Patent US5515864A successfully refreshed from Google Patents.
US5515864A
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Method and apparatus for the in vivo measurement of oxygen concentration levels by the indirect determination of fluoescence lifetime
Assignee: Cellview Imaging Inc
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Filing: 1994-04-21
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Granted: 1996-05-14
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Expires: 2014-04-21
Abstract
Abstract Methods are described for the in vivo topographic determination of tissue and bodily fluid oxygen concentration or PO 2 within an imaged tissue, as well as a method for measuring blood or bodily fluid PO 2 with a fiber optic catheter. In the first method, a lipid soluble, biocompatible fluorescent probe substance is administered to an animal body and accumulates within the lipid bilayers of its tissue cells. In the second method, the fluorescent probe substance is conjugated to a large molecular mass protein which causes it to be retained within a bodily fluid such as blood. In the third method describing a catheter design, in which PO 2 is measured at the catheter tip, the tip contains the fluorescent probe substance dissolved in a viscous nonpolar solvent. In all methods described, a fluorescent probe substance is preferred whose fluorescence lifetime is quenched by molecular oxygen, and oxygen concentration or PO 2 is determined by measuring the fluorescence anisotropy of the fluorescent probe substance. The bodily fluid or tissue containing the biocompatible fluorescent probe substance, or the fluorescent probe substance in a nonpolar solvent within a catheter tip, is irradiated with continuous linearly polarized ultraviolet light at a wavelength strongly absorbed by the fluorophore. The emitted fluorescence is resolved into its vector components parallel and perpendicular to the plane of polarization of the excitation light, thereby permitting the calculation of the fluorescence anisotropy of the irradiated specimen. Tissue and/or bodily fluid PO 2 of the imaged specimen, or the PO 2 of a fluid at the tip of a sealed catheter, is determined by applying a mathematical function which relates the fluorescence anisotropy of an O 2 -quenchable fluorophore to oxygen concentration or partial pressure.
Inventors
- 👤 Ralph Zuckerman
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-31 06:38
CPC Classifications
A; A61; A61B; A61B5/00; A61B5/145; A61B5/1455; A61B5/14551; A61B5/14555; G; G01; G01N; G01N21/00; G01N21/62; G01N21/63; G01N21/64; G01N21/6445
Claims Analysis
Total Claims: 21
Claim 1
Independent Claim
161 words
A method of measuring oxygen concentration or partial pressure in bodily fluid or tissue of a living being, the method comprising the steps of: (a) administering a biocompatible, lipid soluble fluorescent probe substance having a fluorescence lifetime (τ) which is quenched by molecular oxygen, to the living being; (b) irradiating the tissue or bodily fluid with continuous, linearly polarized ultraviolet light at wavelengths strongly absorbed by the fluorescent probe substance to result in fluorescence having vector components parallel and perpendicular to the plane of polarization of the ultraviolet light; (c) resolving the fluorescence emitted from the irradiated tissue or bodily fluid of step (b) into vector components parallel and perpendicular to the plane of polarization of the excitation light; (d) calculating fluorescence anisotropy in space or in time of the tissue or fluid so irradiated; and (e) applying a mathematical function which relates fluorescence anisotropy to oxygen concentration or partial pressure, the mathematical function being in the form of an equation.
Claim 2
Independent Claim
161 words
A method of measuring oxygen concentration or partial pressure in bodily fluid or tissue of a living being, the method comprising the steps of: (a) administering a biocompatible, lipid soluble fluorescent probe substance having a fluorescence lifetime (τ) which is quenched by molecular oxygen, to the living being; (b) irradiating the tissue or bodily fluid with continuous, linearly polarized ultraviolet light at wavelengths strongly absorbed by the fluorescent probe substance to result in fluorescence having vector components parallel and perpendicular to the plane of polarization of the ultraviolet light; (c) resolving the fluorescence emitted from the irradiated tissue or bodily fluid of step (b) into vector components parallel and perpendicular to the plane of polarization of the excitation light; (d) calculating fluorescence anisotropy in space or in time of the tissue or fluid so irradiated; and (e) applying a mathematical function which relates fluorescence anisotropy to oxygen concentration or partial pressure, the mathematical function being in the form of an equation.
Claim 3
Dependent Claim
44 words
The method of claim 1 additionally comprising the step of selecting the equation as follows: ##EQU9## wherein α is a Bunsen solubility coefficient, K D is a dynamic quench constant, and τ o is a fluorescence lifetime in the absence of O 2 .
Claim 4
Independent Claim
35 words
The method of either of claims 1 or 2 additionally comprising the step of selecting the biocompatible fluorescent probe substance to have an O 2 -quenchable fluorescence lifetime ranging between about 30 to 135 nanoseconds.
Claim 5
Dependent Claim
113 words
The method of claim 1 additionally comprising the step of selecting the equation as follows: ##EQU10## wherein A is fluorescence anisotropy defined as ##EQU11## wherein I.sub.∥ and I.sub.⊥ are intensities of fluorescence emission having electric vectors respectively parallel and perpendicular to that of linearly polarized exciting radiation, G is an empirical correction factor used to correct for transmission efficiency in parallel and perpendicular planes, R is mean molecular rotation time in radians/sec, A o is fluorescence anisotropy in a "frozen" state, in the absence of Brownian rotation, α is a Bunsen solubility coefficient, K D is a dynamic quench constant, and τ o is fluorescence lifetime in the absence of O 2 .
Claim 6
Independent Claim
29 words
The method of any one of claims 1, 2, or 4 wherein a topographic distribution of tissue or blood PO 2 of an imaged tissue and vasculature is determined.
Claim 7
Dependent Claim
22 words
The method of claim 5 wherein the biocompatible, fluorescent probe substance accumulates within lipid bilayers of tissue and red blood cell membranes.
Claim 8
Dependent Claim
15 words
The method of claim 6 wherein fluorescence anisotropy values of less than 0.4 are obtained.
Claim 9
Dependent Claim
38 words
The method of claim 5 wherein the fluorescent probe substance is conjugated to a large molecular mass protein and is retained within the vasculature of the imaged tissue, or is retained within a bodily fluid other than blood.
Claim 10
Dependent Claim
19 words
The method of claim 8 additionally comprising the step of selecting the large molecular mass protein to be albumin.
Claim 11
Independent Claim
27 words
The method of claims 5 additionally comprising the step of selecting the biocompatible, lipid soluble fluorescent probe substance to be pyrenebutyric acid or a salt form thereof.
Claim 12
Dependent Claim
19 words
The method of claim 5 wherein the biocompatible, lipid soluble fluorescent probe substance is applied topically in a solution.
Claim 13
Dependent Claim
14 words
The method of claim 11 wherein the solution is physiologic saline or dimethyl sulfoxide.
Claim 14
Independent Claim
93 words
The method of any one of claims 1 and 4 wherein PO 2 - or oxygen concentration-dependent anisotropy of the tissue or bodily fluid in the presence of the fluorescent probe substance (A(PO 2 ) or A probe ) may be determined from anisotropy of the tissue or bodily fluid in the absence of the fluorescent probe substance (A tissue ) by an equation as follows: ##EQU12## wherein A total is total anisotropy (A probe plus A tissue ) of an imaged specimen, and a is an empirical constant determined in prior calibration.
Claim 15
Independent Claim
39 words
The method of any one of claims 1, 2, or 4 wherein a topographic distribution of tissue or fluid PO 2 is determined tomographically within a volume of an imaged imaged tissue by means of optical serial sectioning methodologies.
Claim 16
Dependent Claim
20 words
The method of claim 14 additionally comprising the step of selecting the optical serial sectioning methodology to be confocal microscopy.
Claim 17
Dependent Claim
21 words
The method of claim 14 additionally comprising the step of selecting the optical serial sectioning methodology to be digital deconvolution procedures.
Claim 18
Dependent Claim
17 words
The method of claim 1 wherein the lipid soluble fluorescent probe substance is administered intravenously in solution.
Claim 19
Dependent Claim
33 words
The method of claim 1 wherein the lipid soluble fluorescent probe substance is administered intravenously by incorporating the fluorescent probe substance in liposomes and then intravenously administering the liposomes to the living being.
Claim 20
Independent Claim
26 words
The method of either of claims 1 or 8 wherein the fluorescent probe substance is conjugated to a large molecular mass protein and is administered intravenously.
Claim 21
Dependent Claim
24 words
The method of claim 1 wherein the biocompatible fluorescent probe substance is exposed to linearly polarized light having wavelengths less than about 400 nm.
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USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1999-05-14 | 1999-11-13 | 2000-05-14 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 2003-05-14 | 2003-11-13 | 2004-05-14 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 2007-05-14 | 2007-11-13 | 2008-05-14 | $7700.00 | Window Closed / Lapsed |