
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
- G01N21/80—Indicating pH value
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N2021/6417—Spectrofluorimetric devices
- G01N2021/6419—Excitation at two or more wavelengths
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N2021/6417—Spectrofluorimetric devices
- G01N2021/6421—Measuring at two or more wavelengths
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
- G01N2021/6434—Optrodes
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/645—Specially adapted constructive features of fluorimeters
- G01N2021/6484—Optical fibres
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/7703—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator using reagent-clad optical fibres or optical waveguides
- G01N2021/7706—Reagent provision
- G01N2021/772—Tip coated light guide
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/7703—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator using reagent-clad optical fibres or optical waveguides
- G01N2021/7706—Reagent provision
- G01N2021/773—Porous polymer jacket; Polymer matrix with indicator
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N2021/7769—Measurement method of reaction-produced change in sensor
- G01N2021/7786—Fluorescence
Definitions
- the present invention generally relates to apparatuses, methods and systems for optical sensing, e.g., a pH probe for acidity and alkalinity sensing in human tissue.
- Autofluorescence is the natural emission of light by compounds (including biological molecules) or substrates upon exposure to light, and typically competes with signals originating from artificially added fluorophores, e.g., in stains and dyes.
- Autofluorescence is nearly always present in regions of interest to medical imaging, e.g., collagen, elastin, and large proteins. Autofluorescent molecules can also be present in non-living materials, e.g., paper and textiles.
- fluorophores are applied to samples to enable sensing or visualization of specific properties, chemical concentrations and/or structures in the samples; however, optical excitation of the fluorophores—which is used to generate fluorescent optical signals representing the properties and/or structures of interest— causes autofluorescence in many samples, which presents as a background signal that can limit use or sensitivity of the signal from the applied or added fluorophores.
- Autofluorescence can be reduced using long-wavelength excitation sources, or pre-bleaching of samples, but is difficult to remove through electronic signal processing, and can vary significantly across a single sample, potentially introducing large errors if uniform background subtraction is applied.
- the present invention provides a method for optical sensing, including:
- the present invention also provides a system and apparatus for optical sensing, including an automated probe configured to perform the method above.
- the present invention also provides a system for optical sensing, the system including:
- a processor that generates electronic signals representing the measured driving property and/or a comparison of the measured driving property with a predetermined value based on electronic signals from the optical detectors.
- Figure 1 is a schematic diagram of a first embodiment of a system for optical sensing
- Figure 2 is a schematic diagram of a second embodiment of the system
- Figure 3 is a scanning electron microscope (SEM) image of a fluorosensor probe in the system
- Figure 4 is a diagram of a fluorophore in the system exposed to an acidic environment
- Figure 5 is a diagram of the fluorophore exposed to an alkaline environment
- Figure 6 is a graph of example emission spectra of the fluorophore in buffer environments with different pH values
- Figure 7 is a graph of example fluorescence ratios of the fluorophore for the different pH values, showing the retention of the fluorescence ratio signal after removal of the probe from the buffer;
- Figure 8 is a graph of example emission spectra of the fluorophore when in contact with a sample, and when removed from the sample;
- Figure 9 is a scatter plot of example normalised fluorescence ratios of the fluorophore when removed from tumourous or normal samples after exposure to their environmental pH.
- Figures 10A and 10B are schematic diagrams of an automated probe of the system.
- Described herein is an apparatus and method for optical sensing.
- the method includes the steps of: (1) exposing a fluorophore to a sample having a driving property (i.e., a detectable property, which is generally a chemical property, which may in turn relate to biological properties) to which the fluorophore is sensitive; (2) removing the fluorophore from the sample so that the fluorophore is not exposed to the property; and (3) detecting an optical signal from the removed fluorophore that represents the property of the sample.
- a driving property i.e., a detectable property, which is generally a chemical property, which may in turn relate to biological properties
- the removing step includes lifting the fluorophore upwards and/or away from the sample, and the detecting step includes determining a value of the property from the optical signal, i.e., measuring the property.
- the method may be described as a "lift- and-measure" technique.
- the optical signals include light with optical wavelengths, including from ultraviolet (UV) wavelengths to near-infrared (NIR) wavelengths.
- UV ultraviolet
- NIR near-infrared
- the chemical property can be acidity and alkalinity, and the value can be a pH value.
- the method can be a pH-sensing method, or a pH-detection method.
- the chemical property can be: (1) a concentration of hydrogen peroxide (H202); (2) an ion concentration, including of calcium, potassium, aluminium, sodium, magnesium, zinc, and/or other metal ions; or (3) a nitric oxide concentration.
- H202 hydrogen peroxide
- ion concentration including of calcium, potassium, aluminium, sodium, magnesium, zinc, and/or other metal ions
- the step of exposing the fluorophore and the sample to each other includes placing the fluorophore in contact with the sample, e.g., by manually placing the fluorophore in contact with the sample, including using a handheld housing of the apparatus.
- the fluorophore is thus in an environment of the sample that has the property.
- the step of exposing the fluorophore includes waiting for the fluorophore to respond to the property for a selected time period (an "equilibrium time") such that the fluorophore responds to a value of the driving property, e.g., a high-pH state or a low-pH state for an aqueous environment, corresponding to a structure of the fluorophore.
- Exposing the fluorophore includes allowing the fluorophore to equilibrate in sensitive proximity with the property of the sample, i.e., in the environment.
- the state (e.g., equilibrium state) of the fluorophore determines a signal value in the optical signal, thus the optical signal represents the state of the fluorophore, which in turn represents the value of the property.
- the method includes exposing the fluorophore for a preselected exposure time, between a minimum time and a maximum time, that is determined for the application based on the sensitivity of the particular tip and the driving property being detected.
- the exposure time is dependent on two parameters, and the values of these parameters can be pre-determined for each application: a diffusion speed of the driving property (which can be a species of interest) to the fluorophores (which can be through a matrix around the fluorophores); and a reaction time for the fluorophore to change its optical response after expose to the driving property commences.
- a diffusion speed of the driving property which can be a species of interest
- the fluorophores which can be through a matrix around the fluorophores
- a reaction time for the fluorophore to change its optical response after expose to the driving property commences For an example pH probe, the exposure time is between 1 second and 10 seconds, depending on a thickness of a layer containing the fluorophores.
- the method includes a step of illuminating the fluorophore with an optical beam from a light source, which can be a laser or light-emitting diode, LED.
- the method includes a step of determining and monitoring one or more portions of an emission spectrum, from the removed fluorophore in the optical signal, using an optical detector.
- the light source and the optical detector are remote from the fluorophore: the optical beam (i.e., the illumination for the fluorophore) and the optical signals (from the fluorophore) travel along one or more optical guides in the form of optical fibres, e.g., the same optical fibre, with the optical signals being directed to the optical detector by a splitter.
- the optical guides can be in the form of ⁇ - ⁇ or 200- ⁇ optical fibre patch cables.
- the optical signals can be detected by a fluorescence microscope.
- the fluorophore is mounted on or embedded in a sensor surface to form a fluorosensor of the apparatus.
- the fluorophore includes a population of the fluorescent molecules exposed at the sensing surface of the fluorosensor.
- the sensor surface is an end (or a "tip") of an optical guide of the apparatus that carries the optical signal.
- the optical guide can be provided by an optical fibre.
- the handheld housing of the apparatus mechanically communicates with the fluorosensor to allow manual control of the location or placement of the fluorosensor.
- the handheld housing can be the optical guide itself (e.g., by the optical fibre) and/or alternative or additional support members (e.g., including a supportive coating or tube around the optical guide).
- the optical guide with the fluorophore is thus a probe, e.g., a pH probe or a fibre-tip probe.
- the fibre probe is flexible and robust, and has a small measurement area corresponding to the area of the optical fibre tip, e.g., a ⁇ - ⁇ or 200- ⁇ diameter fibre.
- the fluorophore is mounted in a matrix, which could be a polymer.
- the fluorophore in the matrix can be applied as a coating to the sensor surface.
- the method includes depositing the fluorophore-doped matrix on the sensor surface.
- the matrix can be an acrylamide polymer on the optical fibre tip.
- the pH probe can measure pH rapidly, e.g., in less than one minute, can be simple to use, and can avoid leaving a residue or stain that would affect later pathology testing.
- the fluorescent molecules may be chemically attached to the sensor surface, or embedded into it using silanisation.
- the sample can be a human tissue sample, including a cancerous or noncancerous (i.e., "regular") tissue sample.
- the method can be a cancer-detection method.
- the method includes a step of estimating whether the sample is cancerous or noncancerous based on the driving property (including the pH).
- the step of estimating whether the sample is cancerous or non-cancerous includes comparing the determined value (of the driving property) with one or more predetermined values corresponding to previously-determined cancerous and non-cancerous samples (i.e., respective different sample types).
- the previously-determined samples are for the same tissue type as the sample exposed to the fluorophore.
- the method can be used as an aid for margin detection during cancer surgery, including by measuring the tissue pH, and comparing with the values from regular tissue, quickly and accurately.
- Cancer detection at the margins during surgery allows the removed tissue sections to be enlarged if required during the surgery, thus reducing the likelihood of subsequent surgery.
- Extracellular pH in the vicinity of cancer cells is generally lower than extracellular pH in the vicinity of normal cells of the same tissue type in the same patient.
- the described method allows for pH detection in smaller areas (e.g., as small as the optical fibre tip) than existing
- electrochemical surface pH probes which typically have a tip diameter in the order of 10 mm.
- the described method may also be used for non-invasive pH detection in embryology, dentistry, gastrointestinal investigations, water-quality monitoring (e.g., in aquaria), and food-quality monitoring (e.g., of relatively dry surfaces)
- the fluorophore can be 5,6-carboxynapthofluorescein (CNF), which changes colour with a change in the environmental pH.
- CNF 5,6-carboxynapthofluorescein
- fluorophores for the pH sensing can include any one or more of the following bases (and their derivatives):
- the fluorophores can have a plurality of emission bands, including two, allowing ratiometric detection to alleviate potential issues from photobleaching or coupling, and allowing for measurements to be performed over a long period without the introduction of systematic errors.
- the ratiometric detection allows for each driving property value to be generated with little dependence on the excitation power or fluorophore density. If the excitation intensity is increased, the emission bands will increase proportionally, removing the intensity dependence that can restrict precision of typical fluorophore measurements.
- the ratiometric detection also minimises variations in signal response between different example fluorosensors (or "probes"), simplifying fabrication requirements of large numbers of fluorosensors.
- the method includes generating calibration relationships between pre-prepared samples with pre-selected environmental property values or levels, and respective detected optical signal levels, for each fluorosensor in order to compensate for different tip sensitivities, e.g., doping concentration, polymer thickness, etc..
- the calibration relationships are generated using phosphate buffers with different pH values.
- Described herein is a system for performing the method, the system including:
- a processor including processing circuit, and a program of machine- readable instructions executed by the processing circuit to generate, record and display electronic signals representing the measured driving property, and/or a comparison of the measured driving property with a predetermined value (e.g., an indication of tissue type) based on electronic signals from the optical detectors.
- a predetermined value e.g., an indication of tissue type
- the optical detectors can be provided by a spectrum analyser and/or photodiodes.
- the processing circuit can include a microprocessor for processing the electronic signals from the optical detector.
- the system can have an outer protective housing configured for use in an operating theatre, and all of the operational components can be configured and sized to fit within the protective housing, e.g., on a trolley in a hospital operating theatre.
- a first system 100 for photopolymerisation of the matrix onto a fiber tip, and for pH measurement applications could include:
- a fixing (or "photopolymerising") light source for generating a fixing light beam e.g., a 405-nm or 372-nm laser
- one or more excitation light source/s for generating an excitation light beam e.g., a blue 473 nm laser for a CNF flurorphore
- one or more optical detectors which can include a spectrometer 302 (e.g., for detecting emission of the CNF at two peaks, at
- one or more optical guide/s in the form of silica multi-mode fibres e.g., 100 ⁇ or 200 ⁇ optical fibre patch cables
- silica multi-mode fibres e.g., 100 ⁇ or 200 ⁇ optical fibre patch cables
- the optical components include mirrors to direct the light beams, a dichroic filter to combine the fixing and excitation light beams, a long pass filter to separate the excitation and fluorescent light beams, and microscope objectives to couple into and out of the fibres.
- a second system 200 for measurements using a coated tip includes:
- an excitation light source 308 in the form of an LED (i) an excitation light source 308 in the form of an LED; (ii) the fluorosensor 300 with a dual-emission fluorophore for ratiometric detection;
- optical detectors in the form of two photodetectors (PDs) Dl, D2 or a
- a dichroic filter 304 configured to separate the two emission bands to the optical detectors
- optical guides in the form of a bifurcated fibre 306 fibre splitter to connect the LED, fluorosensor 300 and detector.
- the fluorosensor 300 includes a polymer coating on a cleaved end of an optical fibre.
- the fibre is cleaved to expose a fresh tip surface for polymer attachment.
- the first system 100 and/or the second system 200 can include an apparatus in the form of a timed automated probe 110 that performs the method for sensing, described hereinafter containing the fibre and the fluorophore, which can be referred to as a "timed probe” or "automated probe”.
- the timed probe 110 is controlled by computer-readable instructions to place the fluorosensor 300 in contact with the sample for a pre-determined exposure time (predetermined for the type of fluorosensor 300 and the application), and to remove (by lifting) fluorosensor 300 from the sample.
- the first system 100 and second system 200 can include a controller that follows the computer-readable instructions to operate the timed probe 110, and to perform the optical measurements once the fluorosensor 300 is removed from the sample.
- the timed probe 110 can measure a force of the fluorosensor 300 touching the sample, and can control this force so that the same force is applied for each exposure, thus potentially improving consistency between measurements.
- the timed probe 110 can consist of a fiber probe 101 mounted in a handheld rugged outer housing 105.
- the timed probe 110 includes an aperture 103 that is placed in contact with an area to be measured by the operator.
- a motorized mechanism 102 (including a motor and a fibre mount) is controlled (by the controller) to move the fluorosensor 300 from an initial retracted position 104 (retracted inside the outer housing 105) to an exposed position 106 (in contact with the tissue surface) for a preselected fixed amount of time.
- the motorized mechanism 102 is then controlled (by the controller) to retract the fluorosensor 300 from the surface back to the retracted position 104.
- the optical components are then controlled (by the controller) to perform the optical measurements for the preselected measurement duration. Once the measurements are complete, the first system 100 and second system 200 then alert the operator that the fluorosensor 300 is ready for the next measurement to be performed.
- the method includes:
- the design process for the fluorosensor 300 for a pre-defined application (which can be cancer detection in human tissue cells) includes:
- fluorophore is to be doped; selecting the light source/s (which can be LEDs or lasers) and the optical detector/s (which can be photodiodes or CCD spectrometers) to correspond to the optical excitation band/s (which can include 473 nm) and the optical emission band/s (which can be 565 nm and 705 nm) of the fluorophore; selecting the optical guide/s (which can be optical fibres) to carry the optical excitation light from the light source/s to the fluorophore, and the fluorescent light from the fluorophore to the optical detector/s; selecting a concentration and a volume (which includes the coating thickness and the area of the doped matrix in the fluorosensor 300) for the fluorophore in the fluorosensor 300 based on a pre-determined desired signal intensity, and sensitivity to the driving property (e.g., determined by testing a range of values); determining an optimal exposure time for the fluorophore to be exposed to the sample to allow
- a doped polymer mixture solution at the selected concentration (e.g., for CNF in cellular cancer detection, mixing by weight, 27% acrylamide, 3% bis-acrylamide, 70% pH 6*5 potassium phosphate buffer, 0*4 mg/mL CNF and 40 ⁇ 7 ⁇ _, triethylamine);
- the sensing process broadly includes exciting the dye with a light source, and collecting and analysing the emitted fluorescence light from the dye.
- An example of this process with CNF for pH sensing could include:
- the optical source exciting the fluorophore e.g., the 473 nm laser exciting the CNF fluorophore
- the sensor surface receiving at least a portion of the fluorescent emission, coupling the fluorescent emission into the optical guide (the fluorescent emission travels in a back-propagating mode in the optical guide/s), which directs the fluorescent emission to the optical detector/s;
- wavelength band to resist excess excitation light reaching the detector/s e.g., using a 473 nm long-pass filter
- the optical detector/s detecting the emission from the fluorophore (which can include the plurality of bands, e.g., at 565 nm and 705 nm for CNF);
- driving property from the detected emission e.g., a value based on an intensity ratio of the 565 nm and 705 nm peaks for CNF, e.g., including integrating the detected signal intensity under the two peaks— from 500-635 nm and from 635-900 nm— and dividing the area of the first peak by the area of the second peak to give the fluorescence ratio for that measurement).
- the calibration process includes:
- the operational process for an application of measuring surface pH in tissue samples includes:
- the computer processor comparing the estimated value to the predetermined thresholds (e.g., the pre-determined pH thresholds for cancerous versus non-cancerous cells), and generating an alert or display (e.g., alerting a surgeon to a likelihood of cancer in each sensing location); and
- the predetermined thresholds e.g., the pre-determined pH thresholds for cancerous versus non-cancerous cells
- the fluorosensor 300 (referred to as a "probe") was prepared using CNF on an optical fibre.
- the probe response to pH was recorded with a series of PBS buffer solutions.
- the horizontal axis is wavelength (in nm) and the vertical axis is intensity (in counts)
- the ratiometric response of the probe in air, after dipping into the buffer solutions and removal increased
- the structures of the fluorophore CNF both after and before reaction with hydrogen ions are shown in Figures 4 and 5 respectively.
- the fluorophore When the fluorophore is exposed to the sample, the fluorophore equilibrates to one of the structures, and remains in this form (e.g., the protonated form or lactone form for CNF) ever after removal, for the determined maximum removed time.
- the fluorosensor 300 includes a population of many fluorophore molecules, and the fluorescent emission in each of the bands represents which fraction of the molecules on average is in the state corresponding to that band.
- the fluorosensor 300 (referred to as a "probe") was prepared using CNF on an optical fibre. Sheep tissue samples were spiked with 1M hydrochloric acid or 1M sodium hydroxide to obtain acidic or basic samples respectively. Emission spectra were recorded with the probe both touching and lifted from the tissue surface. As shown in Figure 8, where the vertical axis is intensity (CPS) and the horizontal axis is wavelength (nm), the autofluorescence background can form a significant fraction of the total signal strength and can form a significant contribution to the apparent measured ratio for the emission spectra of the acidic samples and the basic samples when the probe was touching the samples. In contrast, the autofluoresence was substantially reduced for the removed acidic spectrum and removed basic spectrum.
- CPS intensity
- nm wavelength
- the fluorosensor 300 (or "probe") was prepared using CNF on an optical fibre. Surgically excised human breast cancer and melanoma tissue samples were measured with the optical fibre pH probe at numerous locations over the sample surface, with the tissue type of each location confirmed later by histopathology. Tissue surface measurements were obtained from four melanoma and four breast cancer samples. Results were normalised to the mean value of the normal tissue results to simplify comparison between samples. Statistical data were plotted for normal and tumour datasets, showing the mean, median, and standard deviation. The mean and standard deviation values are shown in Table 1 hereinafter, along with descriptions of the specimen and tumour type for each of the measured samples.
- the tumour samples were generally more acidic than the normal tissue samples by a statistically significant difference in detected pH.
- the necrotic tumour samples displayed a similar pH to regular cancer samples, while fibrosis samples showed a similar pH to normal tissue.
- the tumour (grey) and normal (black) measurement datasets for each of the eight tissue samples are shown by individual location measurements (circles). Individual measurements of fibrosis (black triangle), necrotic tumour (grey triangle) and those near normal-tumour margins (square) are also plotted, without any corresponding statistical features, for comparison.
- Cancerous tissue could be differentiated from normal tissue in the fresh human tissue biopsies by measurement of the tissue pH using the optical probe, allowing for measurements to be performed rapidly with high spatial resolution (e.g., 200 ⁇ in diameter, equivalent to an area in the order of five to ten cells wide, with the potential to reduce this to the measurement of single cells by reducing the size of the fibre probe).
- high spatial resolution e.g. 200 ⁇ in diameter, equivalent to an area in the order of five to ten cells wide, with the potential to reduce this to the measurement of single cells by reducing the size of the fibre probe.
- a multi-core imaging fibre with a plurality of fluorosensors on the ends of the respective cores, can be used to increase sample throughput by measuring the tissue pH of a respective plurality of sensor locations simultaneously (i.e., in parallel) with a corresponding plurality of respective optical detectors (i.e., parallel detectors), which may be a charge-coupled device (CCD) or camera.
- CCD charge-coupled device
- Measurement time can be controlled by varying the thickness of the fluorophore layer (e.g., the polymer layer). Using a thinner layer gives a faster response; however the signal intensity is reduced.
- the fluorophore layer e.g., the polymer layer
Abstract
A method for optical sensing, the method including: exposing a fluorophore to a sample having a chemical driving property to which the fluorophore is sensitive; removing the fluorophore from the sample so that the fluorophore is not exposed to the driving property; and detecting an optical signal from the removed fluorophore that represents the driving property.
Description
APPARATUS, METHOD AND SYSTEM FOR DETECTING A CHEMICAL DRIVING PROPERTY USING A FLUOROPHORE
TECHNICAL FIELD
[001] The present invention generally relates to apparatuses, methods and systems for optical sensing, e.g., a pH probe for acidity and alkalinity sensing in human tissue.
BACKGROUND
[002] Autofluorescence is the natural emission of light by compounds (including biological molecules) or substrates upon exposure to light, and typically competes with signals originating from artificially added fluorophores, e.g., in stains and dyes.
Autofluorescence is nearly always present in regions of interest to medical imaging, e.g., collagen, elastin, and large proteins. Autofluorescent molecules can also be present in non-living materials, e.g., paper and textiles.
[003] Autofluorescence is problematic in many optical sensing applications, including making optical measurements of chemical or biological properties, and biological properties of samples (e.g., tissue samples), and/or imaging samples based on these chemical properties and biological properties.
[004] In many optical sensing and imaging applications, fluorophores are applied to samples to enable sensing or visualization of specific properties, chemical concentrations and/or structures in the samples; however, optical excitation of the fluorophores— which is used to generate fluorescent optical signals representing the properties and/or structures of interest— causes autofluorescence in many samples, which presents as a background signal that can limit use or sensitivity of the signal from the applied or added fluorophores.
[005] Autofluorescence can be reduced using long-wavelength excitation sources, or pre-bleaching of samples, but is difficult to remove through electronic signal processing,
and can vary significantly across a single sample, potentially introducing large errors if uniform background subtraction is applied.
[006] Accordingly, autofluorescence remains a limitation to broader applications of optical sensing, including for medical sensing of tissue samples.
[007] In cancer surgery, margins of removed tissue sections are assessed
pathologically to determine whether the entire cancer has been removed, and
approximately 15-20% of margins may show incomplete removal, thus requiring subsequent cancer surgery to remove the remaining cancer. Existing pathological methods used to determine tissue type during surgery can compromise post-operative pathology, have a lag of minutes to hours before the surgeon receives the results of the tissue analysis, and are restricted to excised tissue.
[008] It is desired to address or ameliorate one or more disadvantages or limitations associated with the prior art, or to at least provide a useful alternative.
SUMMARY
[009] The present invention provides a method for optical sensing, including:
(i) exposing a fluorophore to a sample having a chemical driving property to which the fluorophore is sensitive.
(ii) removing the fluorophore from the sample so that the fluorophore is not exposed to the driving property; and
(iii) detecting an optical signal from the removed fluorophore that represents the driving property.
[010] The present invention also provides a system and apparatus for optical sensing, including an automated probe configured to perform the method above.
[011] The present invention also provides a system for optical sensing, the system including:
(i) one or more fluorosensors;
(ii) one or more optical sources to cause the fluorosensors to fluoresce;
(iii) one or more optical detectors to detect the fluorescence; and
(iv) a processor that generates electronic signals representing the measured driving property and/or a comparison of the measured driving property with a predetermined value based on electronic signals from the optical detectors.
BRIEF DESCRIPTION OF THE DRAWINGS
[012] Some embodiments of the present invention are described hereinafter, by way of non-limiting example only, with reference to the accompanying drawings, in which:
[013] Figure 1 is a schematic diagram of a first embodiment of a system for optical sensing;
[014] Figure 2 is a schematic diagram of a second embodiment of the system;
[015] Figure 3 is a scanning electron microscope (SEM) image of a fluorosensor probe in the system;
[016] Figure 4 is a diagram of a fluorophore in the system exposed to an acidic environment;
[017] Figure 5 is a diagram of the fluorophore exposed to an alkaline environment;
[018] Figure 6 is a graph of example emission spectra of the fluorophore in buffer environments with different pH values;
[019] Figure 7 is a graph of example fluorescence ratios of the fluorophore for the different pH values, showing the retention of the fluorescence ratio signal after removal of the probe from the buffer;
[020] Figure 8 is a graph of example emission spectra of the fluorophore when in contact with a sample, and when removed from the sample;
[021] Figure 9 is a scatter plot of example normalised fluorescence ratios of the fluorophore when removed from tumourous or normal samples after exposure to their environmental pH; and
[022] Figures 10A and 10B are schematic diagrams of an automated probe of the system.
DETAILED DESCRIPTION
Overview
[023] Described herein is an apparatus and method for optical sensing. The method includes the steps of: (1) exposing a fluorophore to a sample having a driving property (i.e., a detectable property, which is generally a chemical property, which may in turn relate to biological properties) to which the fluorophore is sensitive; (2) removing the fluorophore from the sample so that the fluorophore is not exposed to the property; and (3) detecting an optical signal from the removed fluorophore that represents the property of the sample.
[024] Removing the fluorophore from the sample before detecting the optical signal mitigates the effects of autofluorescence from the sample, and thus the optical signal from the removed fluorophore can be detected with a better signal-to-noise ratio (SNR) than a non-removed optical signal from a non-removed fluorophore.
[025] The removing step includes lifting the fluorophore upwards and/or away from the sample, and the detecting step includes determining a value of the property from the
optical signal, i.e., measuring the property. Thus, the method may be described as a "lift- and-measure" technique.
[026] The optical signals include light with optical wavelengths, including from ultraviolet (UV) wavelengths to near-infrared (NIR) wavelengths.
[027] The chemical property can be acidity and alkalinity, and the value can be a pH value. Thus, the method can be a pH-sensing method, or a pH-detection method.
Alternatively, the chemical property can be: (1) a concentration of hydrogen peroxide (H202); (2) an ion concentration, including of calcium, potassium, aluminium, sodium, magnesium, zinc, and/or other metal ions; or (3) a nitric oxide concentration.
[028] The step of exposing the fluorophore and the sample to each other includes placing the fluorophore in contact with the sample, e.g., by manually placing the fluorophore in contact with the sample, including using a handheld housing of the apparatus. The fluorophore is thus in an environment of the sample that has the property. The step of exposing the fluorophore includes waiting for the fluorophore to respond to the property for a selected time period (an "equilibrium time") such that the fluorophore responds to a value of the driving property, e.g., a high-pH state or a low-pH state for an aqueous environment, corresponding to a structure of the fluorophore. This state can be described as an equilibrium state. Exposing the fluorophore includes allowing the fluorophore to equilibrate in sensitive proximity with the property of the sample, i.e., in the environment. The state (e.g., equilibrium state) of the fluorophore determines a signal value in the optical signal, thus the optical signal represents the state of the fluorophore, which in turn represents the value of the property. The method includes exposing the fluorophore for a preselected exposure time, between a minimum time and a maximum time, that is determined for the application based on the sensitivity of the particular tip and the driving property being detected. The exposure time is dependent on two parameters, and the values of these parameters can be pre-determined for each application: a diffusion speed of the driving property (which can be a species of interest) to the fluorophores (which can be through a matrix around the fluorophores); and a reaction time for the fluorophore to change its optical response after expose to the driving property commences.
For an example pH probe, the exposure time is between 1 second and 10 seconds, depending on a thickness of a layer containing the fluorophores.
[029] The method includes a step of illuminating the fluorophore with an optical beam from a light source, which can be a laser or light-emitting diode, LED. The method includes a step of determining and monitoring one or more portions of an emission spectrum, from the removed fluorophore in the optical signal, using an optical detector. The light source and the optical detector are remote from the fluorophore: the optical beam (i.e., the illumination for the fluorophore) and the optical signals (from the fluorophore) travel along one or more optical guides in the form of optical fibres, e.g., the same optical fibre, with the optical signals being directed to the optical detector by a splitter. The optical guides can be in the form of ΙΟΟ-μιη or 200-μιη optical fibre patch cables.
Alternatively, the optical signals can be detected by a fluorescence microscope.
[030] The fluorophore is mounted on or embedded in a sensor surface to form a fluorosensor of the apparatus. The fluorophore includes a population of the fluorescent molecules exposed at the sensing surface of the fluorosensor. The sensor surface is an end (or a "tip") of an optical guide of the apparatus that carries the optical signal. The optical guide can be provided by an optical fibre. The handheld housing of the apparatus mechanically communicates with the fluorosensor to allow manual control of the location or placement of the fluorosensor. The handheld housing can be the optical guide itself (e.g., by the optical fibre) and/or alternative or additional support members (e.g., including a supportive coating or tube around the optical guide). The optical guide with the fluorophore is thus a probe, e.g., a pH probe or a fibre-tip probe. The fibre probe is flexible and robust, and has a small measurement area corresponding to the area of the optical fibre tip, e.g., a ΙΟΟ-μιη or 200-μιη diameter fibre. The fluorophore is mounted in a matrix, which could be a polymer. The fluorophore in the matrix can be applied as a coating to the sensor surface. The method includes depositing the fluorophore-doped matrix on the sensor surface. The matrix can be an acrylamide polymer on the optical fibre tip. The pH probe can measure pH rapidly, e.g., in less than one minute, can be simple to use, and can avoid leaving a residue or stain that would affect later pathology testing. The
fluorescent molecules may be chemically attached to the sensor surface, or embedded into it using silanisation.
[031] The sample can be a human tissue sample, including a cancerous or noncancerous (i.e., "regular") tissue sample. Thus, the method can be a cancer-detection method. The method includes a step of estimating whether the sample is cancerous or noncancerous based on the driving property (including the pH). The step of estimating whether the sample is cancerous or non-cancerous includes comparing the determined value (of the driving property) with one or more predetermined values corresponding to previously-determined cancerous and non-cancerous samples (i.e., respective different sample types). The previously-determined samples are for the same tissue type as the sample exposed to the fluorophore. Thus the method can be used as an aid for margin detection during cancer surgery, including by measuring the tissue pH, and comparing with the values from regular tissue, quickly and accurately. Cancer detection at the margins during surgery allows the removed tissue sections to be enlarged if required during the surgery, thus reducing the likelihood of subsequent surgery. Extracellular pH in the vicinity of cancer cells is generally lower than extracellular pH in the vicinity of normal cells of the same tissue type in the same patient. The described method allows for pH detection in smaller areas (e.g., as small as the optical fibre tip) than existing
electrochemical surface pH probes (which typically have a tip diameter in the order of 10 mm).
[032] The described method may also be used for non-invasive pH detection in embryology, dentistry, gastrointestinal investigations, water-quality monitoring (e.g., in aquaria), and food-quality monitoring (e.g., of relatively dry surfaces)
[033] The fluorophore can be 5,6-carboxynapthofluorescein (CNF), which changes colour with a change in the environmental pH.
[034] Other fluorophores for the pH sensing can include any one or more of the following bases (and their derivatives):
(i) 3,6-Diacetoxyphthalonitrile;
(ii) 8-Hydroxypyrene-l,3,6-Trisulfonic Acid;
(iii) 9-Amino-6-Chloro-2-Methoxyacridine;
(iv) 9-Aminoacridine hydrochloride monohydrate 98%;
(v) Carboxyfluorescein;
(vi) Carboxynaphthofluorescein;
(vii) 7'-Dichlorofluorescin diacetate (DCFDA);
(viii) Fluorescein;
(ix) 8-Hydroxypyrene-l,3,6-trisulfonic acid trisodium salt (HPTS);
(x) LysoSensor Yellow/Blue/Green;
(xi) 2',7'-Bis-(2-Carboxyethyl)-5-(and-6)-Carboxyfluorescein) (BCECF);
(xii) Seminaphthofluorescein (SNAFL); and
(xiii) Seminaphtharhodafluor (SNARF).
[035] The fluorophores can have a plurality of emission bands, including two, allowing ratiometric detection to alleviate potential issues from photobleaching or coupling, and allowing for measurements to be performed over a long period without the introduction of systematic errors. The ratiometric detection allows for each driving property value to be generated with little dependence on the excitation power or fluorophore density. If the excitation intensity is increased, the emission bands will increase proportionally, removing the intensity dependence that can restrict precision of typical fluorophore measurements. The ratiometric detection also minimises variations in signal response between different example fluorosensors (or "probes"), simplifying fabrication requirements of large numbers of fluorosensors.
[036] The method includes generating calibration relationships between pre-prepared samples with pre-selected environmental property values or levels, and respective detected
optical signal levels, for each fluorosensor in order to compensate for different tip sensitivities, e.g., doping concentration, polymer thickness, etc.. The calibration relationships are generated using phosphate buffers with different pH values.
[037] Described herein is a system for performing the method, the system including:
(i) one or more fluorosensors;
(ii) one or more optical sources to cause the fluorosensors to fluoresce;
(iii) one or more optical detector to detect the fluorescence; and
(iv) a processor (including processing circuit, and a program of machine- readable instructions executed by the processing circuit) to generate, record and display electronic signals representing the measured driving property, and/or a comparison of the measured driving property with a predetermined value (e.g., an indication of tissue type) based on electronic signals from the optical detectors.
[038] The optical detectors can be provided by a spectrum analyser and/or photodiodes. The processing circuit can include a microprocessor for processing the electronic signals from the optical detector. The system can have an outer protective housing configured for use in an operating theatre, and all of the operational components can be configured and sized to fit within the protective housing, e.g., on a trolley in a hospital operating theatre.
System 100, 200
[039] As shown in Figure 1, a first system 100 for photopolymerisation of the matrix onto a fiber tip, and for pH measurement applications could include:
(i) a fixing (or "photopolymerising") light source for generating a fixing light beam (e.g., a 405-nm or 372-nm laser);
(ii) one or more excitation light source/s for generating an excitation light beam (e.g., a blue 473 nm laser for a CNF flurorphore);
(iii) one or more optical detectors, which can include a spectrometer 302 (e.g., for detecting emission of the CNF at two peaks, at
approximately 565 nm and 705 nm);
(iv) a fluorosensor 300;
(v) one or more optical guide/s in the form of silica multi-mode fibres (e.g., 100 μηι or 200 μιη optical fibre patch cables) for guiding the excitation light to the fluorosensor 300, and for guiding at least a portion of the emitted fluorescent light back from the fluorosensor 300;
(vi) one or more optical guide/s in the form of the silica multi-mode fibres for guiding the fluorescent light to the optical detector/s;
(vii) one or more handheld housings in the form of the optical fibres with graspable coatings, and
(viii) free-space optical components for guiding the fixing light and the excitation light into optical guides, and for guiding the fluorescent light from the fluorosensor 300 to the optical detector/s.
[040] The optical components include mirrors to direct the light beams, a dichroic filter to combine the fixing and excitation light beams, a long pass filter to separate the excitation and fluorescent light beams, and microscope objectives to couple into and out of the fibres.
[041] As shown in Figure 2, a second system 200 for measurements using a coated tip includes:
(i) an excitation light source 308 in the form of an LED;
(ii) the fluorosensor 300 with a dual-emission fluorophore for ratiometric detection;
(iii) optical detectors in the form of two photodetectors (PDs) Dl, D2 or a
spectrometer for the two respective different emission bands of the fluorophore;
(iv) a dichroic filter 304 configured to separate the two emission bands to the optical detectors; and
(v) optical guides in the form of a bifurcated fibre 306 fibre splitter to connect the LED, fluorosensor 300 and detector.
[042] As shown in Figure 3, the fluorosensor 300 includes a polymer coating on a cleaved end of an optical fibre. The fibre is cleaved to expose a fresh tip surface for polymer attachment.
Timed Probe
[043] The first system 100 and/or the second system 200 can include an apparatus in the form of a timed automated probe 110 that performs the method for sensing, described hereinafter containing the fibre and the fluorophore, which can be referred to as a "timed probe" or "automated probe".
[044] The timed probe 110 is controlled by computer-readable instructions to place the fluorosensor 300 in contact with the sample for a pre-determined exposure time (predetermined for the type of fluorosensor 300 and the application), and to remove (by lifting) fluorosensor 300 from the sample. The first system 100 and second system 200 can include a controller that follows the computer-readable instructions to operate the timed probe 110, and to perform the optical measurements once the fluorosensor 300 is removed from the sample. The timed probe 110 can measure a force of the fluorosensor 300 touching the sample, and can control this force so that the same force is applied for each exposure, thus potentially improving consistency between measurements.
[045] As shown in Figures 10A and 10B, the timed probe 110 can consist of a fiber probe 101 mounted in a handheld rugged outer housing 105. The timed probe 110 includes an aperture 103 that is placed in contact with an area to be measured by the operator. A motorized mechanism 102 (including a motor and a fibre mount) is controlled (by the controller) to move the fluorosensor 300 from an initial retracted position 104 (retracted inside the outer housing 105) to an exposed position 106 (in contact with the tissue surface) for a preselected fixed amount of time. The motorized mechanism 102 is then controlled (by the controller) to retract the fluorosensor 300 from the surface back to the retracted position 104. The optical components are then controlled (by the controller) to perform the optical measurements for the preselected measurement duration. Once the measurements are complete, the first system 100 and second system 200 then alert the operator that the fluorosensor 300 is ready for the next measurement to be performed.
Method
[046] The method includes:
(i) a design process;
(ii) a fabrication process;
(iii) a calibration process; and
(iv) an operational process.
[047] The design process for the fluorosensor 300 for a pre-defined application (which can be cancer detection in human tissue cells) includes:
(i) selecting the fluorophore (which can be CNF) to sense a driving
property (which can be pH) that is relevant to the pre-defined application;
(ii) selecting the matrix (which can be a polymer) in which the
fluorophore is to be doped;
selecting the light source/s (which can be LEDs or lasers) and the optical detector/s (which can be photodiodes or CCD spectrometers) to correspond to the optical excitation band/s (which can include 473 nm) and the optical emission band/s (which can be 565 nm and 705 nm) of the fluorophore; selecting the optical guide/s (which can be optical fibres) to carry the optical excitation light from the light source/s to the fluorophore, and the fluorescent light from the fluorophore to the optical detector/s; selecting a concentration and a volume (which includes the coating thickness and the area of the doped matrix in the fluorosensor 300) for the fluorophore in the fluorosensor 300 based on a pre-determined desired signal intensity, and sensitivity to the driving property (e.g., determined by testing a range of values); determining an optimal exposure time for the fluorophore to be exposed to the sample to allow the fluorophore to sense the driving property (e.g., l-20s for CNF); determining an optimal removal time after the initial removal, during which the fluorophore remains removed from the sample, after which the fluorophore is no longer directly affected by the sample, and the state of the fluorophore is sufficiently stable for the emission to be stable (e.g., this is in the range of 2-10 seconds for a CNF-based fibre sensor); determining a maximum removal time from the initial removal, after which the state of the fluorophore may no longer be sufficiently stable for the emission to be stable (e.g., 10 mins for a CNF-based fibre sensor); and
(ix) determining thresholds of relevance to the pre-defined application (e.g., the pre-determined pH thresholds for cancerous versus noncancerous cells in pH detection). The fabrication process involves creating a matrix into which the fluorophore. An example of this process for CNF in a polymer matrix, includes:
(i) preparing a doped polymer mixture solution at the selected concentration (e.g., for CNF in cellular cancer detection, mixing by weight, 27% acrylamide, 3% bis-acrylamide, 70% pH 6*5 potassium phosphate buffer, 0*4 mg/mL CNF and 40 μΙ7πιΙ_, triethylamine);
(ii) ultrasonicating the polymer mixture until solid components are fully
dissolved, to form a polymer solution;
(iii) preparing the sensor surface for the fluorophore (which can include
cleaving the fibre to expose a fresh tip surface for polymer attachment, and/or dipping the sensor surface into a 2% solution of 3- (trimethoxysilyl)propyl methacrylate in pH 3*5 HC1 for 1 hour, removing the fibre and drying with nitrogen gas);
(iv) coating the polymer solution on the sensor surface (which can be the fibre tip), including binding the doped polymer to the sensor surface by photopolymerisation using the fixing light source (which, for an acrylamide polymer, can be the 405-nm laser, held on for 2 seconds at a coupled power of 13 mW);
(v) connecting the light source/s (which can be the blue 473 nm laser for the pH measurements using CNF) and the optical detector/s to the coated sensor surface using the optical guide/s; and
(vi) calibration of the probe for the desired application.
[049] The sensing process broadly includes exciting the dye with a light source, and collecting and analysing the emitted fluorescence light from the dye. An example of this process with CNF for pH sensing could include:
(i) the optical source exciting the fluorophore (e.g., the 473 nm laser exciting the CNF fluorophore);
(ii) the excited fluorophore generating the fluorescent emission (light);
(iii) the sensor surface receiving at least a portion of the fluorescent emission, coupling the fluorescent emission into the optical guide (the fluorescent emission travels in a back-propagating mode in the optical guide/s), which directs the fluorescent emission to the optical detector/s;
(iv) filtering the excitation wavelength band from the fluorescent emission
wavelength band to resist excess excitation light reaching the detector/s (e.g., using a 473 nm long-pass filter);
(v) the optical detector/s detecting the emission from the fluorophore (which can include the plurality of bands, e.g., at 565 nm and 705 nm for CNF); and
(vi) the processing circuit determining an electronic value representing the
driving property from the detected emission (e.g., a value based on an intensity ratio of the 565 nm and 705 nm peaks for CNF, e.g., including integrating the detected signal intensity under the two peaks— from 500-635 nm and from 635-900 nm— and dividing the area of the first peak by the area of the second peak to give the fluorescence ratio for that measurement).
[050] The calibration process includes:
(i) preparing a plurality of standard samples with different detectable values of the driving property (e.g., potassium phosphate buffers with appropriate ratios of monobasic and dibasic potassium hydrogen phosphate to an ionic strength of 0*1 mM, covering a pH range from 6 0 to 8*8);
(ii) performing the sensing process for each of the standard samples, and generating a calibration table of calibration relationships to associate the detectable values of the driving property (in the standard samples) with the respective electronic values representing the driving property from the processing circuit, and obtaining a result after performing the lift and measure procedure.
[051] The operational process for an application of measuring surface pH in tissue samples includes:
(i) preparing the sample surface (e.g., exposing a tissue surface by excision);
(ii) exposing the fluorophore to the sample at one of a plurality of locations ("sensing locations") on the sample surface for the determined optimum exposure time, e.g., manually using a handheld housing that guides the fluorophore;
(iii) removing the fluorophore from the sample so that the fluorophore is not exposed to the driving property (including lifting the probe from the sample surface manually or automatically) for at least the minimum removal time;
(iv) after waiting for the optimum removal time, with the fluorophore
removed from the sample, electronically performing the sensing process for that sensing location before the maximum removal time is reached;
(v) generating an estimated value of the driving property at that sensing location using the calibration table;
(vi) automatically recording the estimated value in computer-readable
storage;
(vii) the computer processor comparing the estimated value to the predetermined thresholds (e.g., the pre-determined pH thresholds for
cancerous versus non-cancerous cells), and generating an alert or display (e.g., alerting a surgeon to a likelihood of cancer in each sensing location); and
(viii) moving to a next one of the sensing locations, and repeating steps (iii) to (viii) until all of the sensing locations have been sensed.
Experimental Example 1
[052] In a first experimental example, the fluorosensor 300 (referred to as a "probe") was prepared using CNF on an optical fibre. The probe response to pH was recorded with a series of PBS buffer solutions. As shown in Figure 6, where the horizontal axis is wavelength (in nm) and the vertical axis is intensity (in counts), the ratiometric response of the probe in air, after dipping into the buffer solutions and removal, increased
monotonically with increasing pH. A fluorescence ratio for each ratiometric response was generated by integrating the two fluorescence bands. As shown in Figure 7, where the horizontal axis is pH in units, and the vertical axis is the ratio, the fluorescence ratios for the exposed fluorosensors (circles) were similar to the fluorescence ratios for the removed fluorosensors (squares), thus showing the removed fluorophores remained sensitive to the sample pH even after removal from the direct exposure. The fluorescence ratios in Figure 7 formed calibration curves for the probe. Upon removal of the fibre from the solution, changes in the fluorescence spectra were observed for 1-5 seconds, e.g., due to evaporation of the solvent from the tip of the fibre. After this rapid change, the signal was observed to be stable for at least 10 minutes.
[053] The structures of the fluorophore CNF both after and before reaction with hydrogen ions (i.e., in a low pH state, which is the protonated acid form, and in a high pH state, which is the closed lactone form) are shown in Figures 4 and 5 respectively. When the fluorophore is exposed to the sample, the fluorophore equilibrates to one of the structures, and remains in this form (e.g., the protonated form or lactone form for CNF) ever after removal, for the determined maximum removed time. The fluorosensor 300 includes a population of many fluorophore molecules, and the fluorescent emission in each
of the bands represents which fraction of the molecules on average is in the state corresponding to that band.
Experimental Example 2
[054] In a second experimental example, the fluorosensor 300 (referred to as a "probe") was prepared using CNF on an optical fibre. Sheep tissue samples were spiked with 1M hydrochloric acid or 1M sodium hydroxide to obtain acidic or basic samples respectively. Emission spectra were recorded with the probe both touching and lifted from the tissue surface. As shown in Figure 8, where the vertical axis is intensity (CPS) and the horizontal axis is wavelength (nm), the autofluorescence background can form a significant fraction of the total signal strength and can form a significant contribution to the apparent measured ratio for the emission spectra of the acidic samples and the basic samples when the probe was touching the samples. In contrast, the autofluoresence was substantially reduced for the removed acidic spectrum and removed basic spectrum.
Experimental Example 3
[055] In a third experimental example, the fluorosensor 300 (or "probe") was prepared using CNF on an optical fibre. Surgically excised human breast cancer and melanoma tissue samples were measured with the optical fibre pH probe at numerous locations over the sample surface, with the tissue type of each location confirmed later by histopathology. Tissue surface measurements were obtained from four melanoma and four breast cancer samples. Results were normalised to the mean value of the normal tissue results to simplify comparison between samples. Statistical data were plotted for normal and tumour datasets, showing the mean, median, and standard deviation. The mean and standard deviation values are shown in Table 1 hereinafter, along with descriptions of the specimen and tumour type for each of the measured samples. As shown in Figure 9, where the horizontal axis is the sample reference number, and the vertical axis is the normalized fluorescence ratio, the tumour samples were generally more acidic than the normal tissue samples by a statistically significant difference in detected pH. The necrotic tumour samples displayed a similar pH to regular cancer samples, while fibrosis samples showed a similar pH to normal tissue. In Figure 9, the tumour (grey) and normal (black)
measurement datasets for each of the eight tissue samples are shown by individual location measurements (circles). Individual measurements of fibrosis (black triangle), necrotic tumour (grey triangle) and those near normal-tumour margins (square) are also plotted, without any corresponding statistical features, for comparison. Cancerous tissue could be differentiated from normal tissue in the fresh human tissue biopsies by measurement of the tissue pH using the optical probe, allowing for measurements to be performed rapidly with high spatial resolution (e.g., 200 μιη in diameter, equivalent to an area in the order of five to ten cells wide, with the potential to reduce this to the measurement of single cells by reducing the size of the fibre probe).
[056] Table 1 - Summary of measurements in Experimental Example 3
Alternatives
[057] A multi-core imaging fibre, with a plurality of fluorosensors on the ends of the respective cores, can be used to increase sample throughput by measuring the tissue pH of a respective plurality of sensor locations simultaneously (i.e., in parallel) with a
corresponding plurality of respective optical detectors (i.e., parallel detectors), which may be a charge-coupled device (CCD) or camera.
[058] Measurement time can be controlled by varying the thickness of the fluorophore layer (e.g., the polymer layer). Using a thinner layer gives a faster response; however the signal intensity is reduced.
[059] Many modifications will be apparent to those skilled in the art without departing from the scope of the present invention.
Interpretation
[060] The reference in this specification to any prior publication (or information derived from it), or to any matter which is known, is not, and should not be taken as an acknowledgment or admission or any form of suggestion that that prior publication (or information derived from it) or known matter forms part of the common general knowledge in the field of endeavour to which this specification relates.
Claims
1. A method for optical sensing, the method including:
(i) exposing a fluorophore to a sample having a chemical driving property to which the fluorophore is sensitive;
(ii) removing the fluorophore from the sample so that the fluorophore is not exposed to the driving property; and
(iii) detecting an optical signal from the removed fluorophore that represents the driving property.
2. The method of claim 1, wherein removing the fluorophore includes lifting the fluorophore upwards from the sample.
3. The method of claim 1 or 2, wherein the detecting includes determining a value of the driving property from the optical signal.
4. The method of claims 1 to 3, wherein the driving property is any one of: acidity and alkalinity; a concentration of hydrogen peroxide; an ion concentration; and a nitric oxide concentration.
5. The method of claims 1 to 4, wherein exposing the fluorophore includes placing the fluorophore in contact with the sample such that the fluorophore is in an environment of the sample with the driving property.
6. The method of claims 1 to 5, including waiting for the fluorophore to respond to the property for a selected exposure time such that the fluorophore responds to a value of the driving property.
7. The method of the preceding claim, wherein the selected exposure time is a preselected time that is predetermined for the application and for the fluorophore.
8. The method of claims 1 to 7, including illuminating the fluorophore with an optical beam from a remote light source.
9. The method of claims 1 to 8, including monitoring one or more portions of an emission spectrum from the removed fluorophore using a remote optical detector.
10. The method of claims 1 to 9, including guiding the optical signal from the fluorophore to an optical detector using one or more optical fibres.
11. The method of the preceding claim, wherein the optical fibres are optical fibre patch cables.
12. The method of claims 1 to 11, wherein the fluorophore is on or in a sensor surface.
13. The method of the preceding claim, wherein the sensor surface is a tip of an optical fibre.
14. The method of claims 1 to 13, including mounting the fluorophore in a matrix.
15. The method of claim 14, wherein the matrix includes a polymer.
16. The method of claim 14 or 15, including applying the matrix to a sensor surface.
17. The method of claims 14 to 16, wherein the matrix is an acrylamide polymer.
18. The method of claims 14 to 17, including fixing the matrix using light.
19. The method of claims 1 to 18, wherein the sample is a human tissue sample that includes cancerous cells and non-cancerous cells, wherein the method includes
determining whether the sample is cancerous or non-cancerous based on the driving property.
20. The method of claims 1 to 19, including comparing a determined value of the driving property with one or more predetermined values associated with respective different sample types.
21. The method of claims 1 to 20, including detecting the optical signal having a plurality of emission bands, and detecting an intensity ratio between at least two of the emission bands.
22. The method of claims 1 to 21, including: generating calibration relationships between pre-prepared samples and respective detected optical signals for the fluorosensor; and using the calibration relationships to determine a value of the driving property from the optical signal.
23. A system for optical sensing, the system including an automated probe configured to perform the method of claims 1 to 22.
24. A system for optical sensing, the system including: one or more fluorosensors; one or more optical sources to cause the fluorosensors to fluoresce; one or more optical detectors to detect the fluorescence; and a processor that generates electronic signals representing the measured driving property and/or a comparison of the measured driving property with a predetermined value based on electronic signals from the optical detectors.
25. The system of claim 24, wherein the processor records and displays the electronic signals.
26. The system of claim 24 or 25, wherein the optical detectors are provided by a spectrum analyser and/or photodiodes.
27. The system of claims 24 to 26, including a protective housing configured for portable use in a hospital operating theatre.
28. An apparatus for optical sensing, the apparatus including: a fluorophore; an optical guide for guiding an optical beam to illuminate the fluorophore, and for guiding an optical signal from the fluorophore, wherein the fluorophore is on or in a tip of the optical guide; and a handheld housing for manually placing the fluorophore in contact with a sample having a detectable property.
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
AU2016900338A AU2016900338A0 (en) | 2016-02-02 | Apparatus, method and system | |
AU2016900338 | 2016-02-02 |
Publications (1)
Publication Number | Publication Date |
---|---|
WO2017132727A1 true WO2017132727A1 (en) | 2017-08-10 |
Family
ID=59499170
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
PCT/AU2017/050082 WO2017132727A1 (en) | 2016-02-02 | 2017-02-01 | Apparatus, method and system for detecting a chemical driving property using a fluorophore |
Country Status (1)
Country | Link |
---|---|
WO (1) | WO2017132727A1 (en) |
Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US11324425B2 (en) | 2017-11-28 | 2022-05-10 | Alan D. Kersey | Apparatus and method for assessment of cancer margin |
CN115656132A (en) * | 2022-11-09 | 2023-01-31 | 淮阴工学院 | Novel multiplexing optical fiber active oxygen sensor and method for detecting active oxygen concentration |
Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2001006253A2 (en) * | 1999-07-16 | 2001-01-25 | Board Of Regents, The University Of Texas System | Detection system based on an analyte reactive particle |
US20020122612A1 (en) * | 1997-03-14 | 2002-09-05 | Trustees Of Tufts College | Fiber optic sensor with encoded microspheres |
WO2002077291A1 (en) * | 2001-03-23 | 2002-10-03 | The Trustees Of Tufts College | Methods for detecting target analytes and enzymatic reactions |
WO2011103497A1 (en) * | 2010-02-19 | 2011-08-25 | Pacific Biosciences Of California, Inc. | Illumination of integrated analytical systems |
WO2012098241A2 (en) * | 2011-01-20 | 2012-07-26 | City University | Fibre-optic sensor |
-
2017
- 2017-02-01 WO PCT/AU2017/050082 patent/WO2017132727A1/en active Application Filing
Patent Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20020122612A1 (en) * | 1997-03-14 | 2002-09-05 | Trustees Of Tufts College | Fiber optic sensor with encoded microspheres |
WO2001006253A2 (en) * | 1999-07-16 | 2001-01-25 | Board Of Regents, The University Of Texas System | Detection system based on an analyte reactive particle |
WO2002077291A1 (en) * | 2001-03-23 | 2002-10-03 | The Trustees Of Tufts College | Methods for detecting target analytes and enzymatic reactions |
WO2011103497A1 (en) * | 2010-02-19 | 2011-08-25 | Pacific Biosciences Of California, Inc. | Illumination of integrated analytical systems |
WO2012098241A2 (en) * | 2011-01-20 | 2012-07-26 | City University | Fibre-optic sensor |
Non-Patent Citations (2)
Title |
---|
GLICKMAN, R.D. ET AL.: "Intracellular photochemical reactions in the RPE cell exhibit a wavelength dependence that resembles the action spectrum of melanin", PROCEEDINGS OF THE SPIE 3601, LASER -TISSUE INTERACTION X; PHOTOCHEMICAL, PHOTOTHERMAL, AND PHOTOMECHANICAL, vol. 94, 14 June 1999 (1999-06-14) * |
SCHARTNER, E.P. ET AL.: "Cancer Detection in Human Tissue Samples Using a Fiber- Tip pH Probe", CANCER RESEARCH, vol. 76, no. 23, 30 November 2016 (2016-11-30), pages 6795 - 6801, XP055440364, DOI: doi:10.1158/0008-5472.CAN-16-1285 * |
Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US11324425B2 (en) | 2017-11-28 | 2022-05-10 | Alan D. Kersey | Apparatus and method for assessment of cancer margin |
CN115656132A (en) * | 2022-11-09 | 2023-01-31 | 淮阴工学院 | Novel multiplexing optical fiber active oxygen sensor and method for detecting active oxygen concentration |
CN115656132B (en) * | 2022-11-09 | 2023-12-01 | 淮阴工学院 | Multiplexing optical fiber active oxygen sensor and method for detecting active oxygen concentration thereof |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Broussard et al. | Fluorescence resonance energy transfer microscopy as demonstrated by measuring the activation of the serine/threonine kinase Akt | |
US10874333B2 (en) | Systems and methods for diagnosis of middle ear conditions and detection of analytes in the tympanic membrane | |
JP5122481B2 (en) | Improvements in and related to imaging of biological samples | |
van Munster et al. | Fluorescence lifetime imaging microscopy (FLIM) | |
JP4982386B2 (en) | Apparatus and measurement method for measuring signal from fluorescent nanodrop held at surface tension | |
US11219370B2 (en) | Devices and methods for optical pathology | |
US20080294032A1 (en) | Spectral Imaging of Biological Samples | |
WO2005040769A2 (en) | Spectral imaging of biological samples | |
Sordillo et al. | Optical spectral fingerprints of tissues from patients with different breast cancer histologies using a novel fluorescence spectroscopic device | |
CN1302210A (en) | Acetic acid as a signal enhancing contrast agent in fluorescence spectroscopy | |
Schartner et al. | Cancer detection in human tissue samples using a fiber-tip pH probe | |
WO1990012536A1 (en) | Spectral diagnosis of diseased tissue | |
BRPI0714697A2 (en) | Method for the determination in vivo of the amount of nuclear nucleic acids in at least one cell of an animal or human subject, and use of a device | |
Schulmerich et al. | Transcutaneous Raman spectroscopy of bone tissue using a non-confocal fiber optic array probe | |
EP2805151B1 (en) | Optode for determining chemical parameters | |
Zhuo et al. | The layered–resolved microstructure and spectroscopy of mouse oral mucosa using multiphoton microscopy | |
WO2017132727A1 (en) | Apparatus, method and system for detecting a chemical driving property using a fluorophore | |
CN108351302B (en) | Method and device for determining tumor site | |
CN109060746B (en) | Ratio type fluorescence detection method and application | |
US20130096438A1 (en) | Method and device for detecting tumorous tissue in the gastrointestinal tract with the aid of an endocapsule | |
KR101758114B1 (en) | Method for measuring fluorescence lifetime according to energy transfer | |
US11324425B2 (en) | Apparatus and method for assessment of cancer margin | |
Houlne et al. | Imaging and quantitation of a tissue-selective lanthanide chelate using an endoscopic fluorometer | |
US20240035976A1 (en) | Uv excited multi spectral fluorescence based tissue analysis with raman spectroscopy zoom-in scanning | |
Banerjee et al. | Detection of murine intestinal adenomas using targeted molecular autofluorescence |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
121 | Ep: the epo has been informed by wipo that ep was designated in this application |
Ref document number: 17746641 Country of ref document: EP Kind code of ref document: A1 |
|
NENP | Non-entry into the national phase |
Ref country code: DE |
|
122 | Ep: pct application non-entry in european phase |
Ref document number: 17746641 Country of ref document: EP Kind code of ref document: A1 |