Highly multiplexed confocal detection systems and methods of using same
First Claim
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1. A multiplex fluorescence detection system, comprising:
- an excitation illumination source;
an optical train comprising an illumination path and a fluorescence path, wherein;
the illumination path comprises an optical train that comprises multiplex optics that convert a single originating illumination beam from the excitation illumination source into at least 10 discrete illumination beams, and an objective lens that focuses the at least 10 discrete illumination beams onto at least 10 discrete locations on a substrate; and
the fluorescence path comprises collection and transmission optics that receive fluorescent signals from the at least 10 discrete locations, and separately direct the fluorescent signals from each of the at least 10 discrete locations through a confocal filter and focus the fluorescent signals onto a different location on a detector.
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Abstract
Systems and methods for analyzing highly multiplexed sample arrays using highly multiplexed, high density optical systems to illuminate high density sample arrays and/or provide detection and preferably confocal detection off signals emanating from such high density arrays. Systems and methods are applied in a variety of different analytical operations, including analysis of biological and biochemical reactions, including nucleic acid synthesis and derivation of sequence information from such synthesis.
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Citations
59 Claims
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1. A multiplex fluorescence detection system, comprising:
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an excitation illumination source; an optical train comprising an illumination path and a fluorescence path, wherein; the illumination path comprises an optical train that comprises multiplex optics that convert a single originating illumination beam from the excitation illumination source into at least 10 discrete illumination beams, and an objective lens that focuses the at least 10 discrete illumination beams onto at least 10 discrete locations on a substrate; and the fluorescence path comprises collection and transmission optics that receive fluorescent signals from the at least 10 discrete locations, and separately direct the fluorescent signals from each of the at least 10 discrete locations through a confocal filter and focus the fluorescent signals onto a different location on a detector. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40)
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41. A system for detecting fluorescence from a plurality of discrete locations on a substrate, comprising:
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a substrate; an excitation illumination source; a detector; and an optical train positioned to receive an originating illumination beam from the excitation illumination source, and configured to; convert the originating illumination beam into a plurality of discrete illumination beams; focus the plurality of discrete illumination beams onto a plurality of discrete locations on the substrate, wherein the plurality of discrete locations are at a density of greater than 1000 discrete locations per mm2; receive a plurality of discrete fluorescent signals from the plurality of discrete locations; and focus the plurality of discrete fluorescent signals through a confocal filter, onto the detector.
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42. A system for collecting fluorescent signals from a plurality of locations on a substrate, comprising:
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excitation illumination optics configured to simultaneously provide excitation radiation to an area of a substrate that includes the plurality of locations; and fluorescence collection and transmission optics that receive fluorescent signals from the plurality of locations on the substrate, and separately direct the fluorescent signals from each of the plurality of locations through a separate confocal aperture in a confocal filter and image the fluorescent signals onto a detector.
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43. A system for detecting fluorescent signals from a plurality of discrete locations on a substrate, comprising:
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an excitation illumination source; a diffractive optical element positioned to convert a single originating illumination beam from the excitation illumination source into at least 10 discrete beams each propagating at a unique angle relative to the originating beam; an objective for focusing the at least ten discrete beams onto at least 10 discrete locations on a substrate; fluorescence collection and transmission optics; and a detector, wherein the fluorescence collection and transmission optics are positioned to receive fluorescent signals from the plurality of discrete locations and transmit the fluorescent signals to the detector. - View Dependent Claims (58, 59)
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44. A method of detecting a plurality of discrete fluorescent signals from a plurality of discrete locations on a substrate, comprising:
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simultaneously and separately illuminating each of the plurality of discrete locations on the substrate with excitation illumination; simultaneously collecting fluorescent signals from each of the plurality of locations and separately directing each of the fluorescent signals from the plurality of discrete locations through a confocal filter; and separately imaging the fluorescent signals from each of the plurality of discrete locations on the substrate onto a discrete location on a detector. - View Dependent Claims (45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57)
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Specification