US5324401A
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US
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Multiplexed fluorescence detector system for capillary electrophoresis
Assignee: Iowa State University Research Foundation Inc
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Filing: 1993-02-05
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Granted: 1994-06-28
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Expires: 2013-02-05
Abstract
Abstract A fluorescence detection system for capillary electrophoresis is provided wherein the detection system can simultaneously excite fluorescence and substantially simultaneously monitor separations in multiple capillaries. This multiplexing approach involves laser irradiation of a sample in a plurality of capillaries through optical fibers that are coupled individually with the capillaries. The array is imaged orthogonally through a microscope onto a charge-coupled device camera for signal analysis.
Inventors
- 👤 Edward S. Yeung
- 👤 John A. Taylor
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-05 22:56
CPC Classifications
G; G01; G01N; G01N27/00; G01N27/26; G01N27/416; G01N27/447; G01N27/44704; G01N27/44717; G01N27/44721
Claims Analysis
Total Claims: 42
Claim 1
Independent Claim
172 words
A multiplexed fluorescence detection system for use with capillary electrophoresis to detect target species in a sample, comprising: (a) an array of at least two capillaries each having an intake end, an outflow end, and an annular wall wherein at least a portion of the wall is translucent to light having a wavelength of about 200-1,500 nm, the intake end being in fluid communication with the sample so that the sample is drawn into the capillary; (b) an array of at least two optical fibers, each optical fiber having a first and second end, the second end of each optical fiber being coupled to a capillary in the array of capillaries; (c) means for generating coherent light positioned to direct the coherent light onto the first end of the optical fibers, wherein the coherent light is of a wavelength suitable to cause fluorescence in target species present in the capillaries; and (d) means for substantially simultaneously detecting changes in fluorescent emission by the target species through the annular wall of each capillary.
Claim 2
Dependent Claim
172 words
A multiplexed fluorescence detection system for use with capillary electrophoresis to detect target species in a sample, comprising: (a) an array of at least two capillaries each having an intake end, an outflow end, and an annular wall wherein at least a portion of the wall is translucent to light having a wavelength of about 200-1,500 nm, the intake end being in fluid communication with the sample so that the sample is drawn into the capillary; (b) an array of at least two optical fibers, each optical fiber having a first and second end, the second end of each optical fiber being coupled to a capillary in the array of capillaries; (c) means for generating coherent light positioned to direct the coherent light onto the first end of the optical fibers, wherein the coherent light is of a wavelength suitable to cause fluorescence in target species present in the capillaries; and (d) means for substantially simultaneously detecting changes in fluorescent emission by the target species through the annular wall of each capillary.
Claim 3
Dependent Claim
40 words
The detection system of claim 1 wherein the capillaries have an inside diameter of about 20-500 microns and at least one optical fiber is coupled to each capillary in a manner to transmit the coherent light axially into the capillary.
Claim 4
Dependent Claim
53 words
The detection system of claim 2 wherein the optical fiber is coupled to the capillary by insertion of the second end of the optical fiber into the outflow end of the capillary, the optical fiber being of a suitable diameter for affixing the fiber inside the capillary without substantially hindering the sample flow.
Claim 5
Dependent Claim
28 words
The detection system of claim 1 wherein at least one optical fiber is coupled to each capillary and positioned adjacent to and orthogonally in relation to the capillary.
Claim 6
Dependent Claim
13 words
The detection system of claim 1 further comprising a means for focusing light.
Claim 7
Dependent Claim
32 words
The detection system of claim 5 wherein the focusing means is a lens for focusing fluorescent light emitted from the target species in the capillaries to the means for detecting fluorescent light.
Claim 8
Dependent Claim
34 words
The detection system of claim 1 wherein the means for generating coherent light includes collimating means for distributing the coherent light into the first end of the optical fibers in a substantially uniform manner.
Claim 9
Dependent Claim
21 words
The detection system of claim 1 wherein the means for detecting the fluorescent emission comprises a charged-coupled device based imaging system.
Claim 10
Dependent Claim
22 words
The detection system of claim 1 wherein the means for detecting the fluorescent emission comprises a charge injection device based imaging system.
Claim 11
Dependent Claim
23 words
The detection system of claim 1 further comprising a spacer positioned between at least two capillaries to reduce optical crosstalk between the capillaries.
Claim 12
Dependent Claim
19 words
The detection system of claim 1 wherein the means for generating coherent light delivers about 0.05-10 mW per fiber.
Claim 13
Dependent Claim
17 words
The detection system of claim 1 wherein the coherent light has a wavelength of about 250-700 nm.
Claim 14
Dependent Claim
16 words
The detection system of claim 1 wherein the array of capillaries comprises at least 100 capillaries.
Claim 15
Dependent Claim
16 words
The detection system of claim 1 wherein the array of capillaries comprises at least 1000 capillaries.
Claim 16
Dependent Claim
24 words
The detection system of claim 1 wherein at least one of the capillaries contains a gel that is suitable for capillary gel electrophoresis systems.
Claim 17
Dependent Claim
16 words
The detection system of claim 1 wherein at least one of the capillaries contains no gel.
Claim 18
Dependent Claim
116 words
A multiplexed method for detecting the presence of fluorescent target species in a sample in a plurality of capillaries using capillary electrophoresis, comprising the steps of: (a) introducing samples containing a fluorescent species into intake ends of a plurality of capillaries, each capillary having an annular wall, and an outflow end, wherein the samples migrate through the capillaries towards the outflow ends; (b) irradiating the samples in the capillaries with coherent light transmitted through optical fibers, the optical fibers having first ends coupled to a coherent light source and second ends individually coupled to the capillaries; (c) substantially simultaneously detecting change in fluorescent emission by the fluorescent species through the wall of each of the capillaries.
Claim 19
Dependent Claim
116 words
A multiplexed method for detecting the presence of fluorescent target species in a sample in a plurality of capillaries using capillary electrophoresis, comprising the steps of: (a) introducing samples containing a fluorescent species into intake ends of a plurality of capillaries, each capillary having an annular wall, and an outflow end, wherein the samples migrate through the capillaries towards the outflow ends; (b) irradiating the samples in the capillaries with coherent light transmitted through optical fibers, the optical fibers having first ends coupled to a coherent light source and second ends individually coupled to the capillaries; (c) substantially simultaneously detecting change in fluorescent emission by the fluorescent species through the wall of each of the capillaries.
Claim 20
Dependent Claim
55 words
The method of claim 17 wherein the step of irradiating comprises transmitting coherent light axially into a capillaries by inserting the second ends of the optical fibers into the outflow ends of the capillaries, the optical fiber having a diameter suitable for affixing the fiber in position in the capillary without substantially hindering sample flow.
Claim 21
Dependent Claim
34 words
The method of claim 17 wherein the step of irradiating comprises irradiating coherent light orthogonally into the capillaries through an optical fiber that is coupled adjacent to, and perpendicular in relation to the capillary.
Claim 22
Dependent Claim
25 words
The method of claim 17 wherein the step of detecting fluorescent emission comprises using a charged-coupled device based imaging system for detecting the fluorescent emissions.
Claim 23
Dependent Claim
26 words
The method of claim 17 wherein the step of detecting fluorescent emission comprises using a charge injection device based imaging system for detecting the fluorescent emission.
Claim 24
Dependent Claim
37 words
The method of claim 17 further comprising a step of using a lens for evenly distributing the coherent light generated by a laser axially into a plurality of optical fibers at an end distal to the capillaries.
Claim 25
Dependent Claim
23 words
The method of claim 17 wherein the step of irradiating comprises irradiating coherent light at a power of about 0.05-10 mW per fiber.
Claim 26
Dependent Claim
21 words
The method of claim 17 wherein the step of irradiating comprises irradiating coherent light with a wavelength of about 250-700 nm.
Claim 27
Dependent Claim
24 words
The method of claim 17 wherein the step of introducing samples into capillaries comprises introducing samples into an array of at least 100 capillaries.
Claim 28
Dependent Claim
24 words
The method of claim 17 wherein the step of introducing samples into capillaries comprises introducing samples into an array of at least 1000 capillaries.
Claim 29
Dependent Claim
33 words
The method of claim 17 wherein the step of introducing samples into capillaries comprises introducing a sample into at least one capillary that contains a gel that is suitable for capillary gel electrophoresis.
Claim 30
Dependent Claim
26 words
The method of claim 17 wherein the step of introducing samples into capillaries comprises introducing a sample into at least one capillary that contains no gel.
Claim 31
Dependent Claim
138 words
A multiplexed method of detecting the presence of macromolecule in a sample present in a plurality of capillaries in capillary electrophoresis, comprising the steps of: (a) linking a fluorophore to the macromolecule in a sample to transform the macromolecule into a fluorescent species with fluorescence that can be excited by light of suitable wavelength; (b) introducing samples containing fluorescent species into each of a plurality of capillaries, each capillary having an annular wall, an intake end and an outflow end; (c) irradiating each sample in each capillary with coherent light to cause fluorescence of the macromolecule, the coherent light being transmitted though optical fibers individually coupled to the capillaries; and (d) substantially simultaneously detecting changes in fluorescent emission through the wall of each of the capillaries by the fluorescent species using a charge transfer device based imaging system.
Claim 32
Dependent Claim
138 words
A multiplexed method of detecting the presence of macromolecule in a sample present in a plurality of capillaries in capillary electrophoresis, comprising the steps of: (a) linking a fluorophore to the macromolecule in a sample to transform the macromolecule into a fluorescent species with fluorescence that can be excited by light of suitable wavelength; (b) introducing samples containing fluorescent species into each of a plurality of capillaries, each capillary having an annular wall, an intake end and an outflow end; (c) irradiating each sample in each capillary with coherent light to cause fluorescence of the macromolecule, the coherent light being transmitted though optical fibers individually coupled to the capillaries; and (d) substantially simultaneously detecting changes in fluorescent emission through the wall of each of the capillaries by the fluorescent species using a charge transfer device based imaging system.
Claim 33
Dependent Claim
21 words
The method of claim 29 wherein the step of detecting changes in fluorescence comprises using a charged-coupled device based imaging system.
Claim 34
Dependent Claim
22 words
The method of claim 29 wherein the step of detecting change in fluorescence comprises using a charged injection device based imaging system.
Claim 35
Dependent Claim
24 words
The method of claim 29 further comprising a step of introducing a light absorber into the capillaries for reducing photochemical bleaching of fluorescent species.
Claim 36
Dependent Claim
24 words
The method of claim 29 wherein the step of introducing samples into capillaries comprises introducing samples into an array of at least 100 capillaries.
Claim 37
Dependent Claim
54 words
The method of claim 29 wherein the step of linking a fluorophore to the macromolecule comprises linking a fluorophore to the macromolecule in a material selected from the group consisting of proteins, amino acids, polypeptides, carbohydrate, polysaccharides, oligonucleotides, nucleic acids, RNA, DNA, bacteria, viruses, chromosomes, genes, organelles, as well as fragments and combinations thereof.
Claim 38
Dependent Claim
42 words
The method of claim 29 wherein the step of linking a fluorophore to the macromolecule comprises linking the macromolecule to a fluorophore selected from the group consisting salicylate, 3,3'-diethylthiadicarbocyanine iodide (DTDCI), dansyl chloride, fluorescein, fluorescein isothiocyanate, ethidium bromide, rhodamine, and combinations thereof.
Claim 39
Dependent Claim
105 words
A multiplexed method of detecting the presence of macromolecules in samples present in a plurality of capillaries in capillary electrophoresis, comprising the steps of: (a) providing a buffer solution containing a fluorophore; (b) introducing samples into a plurality of capillaries in a capillary electrophoresis system, each capillary containing the buffer; (c) irradiating the samples in the capillaries with coherent light to cause fluorescence, the coherent light being transmitted though optical fibers individually coupled to the capillaries; and (d) substantially simultaneously detecting changes in fluorescent emission through the wall of each the capillaries by the fluorescent species by using a charge transfer device based imaging system.
Claim 40
Dependent Claim
105 words
A multiplexed method of detecting the presence of macromolecules in samples present in a plurality of capillaries in capillary electrophoresis, comprising the steps of: (a) providing a buffer solution containing a fluorophore; (b) introducing samples into a plurality of capillaries in a capillary electrophoresis system, each capillary containing the buffer; (c) irradiating the samples in the capillaries with coherent light to cause fluorescence, the coherent light being transmitted though optical fibers individually coupled to the capillaries; and (d) substantially simultaneously detecting changes in fluorescent emission through the wall of each the capillaries by the fluorescent species by using a charge transfer device based imaging system.
Claim 41
Dependent Claim
154 words
A multiplexed method of detecting the presence of DNA fragments in samples present in a plurality of capillaries in capillary electrophoresis, comprising the steps of: (a) linking a fluorophore to a DNA fragment in a sample to transform it into a fluorescent species; (b) introducing samples containing fluorescent species into a plurality of capillaries in a capillary electrophoresis system, the system comprises an array of at least about 100 capillaries each having an inside diameter of about 20-500 microns, and an annular wall; (c) irradiating the samples in the capillaries with coherent light having a wavelength of about 250-700 nm to cause native fluorescence of the DNA fragment, the coherent light being transmitted though optical fibers individually coupled to the capillaries; and (d) substantially simultaneously detecting change in fluorescence emitted by the fluorescent species through the wall of each of the capillaries by focusing the fluorescence onto a charge transfer device based imaging system.
Claim 42
Dependent Claim
154 words
A multiplexed method of detecting the presence of DNA fragments in samples present in a plurality of capillaries in capillary electrophoresis, comprising the steps of: (a) linking a fluorophore to a DNA fragment in a sample to transform it into a fluorescent species; (b) introducing samples containing fluorescent species into a plurality of capillaries in a capillary electrophoresis system, the system comprises an array of at least about 100 capillaries each having an inside diameter of about 20-500 microns, and an annular wall; (c) irradiating the samples in the capillaries with coherent light having a wavelength of about 250-700 nm to cause native fluorescence of the DNA fragment, the coherent light being transmitted though optical fibers individually coupled to the capillaries; and (d) substantially simultaneously detecting change in fluorescence emitted by the fluorescent species through the wall of each of the capillaries by focusing the fluorescence onto a charge transfer device based imaging system.
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