Optics: measuring and testing – By dispersed light spectroscopy – With sample excitation
Reexamination Certificate
2002-09-30
2004-11-30
Evans, F. L. (Department: 2877)
Optics: measuring and testing
By dispersed light spectroscopy
With sample excitation
C250S458100, C422S082080, C435S006120, C436S172000
Reexamination Certificate
active
06825929
ABSTRACT:
FIELD OF THE INVENTION
This invention relates to arrays, particularly biopolymer arrays such as DNA or protein arrays, which are useful in diagnostic, screening, gene expression analysis, and other applications.
BACKGROUND OF THE INVENTION
For the purposes of the discussion below, cited art is not admitted to constitute prior art to the present application.
Polynucleotide arrays (such as DNA or RNA arrays) and peptide arrays, are known and may be used, for example, as diagnostic or screening tools. Such arrays include regions (sometimes referenced as spots or features) of usually different sequence polynucleotides or peptides arranged in a predetermined configuration on a substrate. The array is “addressable” in that different features have different predetermined locations (“addresses”) on a substrate carrying the array.
Biopolymer arrays can be fabricated using in situ synthesis methods or deposition of the previously obtained biopolymers. The in situ fabrication methods include those described in U.S. Pat. No. 5,449,754 for synthesizing peptide arrays, and in U.S. Pat. No. 6,180,351 and WO 98/41531 and the references cited therein for polynucleotides. In situ methods also include photolithographic techniques such as described, for example, in WO 91/07087, WO 92/10587, WO 92/10588, and U.S. Pat. No. 5,143,854. The deposition methods basically involve depositing biopolymers at predetermined locations on a substrate which are suitably activated such that the biopolymers can link thereto. Biopolymers of different sequence may be deposited at different feature locations on the substrate to yield the completed array. Procedures known in the art for deposition of biopolymers, particularly DNA such as whole oligomers or cDNA, are described, for example, in U.S. Pat. No. 5,807,522 (touching drop dispensers to a substrate), and in PCT publications WO 95/25116 and WO 98/41531, and elsewhere (use of a pulse jet in the form of a piezoelectric inkjet head).
Further details of large scale fabrication of biopolymer arrays by depositing either previously obtained biopolymers or by the in situ method, are disclosed in U.S. Pat. Nos. 6,242,266, 6,232,072, 6,180,351, and U.S. Pat. No. 6,171,797.
In array fabrication, the quantities of DNA available for the array are usually very small and expensive. Sample quantities available for testing are usually also very small and it is therefore desirable to simultaneously test the same sample against a large number of different probes on an array. These conditions require the manufacture and use of arrays with large numbers of very small, closely spaced features.
The arrays, when exposed to a sample, will exhibit a binding pattern. The array can be read by observing this binding pattern by, for example, labeling all targets such as polynucleotide targets (for example, DNA), in the sample with a suitable label (such as a fluorescent compound), scanning an illuminating beam across the array and accurately detecting the fluorescent signal from the different features of the array. Assuming that the different sequence polynucleotides were correctly deposited in accordance with the predetermined configuration, then the observed binding pattern will be indicative of the presence and/or concentration of one or more polynucleotide components in the sample. Peptide or arrays of other chemical moieties can be used in a similar manner.
Techniques and apparatus for scanning chemical arrays are described, for example, in U.S. Pat. Nos. 6,406,849, 6,371,370, 6,355,921, 5,763,870 and U.S. Pat. No. 5,945,679. Apparatus which reads an array by scanning an illuminating beam by the foregoing technique are often referred to as scanners and the technique itself often referred to as scanning. Conventionally, such scanning has been done by illuminating array features on a front surface of the substrate one pixel at a time.
Array scanners typically use one or more laser beams of different waveband as light sources, which are scanned over pixels covering the array features. The lasers are generally set to provide as much light as possible to a scanned array, and consequently the relative intensities of such different waveband light sources are close to equal, or about 2/1 or less. detectors (typically fluorescence detectors) each with a very high light sensitivity is normally desirable to achieve maximum signal-to-noise in detecting hybridized molecules, particularly in array scanners used for DNA sequencing or gene expression studies. At present, photomultiplier tubes (“PMTs”) are still the detector of choice although charge coupled devices (“CCDs”) and avalanche photodiodes (“APDs”) can also be used. PMTs and APDs are typically used for temporally sequential scanning of array features, while CCDs permit scanning many features in parallel (for example, one line of features simultaneously, in which case an illuminating line may be used).
A difficulty in reading chemical arrays is that given the large numbers of features which may be present, for example, many thousands of features, and given the various samples that may be exposed to the array, the fluorescent signals detected from different features can vary over a wide range of intensities. In order to obtain the maximum signal from a feature it is desirable to illuminate the array features with light of an intensity and for a time, as high as possible so as to obtain a detectable signal even from features which may only have a few labels present. On the other hand though, in this situation a detected signal from a feature having many labels may be so strong as to cause detector saturation (that is, further increases in signal from the array feature no longer cause an increase in detector signal output). In this event, meaningful measurements from such features are lost. The range over which meaningful signals may be obtained from a feature by an array reader may be referenced as its dynamic range.
It would be desirable then, to provide an array reader with a high dynamic range covering both features which may produce a low or high detectable signal, and which is not overly complex to construct.
SUMMARY OF THE INVENTION
The present invention provides a method of interrogating an addressable array having a plurality of different chemical features. The method may include simultaneously illuminating different regions of the array with light of different intensities (which, for example can, but need not be, of a same waveband). Light emitted from the different regions in response to the illuminating (which light can, but need not be, of a same waveband) may be simultaneously detected with different detectors.
The present invention further provides an apparatus which includes a light system to provide the simultaneous illumination to the different regions of the array, and different light detectors for the simultaneous detection from the different regions, both as mentioned above.
The present invention further provides an addressable array unit. The addressable array unit may have a plurality of different chemical features (such as different polynucleotide or poly-amino acid features). The array unit may also include instructions that different regions of the array can be simultaneously illuminated with light of different intensities and that light emitted from the different regions in response to the illuminating can be simultaneously detected from those regions with different detectors. The instructions may further include an indication of the relative light intensities for the simultaneously illuminated regions or detector sensitivities for the simultaneously detected regions. Alternatively, rather than the array unit including any instructions, it may instead include a code (for example, a machine readable code) linked to such instructions or a combination of part of the instructions and a code linked to the remainder. Such a code can, for example, be located on a substrate carrying the array or an element which carries the substrate.
The present invention also provides a method for receiving a code as already described, and
Agilent Technologie,s Inc.
Evans F. L.
Stewart Gordon M.
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