Optics: measuring and testing – Document pattern analysis or verification
Reexamination Certificate
1999-11-04
2004-08-31
Nguyen, Tu T. (Department: 2877)
Optics: measuring and testing
Document pattern analysis or verification
Reexamination Certificate
active
06784982
ABSTRACT:
BACKGROUND
A DNA chip is a rigid flat surface, typically glass or silicon, with short chains of related nucleic acids spotted in rows and columns on it. Hybridization between a fluorescently-labeled DNA and specific locations on the chip can be detected and analyzed by computer-based instrumentation. The information derived from the results of hybridization to DNA chips is stimulating advances in drug development, gene discovery, gene therapy, gene expression, genetic counseling, and plant biotechnology.
Among the technologies for creating DNA chips are photolithograpy, “on-chip” synthesis, piezoelectric printing, and direct printing. Chip dimensions, the number of sites of DNA deposition (sometimes termed “addresses”) per chip, and the width of the DNA spot per “address” are dependent upon the technologies employed for deposition. The most commonly used technologies produce spots with diameters of 50-300 &mgr;m. Photolithography produces spots that can have diameters as small as 1 micron. Technologies for making such chips are known to those skilled in these arts and arc described, for instance, in U.S. Pat. Nos. 5,925,525, 5,919,523, 5,837,832, and 5,744,305, which are incorporated herein by reference.
Hybridization to DNA chips can be monitored by fluorescence optics, by radioisotope detection, and by mass spectrometry. The most widely-used method for detection of hybridization employs fluorescently-labeled DNA, and a computerized system featuring a confocal fluorescence microscope (or an epifluorescence microscope), a movable microscope stage, and DNA detection software. Technical characteristics of these microscope systems are described in U.S. Pat. Nos., 5,293,563, 5,459,325, and 5,552,928, which are incorporated herein by reference. Further descriptions of imaging fluorescently immobilized biomolecules and analysis of the images are set forth in U.S. Pat. Nos. 5,874,219, 5,871,628, 5,834,758, 5,631,734, 5,578,832, 5,552,322, and 5,556,539 which are incorporated herein by reference.
In brief, these conventional approaches to visualizing the surface of a DNA chip involve placing the chip on a microscope stage, moving the stage to put the sample into focus with a microscope objective, and triggering a digital camera or similar device to capture an image. An objective is a device made of a group of lenses that have a sophisticated design that collects light from the sample, magnifies the image of the sample, and minimizes the unavoidable image and color distortion caused by the passage of the light through the objective. The light-collected from the sample passes through the objective and through a set of mirrors and lenses until is delivered to an eyepiece or the camera. The light path is the path that the light takes from the point where it leaves the surface of the sample until it reaches an imaging device such as an eyepiece or camera. The microscopes are integral with light sources that direct light on to the sample.
These microscopes also have sets of optical filters that allow for viewing of fluorescent images. The DNA that is hybridized to the surface of the DNA chip is typically labeled with fluorescent molecules that absorb light at one wavelength and then emit a different wavelength. The microscope is equipped with sets of optical filters that block the wavelengths of light from the light source but allow the light emitted by the fluorescent molecules to pass through the light path to reach the eyepiece or camera. The light source is typically integral with the microscope and is an important part of the imaging system.
These conventional microscopes are sophisticated and expensive instruments that require training and maintenance. A single microscope objective typically has multiple lenses. A lens, as used herein, means a transparent solid material shaped to magnify, reduce, or redirect light rays. A light filter or mirror is distinct from a lens. Furthermore, use of a microscope requires a dedicated workspace that is approximately the size of a typical desk. Conventional microscopes have a light path that is several centimeters long that transmits the collected light through air and other assorted optical devices within the light path. One of the challenges in microscopy is making the microscope as efficient as possible in capturing all of the light that leaves the sample surface so that an optimal image may be made.
The costly instrumentation conventionally used to image DNA clips impedes the broad usage of DNA chip technologies.
What is needed is an inexpensive, low-maintenance alternative spot detection method for DNA chip analysis that is easy to use and requires a minimum of space and maintenance.
Integrated electronic circuit arrays for light-detection (herein referred to as members of the group of detectors called electronic light detector arrays) and analysis are readily available. They generally are based on CCD (charge-coupled device) or CMOS (complementary metal oxide semiconductor) technologies. Both CCD and CMOS imaging detectors are two-dimensional arrays of electronic light sensors. Each array consists of a set of known, unique positions that are also called addresses. Each address in a CCD or CMOS array is occupied by a sensor that covers an area that is typically shaped as a box or a rectangle; this area or the area occupied by a single sensor is referred to as a pixel. Herein, a light-detecting sensor located on a pixel is called a detector pixel. A detector pixel may be in a CCD sensor, a CMOS sensor, or other device that detects or measures light. The sizes of detector pixels vary widely and may have a diameter or length of 0.2 &mgr;m, which is the theoretical limit of resolution of the light microscope. Thus an invention that directly employs electronic detection instead of a conventional optical system is potentially as powerful as any light microscope. Light, as used herein, means any electromagnetic emission of at least 120 nm wavelength and includes ultraviolet, visible, and infrared light.
CCDs, widely used in consumer and scientific applications such as digital recorders and digital cameras, are most sensitive, and may be made with detector pixels that are smaller than those of CMOS devices. CMOS devices are now beginning to be incorporated in recorders and cameras because they are less expensive to produce. CMOS devices also are easier to interface with external control systems than CCDs. Some readily-available CMOS devices are capable of acquiring, digitizing, and transmitting an image without additional circuitry, while CCD arrays require two or more additional circuit elements to accomplish the same tasks.
SUMMARY OF THE INVENTION
The present invention describes an inexpensive device and method for resolving the light spots emitted by a light-transparent DNA chip. The method is direct mapping of the light emitted by a single DNA spot onto corresponding detector pixels of an electronic light detector array system. One method is to put the DNA chip in direct physical contact with the electronic light detector array system. In a modification of this basic method, a simple optical system, such as a single mapping lens, maps an enlarged or reduced version of the DNA array onto the electronic light detector array. Computer software processes the data from the electronic light detector array system. The data may be treated as a two-dimensional map or otherwise processed as an array.
Implementation of the described method would replace the expensive optical detection systems currently employed for DNA chip analysis with an inexpensive system. This system comprises an electronic light detector array, a filter, and, optionally, a mapping lens system. The invention enables the DNA chip to be mapped onto the electronic light detector array. Thus each position on the DNA chip surface has a corresponding position or set of positions on the detector array whereby a fluorescence at an address on the DNA chip surface is projected onto a known pixel or set of pixels.
Direct mapping is inexpensive. It eliminates the need for a complicated microscope
Blumenfeld Martin
Sanders Mark A.
Talghader Joseph J.
Fish & Richardson P.C.P.A.
Nguyen Tu T.
Regents of the University of Minnesota
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