| PATENT NUMBER | This data is not available for free |
| PATENT GRANT DATE | April 2, 2002 |
| PATENT TITLE |
Apparatus and methods for arraying solution onto a solid support |
| PATENT ABSTRACT |
A method for depositing biomolecule onto a solid support, the method including the steps of: immersing a tip of a spring probe into a solution of biomolecule; removing said tip from said solution to provide biomolecule solution adhered to said tip; and contacting said biomolecule solution with a solid support to thereby transfer biomolecule solution from said tip to said solid support. The spring probe has a planar tip but it otherwise identical to commercial spring probes. The solution of biomolecule contains a thickening agent in addition to biomolecule, where oligonucleotide is a preferred biomolecule. |
| PATENT INVENTORS | This data is not available for free |
| PATENT ASSIGNEE | This data is not available for free |
| PATENT FILE DATE | July 21, 1998 |
| PATENT REFERENCES CITED |
Blanchard et al., (1996), "High-density oligonucleoyide arrays," Biosens. Bioelectron. 11: 687-690. Chee et al., (1996), "Accessing genetic information with high-density DNA arrays," Science 274: 610-614. Chu et al., (1998), "The transcriptional program of sporulation in budding yeast," Science 282, 699-705. Cronin et al., (1996), "Cystic Fibrosis Mutation Detection by Hybridization to Light-Generated DNA Probe Arrays," Human Mutation 7:244-255. DeRisi et al., (1996), "Use of a cDNA microarray to analyze gene expression patterns in human cancer," Nat Genet 14: 457-460. DeRisi et al., (1997), "Exploring the metabolic and genetic control of gene expression on a genomic scale," Science 278: 680-686. de Saizieu et al., (1998), "Bacterial transcript imaging by hybridization of total RNA to oligonucleotide arrays," Nature Biotech. 16: 45-48. Drmanac et al., (1998), "Accurate sequencing by hybridization for DNA diagnostics and individual genomics," Nature Biotech. 16: 54-58. Fodor et al., (1991), "Light-directed, spatially addressable parallel chemical synthesis," Science 251: 767-773. Hacia et al., (1996), "Detection of heterozygous mutations in BRCA1 using high density oligonucleotide arrays and two-colour fluorescence analysis," Nature Genet. 14: 441-447. Heller et al., (1997), "Discovery and analysis of inflammatory disease-related genes using cDNA microarrays," Proc Natl Acad Sci USA. 94:2150-2155. Khrapko et al., (1991), "Hybridization of DNA with oligonucleotides immobilized in gel: a convenient method for detecting single base substitutions," Molecular Biology 25: 581-591. Kozal et al., (1996), "Extensive polymorphisms observed in HIV-1 clade B protease gene using high-density oligonucleotide arrays," Nature Med. 2: 753-759. Lashkari et al., (1997), "Yeast microarrays for genome wide parallel genetic and gene expression analysis," Proc. Natl. Acad. Sci. USA 94: 13057-13062. Lemieux et al., (1998) "Overview of DNA Chip Technology," Molecular Breeding 4: 277-289. Lockhart et al., (1996), "Expression Monitoring by Hybridization to High-Density Oligonucleotide Arrays," Nature Biotechnology 14: 1675-1680. Maier et al., (1994), "Application of robotic technology to automated sequence fingerprint analysis by oligonucleotide hybridisation," Summary. Pease et al., (1994), "Light-generated oligonucleotide arrays for rapid DNA sequence analysis," Proc. Natl. Acad. Sci. USA 91: 5022-5026. Sapolsky and Lipshutz, (1996), "Mapping Genomic Library Clones Using Oligonucleotide Arrays," Genomics 33: 445-456. Schena et al., (1995), "Quantitative monitoring of gene expression patterns with a complementary DNA microarray," Science 270:467-470. Schena, M., (1996), "Genome Analysis with Gene Expression Microarrays," BioEssays 18: 427-431. Schena et al., (1996), "Parallel human genome analysis: microarray-based expression montitoring of 1000 gene," Proc Natl Acad Sci USA93: 10614-10619. Schena et al., (1998), "Microarrays: Biotechnology's discovery platform for functional genomics," Trends Biotech. 16: 301-306. Schena and Davis, (1998), "Parallel Analysis with Biological Chips. in PCR Methods Manual," Academic Press (San Diego), in press. Shalon et al., (1996), "A DNA micro-array system for analyzing complex DNA samples using two-color fluorescent probe hybridization," Genome Research 6: 639-645. Shoemaker et al., (1996), "Quantitative phenotypic analysis of yeast deletion mutants using a highly parallel molecular bar-coding strategy," Nature Genetics 14: 450-456. Wodicka et al., (1997), "Genome-wide expression monitoring in Saccharomyces cerevisiae," Nature Biotech. 15: 1359-1367. Yershov et al. (1996), "DNA analysis and diagnostics on oligonucleotide microchips," Proc. Natl. Acad. Sci. USA 93: 4913-4918. http://cmgm.stanford.edu/pbrown/mguide, Sep. 12, 2000 and http://cmgm.stanford.edu/pbrown/mguide/tips.html, Sep. 12, 2000. Schummer et al., "Inexpensive Handheld Device for the Construction of High-Density Nucleic Acid Arrays," Biotechniques 23(6): 1087-1092, 1997. Hamilton Microlab Arrayer, modular print head, http://chroma.mbt.washington.edu/mod.sub.- www/array.html.[Accessed Nov. 19, 1998]. |
| PATENT PARENT CASE TEXT | This data is not available for free |
| PATENT CLAIMS |
What is claimed is: 1. A spring probe comprising a tubular housing encasing a compression spring, said spring in mechanical communication with a plunger, said plunger having a first region extending out of said housing, said first region comprising a cone-shaped fluted tip terminating as a flat surface, said surface perpendicular to a longitudinal axis of said housing, said cone-shaped tip having in cross-section two exterior sides adjacent said surface which, if said sides extended past said surface, would meet at a point positioned a distance of about 0.00001-0.010 inches beyond said surface. 2. The spring probe of claim 1 where the cone-shaped fluted tip comprises a gold surface. 3. The spring probe of claim 1 wherein said cone-shaped tip has in cross-section two exterior sides adjacent said surface which, if said sides extended past said surface, would meet at a point positioned a distance of about 0.001-0.005 inches beyond said surface. 4. The spring probe of claim 1 wherein said first region comprises a plurality of cone-shaped flutes, the flutes separated by vanes converging toward said flat surface at an angle between approximately 15.degree. and 120.degree.. 5. The spring probe of claim 1 wherein said first region comprises a plurality of cone-shaped flutes, the flutes separated by vanes converging toward said flat surface at an angle between approximately 60.degree. and 90.degree.. 6. A composition comprising a thickening agent at a concentration of about 35 vol % to about 80 vol % based on the total volume of the composition, an oligonucleotide at a concentration ranging from 0.001 .mu.g/mL to 10 .mu.g/mL, and water. 7. The composition of claim 6 wherein the thickening agent is a polyhydric alcohol having at least three hydroxyl groups. 8. The composition of claim 7 wherein the polyhydric alcohol is selected from the group consisting of glycerol, trimethylolpropane, trimethylolethane, pentaerythritol, and saccharides. 9. The composition of claim 8 wherein the saccharide is selected from the group consisting of mannitol, sucrose, fructose, lactose, cellulose and corn syrup. 10. The composition of claim 7 wherein the oligonucleotide is at a concentration of 0.05 .mu.g/mL to 0.5 .mu.g/mL. 11. The composition of claim 6 wherein the thickening agent is glycerol present at a concentration of 40 vol % to 60 vol %. 12. The composition of claim 6 wherein the oligonucleotide is at a concentration ranging from 0.01 .mu.g/mL to 1 .mu.g/mL. 13. The composition of claim 6 wherein the oligonucleotide comprises 15 to 50 nucleotides. 14. The composition of claim 6 wherein the oligonucleotide comprises 50 to 1,000 nucleotides. 15. The composition of claim 6 wherein the oligonucleotide is single stranded. 16. The composition of claim 6 wherein the oligonucleotide is duplex. 17. The composition of claim 6 wherein the oligonucleotide has an amino (--NH.sub.2) group at a 5' end of the oligonucleotide. 18. The composition of claim 17 wherein the oligonucleotide has a hexylamine (--(CH.sub.2).sub.6 --NH.sub.2) group at a 5' end of the oligonucleotide. 19. The composition of claim 17 further comprising trichlorotriazine. 20. The composition of claim 6 having a pH of 7 to 9 and further comprising a buffering agent. 21. The composition of claim 20 wherein the buffering agent is selected from the group consisting of sodium phosphate, sodium borate, sodium carbonate and Tris HCl. 22. The composition of claim 6 having a temperature of 18-25.degree. C. 23. The composition of claim 6 having a viscosity at 20.degree. C. of about 6 to 80 centipoise at 25.degree. C. -------------------------------------------------------------------------------- |
| PATENT DESCRIPTION |
TECHNICAL FIELD This invention relates to microfabrication technology, such as DNA chip-making technology, and more specifically to methods and apparatuses for delivering controlled amounts of a solution to specific, closely spaced locations on a solid support. BACKGROUND OF THE INVENTION In the fields of molecular biology and microbiology it has long been common in the art to make replicate arrays of biological agents to facilitate parallel testing of many samples. For example, the use of sterile velvet cloths and a piston-ring apparatus has long been used to make replicate agar plates of bacterial and yeast colonies on many plates, each containing a different growth medium, as a way of rapidly screening a large number of independent colonies for different growth phenotypes (Lederberg and Lederberg, J. Bacteriol. 63:399, 1952). Likewise, 96-well microtiter plates have long been used to store, in an organized and easily accessed fashion, large numbers of cell lines and virus isolates representing recombinant DNA libraries or monoclonal antibody cell lines. Experimental screening of the 96-well microtiter plates housing a clone collection is commonly accomplished by using a rigid metal or plastic 96-pin device designed so that each pin is spaced relative to the others such that it fits precisely into the microtiter plate. Depending on the task at hand, the 96-pin device is lowered carefully to the surface of an nutrient-agar plate (if the objective was to grow replicate biological samples), into another microtiter plate (to grow or dilute the samples), onto nylon embranes (for molecular screening by DNA or RNA hybridization to identify a particular recombinant clone), or transferred for use in any other screening or procedure that is adaptable to the 96-well microtiter dish format. While multiple prints may be performed from one pin dip into the samples arrayed in the master microtiter dish, the amount of sample deposited during each sequential print drops off. The ability to control the uptake of a solution onto the printing pin, and the deposition of solution onto a printing surface are critical to realizing an aliquotting devise which meets the technical needs of microarray production for the fields of genomics, molecular biology and molecular diagnostics. An important factor in developing a successful printing process is the ability to control the force and speed of movement with which the pin tips contacts the surface being printing upon. As noted by Drmanac and Drmanac (BioTechniques 17:328, 335, 1994), two problems with conventional flat-cylinder pins are that drops can be caught on the sides of a pin leading to irregular printing, and drop splashing can occur when the printing pin head is withdrawn too fast from the printing surface. Too much force can lead to extensive damage to the print surface negating the utility of that print array. Too little force may be just as disabling in that variable amounts of sample may be transferred, or the print maybe defective all together. For example, when printing bacterial or viral samples to the surface of a nutrient-agar plate, too much pressure results in disruption of the agar surface, while too little force may result in little or no transfer of a sample. In addition, many nucleic acid hybridization membrane surfaces are fragile and are easily damaged by excess pin head force during sample printing. The advent of large scale genomic projects and the increasing medical use of molecular diagnostics, has prompted the development of large volume throughput methods for screening recombinant DNA libraries representing entire genomes, the performance of large scale DNA sequencing projects, and executing replicative immunological assays, nucleic acid hybridization assays, or polymerase chain reaction assays. The following publications (and the references cited therein), which are exemplary only, provide general and specific overviews of large throughput methods that rely on biomolecular arrays, as well as methods of preparing such arrays: Eggers, M. D. et al. Advances in DNA Sequencing Technology SPIE Vol. 1891:113-126, 1993; Chetverin, A. B. et al. Bio/Technology 12:1093-1099, 1994; Southern, E. M. Nucleic Acids Research 22:1368-1373, 1994; Lipshutz, R. J. et al. BioTechniques 19:442-447, 1995; Schena, M. BioEssays 18:427-431, 1996; Blanchard, A. P. et al. Biosensors & Bioelectronics 11:687-690, 1996; O'Donnell-Maloney, M. J. et al. Genetic Analysis: Biomolecular Engineering 13:151-157, 1996; Regalado, A. Start-Up 24-30, October 1996; and Stipp, D. Fortune pp. 30-41, Mar. 31, 1997. The need for high throughput methodology has led, in some cases, to a change from a 96-well microtiter dish format, to a 384-well (Maier et al., J. Biotechnology 35:191, 1994) or 864-well (Drmanac et al., Electrophoresis 13:120, 1992) format, which can also be used in conjunction with robotic devises (see, e.g., Belgrader et al., BioTechniques 19:426, 1995; Wilke et al., Diagnostic Microbiology and Infect. Disease 21:181, 1995). However, all of these automated techniques require the use of a robotic pin-tool devise that is capable of reproducibly transferring equal volumes of liquid from one arrayed configuration (i.e., 96-well microtiter plate) to another (i.e., 96-spot array on a hybridization filter membrane). Recently, methods have also been developed to synthesize large arrays of short oligodeoxynucleotides (ODNs) bound to a glass surface that represent all, or a subset of all, possible nucleotide sequences (Maskos and Southern, Nucl. Acids Res. 20: 1675, 1992). Once such an ODN array has been made may be used to perform DNA sequencing by hybridization (Southern et al., Genomics 13:1008, 1992; Drmanac et al., Science 260:1649, 1993). The utility of this method of DNA sequencing would be greatly improved if better methods existed for the transfer and arraying of the precise amounts of the biochemical reagents required for the synthesis of large sets ODNs bound to hybridizable surfaces. This would enable greater equality of ODN yield at each position within the array and also increase the nucleotide chain length it is possible to synthesize. The polymerase chain reaction (PCR) has found wide application to many different biological problems. Two major limitations to the commercial utilization of PCR are the high cost of the reagents and the inability to automate the performance of the process. Reagent costs can be lowered if the total volume of each reaction can be decreased, allowing a concomitant decrease in DNA polymerase and nucleotides. An accurate and reliable means to array small volumes of reagents using a robotically controlled pin tool could help solve both of these PCR problems. As noted above, transfer devices have been in use for some time in the fields of microbiology and molecular biology. The types of devises which have been used can be roughly divided into two categories. Pressure devises (e.g., pumps and automatic pipettes), driven by positive and/or negative pressure, which deliver fixed aliquots of liquids sample via a pipette tip to a solid surface or into a microtiter well. Pipette arrays have been constructed that correspond to the standard 96-well microtiter dish format (Reek et al., BioTechniques 19:282, 1995). These devices are most accurate in the 5 .mu.l and above volume range, but are generally ill-suited to smaller volume tasks. Solid surface pin devises transfer liquids based upon pin surface area and the factors regulating liquid surface tension, and have been widely adopted because of their simplicity and ability to transfer small volumes of liquid. These rigid pin devises have been used for several years in robotic devises to print multiple copies of nucleic acid microdot arrays which are then used in hybridization reactions to measure gene expression. Researchers have modified the traditional rigid microarray printing tip so that it contains a micro-channel which functions by capillary action to collect and hold liquid for subsequent printing to a glass surface (Schena et al., Science 270:467, 1995; Schena, BioEssays 18.427, 1996; Shalon et al., Genome Res. 6:639, 1996). Such a print head has been used to print PCR amplified cDNA inserts into micro-arrays using a robotic system. Small volume (2 .mu.l per microdot) hybridization reactions were performed using this system to measure the differential expression of 45 genes by means of simultaneous, two color fluorescence hybridization (Schena et al., (Science 270:467, 1995). There is a need in the art for highly efficient, cost effective means for arraying oligonucleotides and other biomolecules on a planar solid support. The present invention provides these and related advantages as disclosed in more detail herein. SUMMARY OF THE INVENTION In one aspect, the invention provides a spring probe comprising a tubular housing encasing a compression spring. The spring is in mechanical communication with a plunger. The plunger has a first region extending out of the housing, where the first region comprises a cone-shaped tip terminating in a flat surface. The flat surface is perpendicular to a longitudinal axis of the housing. The cone-shaped tip has, in cross-section, two exterior sides adjacent to the surface which, if the sides extended past the surface, would meet at a point positioned a distance of about 0.001-0.005 inches beyond the surface. In another aspect, the invention provides a composition including a thickening agent at a concentration of about 35 vol % to about 80 vol % based on the total volume of the composition, an oligonucleotide at a concentration ranging from 0.001 .mu.g/mL to 10 .mu.g/mL, and water. In another aspect, the invention provides a method for depositing a biomolecule onto a solid support. The method includes the steps of: immersing a tip of a spring probe into a solution of biomolecule; removing the tip from the solution to provide biomolecule solution adhered to the tip; and contacting the biomolecule solution with a solid support to thereby transfer biomolecule solution from the tip to the solid support. The spring probe used in the depositing includes a tubular housing encasing a compression spring, as described above. In another aspect, the invention provides a method for arraying a biomolecule. The method includes the steps of: immersing a tip of a spring probe into a solution of biomolecule; removing the tip from the solution to provide biomolecule solution adhered to the tip; contacting the biomolecule solution with a solid support to thereby transfer biomolecule solution from the tip to the solid support; and repeating the contacting step a plurality of times to provide biomolecule patterned in an array on the solid support. Again, the spring probe having a tubular casing is as described above. Other aspects of this invention will become apparent upon reference to the attached Figures and the following detailed description. |
| PATENT EXAMPLES | This data is not available for free |
| PATENT PHOTOCOPY | Available on request |
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