Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid
Reexamination Certificate
1998-11-16
2002-11-05
Siew, Jeffrey (Department: 1656)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving nucleic acid
C435S091200, C435S283100, C435S285200, C435S287200, C204S450000, C204S451000, C204S454000, C204S507000, C029S825000, C536S022100, C536S023100, C536S024300, C536S024310, C536S024320, C536S024330
Reexamination Certificate
active
06475722
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to improved nucleic acid processing devices of all types, and more particularly to miniaturized DNA processing devices with surface treatments designed to reduce DNA adsorption to device surfaces exposed to DNA containing mediums.
BACKGROUND OF THE INVENTION
A recent development in the fields of analytical chemistry and biotechnologyhas been the miniaturization of devices and systems for the processing and analysis of DNA. See e.g., McCormick, et al., 1997.
Anal. Chem.
69:2626. Similarly, the current trend towards microfabrication has been driven by efforts to parallel the miniaturization accomplished in the semiconductor industry, and to exploit similar microfabrication techniques. See e.g., Ramsey, et al., 1995.
Nature Med.
1:1093. Justifications for miniaturization include reduced cost, increased speed and reliability, distributed access (point-of-care diagnostics), decreased sample and reagent consumption and reduced waste generation.
An example of a microfabricated DNA analysis device is set forth in PCT Publication WO 96/35810, which is hereby incorporated by reference in its entirely. This aforementioned publication describes electrophoresis devices for the separation and observation of biopolymer fragments in an electrophoretic gel. In one embodiment, an electrophores is device is disclosed which possesses miniaturized electrophores is lanes, which are formed by open channels in a flat plate having dimensions down to approximately 25 &mgr;m and closed by a flat cover plate. In a further embodiment, the publication discloses an electrophores is device which includes an integrally-associated, miniaturized reactor for generating biopolymer fragments for subsequent separation by the device. These miniaturized features are capable of being constructed by various micro-machining techniques, including the lithographic and etching methodologies initially developed in the semiconductor industry.
A further example of a microfabricated DNA analysis device is set forth in the commonly-assigned, U.S. patent application Ser. No. 08/623,346, filed Mar. 27, 1996, which is hereby incorporated by reference in its entirety. This aforementioned application discloses an apparatus for the separation of charged particles in a medium according to the differential diffusion properties of the particles within the electrophoretic medium by use of a spatially- and temporally-varying electric potential. Such an apparatus has application to the separation of single-stranded or double-stranded DNA fragments. In one embodiment, the device consists of a series of miniaturized electrodes which are patterned on a substrate and a cover plate which has one or more miniaturized channels (also down to approximately 25 &mgr;m). This device is also described as being fabricated using the techniques initially developed within the semiconductor industry.
Due to their decreased dimensions the ratio of surface to volume in miniaturized or microfabricated DNA processing devices is markedly increased over other conventional devices. See e.g., Shoffner, et al., 1996.
Nuc. Acids Res.
24:375. This increased surface-to-volume ratio increases the significance of effects of surface chemistry in such microfabricated devices. In particular, it is well known in the art that DNA interacts strongly with and adheres to a number of surfaces. See e.g., Hjerten 1985.
J. Chromatography
347:191. The hydrophilic phosphate groups and hydrophobic protonated bases mean that almost any surface is likely to interact. In addition, the harsh processes used during the standard microfabrication process can damage or contaminate the surfaces creating even stronger interaction forces. See e.g., Henck, 1997.
Tribology Letters
3:239. Although this problem is present in larger scale DNA processing devices, it is considerably exacerbated in micro-machined devices with larger surface to volume ratios. It is also a problem in DNA processing systems such as PCR reactors, capillary and plate gel electrophoresis systems.
Surface interactions have been addressed for a microfabricated polymerase chain reaction (“PCR”) device (see e.g., Shoffner, et al., 1996.
Nuc. Acids Res.
24:375. Additionally, several types of surface treatments were investigated in an initial attempt to find PCR “friendly” surfaces, including surface treatment by silanization followed by a polymer treatment, by stoichiometric silicon nitride coating, and by silicon oxide coating. It should be noted, however, that only silicon oxide was demonstrated not to inhibit the PCR reaction; whereas the inhibition of the PCR amplification reaction by the other treatments methodologies was presumed to have been the result of surface binding sites that non-specifically adsorbed molecules involved in the PCR reaction (see e.g., Cheng, 1996.
Nuc. Acids Res.
24:380.
Accordingly, it is apparent that there is a need for surface treatments for surfaces created in micro-machined DNA processing devices that inhibit DNA surface adsorption. Such an inhibition is termed herein surface “passivation.”
It should be noted that citation of references herein is not to be taken as an admission that such references are prior art to the instant invention.
SUMMARY OF THE INVENTION
The present invention discloses the finding that certain surface treatments possess the ability to reduce DNA adsorption. For example, certain surface treatments are based upon: (i) plasma-enhanced, low temperature deposition of silicon oxide or (ii) low pressure chemical vapor deposition (hereinafter designated “LPCVD”) of low temperature silicon oxide. In particular, the present invention discloses conditions (including precursors), deposition process conditions and subsequent process conditions, which provide for minimal adherence of DNA to a treated surface. Similarly, other surface treatments of the present invention are based upon LPCVD deposition of silicon nitride. In particular, the present invention discloses the finding that certain levels of silicon-enrichment possess novel and highly efficacious properties. Furthermore, the present invention discloses the finding that surface treatments, based upon low pH wash solutions, also markedly reduce DNA adsorption. These aforementioned treatments, as discloses herein, have been adapted to microfabrication processes which have, heretofore, only been utilized in the fabrication of miniaturized devices, primarily in the electronics industry.
Accordingly, one embodiment of the present invention discloses methodologies for the administration of these surface treatments to various types of DNA analysis devices. Another embodiment of the present invention discloses devices made by these methodologies which may be applied, for example, to the analysis of nucleic acids. In a preferred embodiment, the devices are improved DNA processing devices possessing surfaces to which have been administered the surface treatments and washes of the present invention. As these treatments reduce DNA adsorption, such improved devices may be advantageously further miniaturized with an attendant increase in the overall surface to volume ratios.
In a further embodiment, the present invention includes methods for assaying the extent of DNA adsorption to untreated and treated surfaces. These methods include, but are not limited to, washing the surfaces with fluorescently-labeled DNA, rinsing, and fluorescence detection of adsorbed DNA by use of a spectrofluorometry.
The instant invention also may be applied to both microfabricated and to larger scale DNA systems and devices. Such systems and devices may perform processing functions including, but not limited to: (i) DNA analysis (e.g., sequencing, separation, hybridization, electrophoresis; (ii) DNA processing (e.g., DNA replication, polymerase chain reaction (“PCR”), Reverse Transcription-basedPCR (RT-PCR), ligase chain reaction (“LCR”), in vitro transcription and translation, strand exchange with or without enzymes); (iii) DNA modifications (e.g., end- or internal-labeling,phosphorylation, de-phos
CuraGen Corporation
Elrifi Ivor R.
Mintz Levin Cohn Ferris Glovsky and Popeo P.C.
Siew Jeffrey
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