Chemistry: electrical and wave energy – Apparatus – Electrolytic
Reexamination Certificate
2002-01-30
2004-06-15
Tung, T. (Department: 1743)
Chemistry: electrical and wave energy
Apparatus
Electrolytic
C204S416000, C204S286100, C435S285200
Reexamination Certificate
active
06749731
ABSTRACT:
TECHNICAL FIELD
The present invention relates to a gene detection chip and detection device capable of detecting and analyzing gene base sequences as well as gene abnormalities such as genetic DNA single base substitution SNPs (single nucleotide polymorphisms: mutations of human genetic code), multiple base substitutions, point mutations, and genetic defects.
BACKGROUND ART
Methods in which probe DNA is immobilized on an electrode, the probe DNA is hybridized with sample DNA, and the resulting hybrid is electrochemically detected have been proposed as a means of detecting the base sequence of gene DNA (see Japanese Patent Laid-open No. 9-288080 and Proceedings of the 57
th
Meeting of the Japan Society of Analytical Chemistry, pp. 137-138, 1996) These methods allow such hybrids to be detected with high sensitivity.
There exist, however, enormous numbers of such single base genetic substitution SNPs, genetic mutations, and the like, so at least 2,000,000 single base genetic substitution SNPs must be identified in order, for example, to map such single base substitution SNPs with a density (resolution) of 15 kB in humans. Genetic point mutations related to existing disorders also exist in large numbers. A means that can be used to comprehensively analyze such single base substitutions or point mutations.
The present invention, which was perfected in view of this situation, provides a gene detection chip and detection device that allow large amounts of genes to be detected (that is, processed with a high throughput) and allow detection and analysis procedures to be carried out with high sensitivity.
BRIEF SUMMARY OF THE INVENTION
The gene detection device of the present invention comprises a plurality of pins that constitute measurement electrodes, and a common electrode that constitutes a counter electrode for these measurement electrodes. At least portions of the surfaces of the abovementioned pins are coated with a resin. Here, a so-called detection chip is included in the meaning of the term “detection device”. Furthermore, a device constructed by attaching a detection chip to a measuring device is also included in the meaning of the term “detection device”.
In the above arrangement, the pins may be configured such that the resin is applied only to part of the surface on which the Au film is formed. Covering only part of the outermost surface of the pins with resin makes it possible to immobilize a gene solely on the uncoated, exposed portions and allows the exposed surface area to be kept constant and the amount of immobilized probe to be controlled, producing detection results that have higher sensitivity. In addition, an arrangement in which the lateral surface of each pin is covered with resin and the gene is immobilized solely on the exposed portion (located at the end opposite from the base end of the pin fixedly supported on the supporting member) is preferred because the exposed surface area can be readily determined based on the thickness of the pin electrodes. The coating resin should be PEEK (polyether ether ketone), a fluororesin or other thermoplastic resin, or an epoxy resin because of considerations related to chemical resistance.
In the above, the abovementioned pins may contact the surface of a supporting member or may be implanted in the surface of a supporting member, and the lateral surfaces of these pins and the portions of the surface of the abovementioned supporting member that are not contacted by these pins or in which these pins are not implanted may be respectively coated with a resin. In this case, the abovementioned resin is preferably a fluororesin. In particular, a copolymer of tetrafluoroethylene and hexafluoropropylene is especially desirable.
In the above, the device may comprise a plate-form member which consists of a resin and which has a plurality of pin holes into which the abovementioned plurality of pins are respectively inserted, and portions of the surfaces of the abovementioned pins may be coated by this plate-form member.
According to the present invention, a plurality of pins can be secured easily and stably because the pins are inserted into and held in place inside the pin holes with the aid of a plate-form member provided with a plurality of pin holes at positions that match the pin positions.
Each pin can be tightly fitted into the corresponding pin hole when inserted thereinto as a result of the fact that the pin hole diameter is the same as or slightly smaller than the outside diameter of the pin. In other words, each pin can be held airtightly by the plate-form member, preventing the solutions used in the detection process from penetrating through the contact areas between the pins and pin holes.
The present invention provides a gene detection device characterized in that the diameters of the pin holes taper off in the direction in which the pins are inserted, and the pins are held in place in the narrowest sections of the pin holes.
According to the present invention, the pin hole diameters vary in tapered fashion, and the pins are inserted from the side with the large diameter and are held in place in the section with the minimum hole diameter, thus facilitating positioning when the pins are inserted into the pin holes and making it easier to conduct operations in which the tabular member is mounted on the pins. Another feature is that because the minimum hole diameter is the same as or slightly less than the pin diameter, the pins are held airtightly by the plate-form member when the pins are inserted into the pin holes, and the solutions used in the detection process can be prevented from penetrating through the contact areas between the pins and pin holes.
It is desirable that the abovementioned pins contact the surface of the supporting member or be implanted in the surface of the supporting member, and that the abovementioned plate-form member adhere tightly to the surface of the abovementioned supporting member.
According to the present invention, the pins are held in place while the plate-form member is tightly bonded to the supporting member, making it possible to prevent the solutions used in the detection process from penetrating through the joint between the plate-form member and the supporting member. The supporting member may also be a circuit substrate whose interior contains electric circuitry.
The plate-form member should preferably contain a thermoplastic resin as the principal component thereof. Specifically, PTFE (polytetrafluoroethylene) and other fluororesins can be cited as examples of suitable materials, as can PEEK (polyether ether ketone). Using a plate-form member composed of a thermoplastic resin makes it possible to provide a gene detection chip that has excellent heat resistance and is highly resistant to the chemicals (alkalis, acids, and the like) used for detection pretreatments. Such enhanced chemical and heat resistance makes this material suitable for processes in which a detection cycle is followed by the removal of sample DNA and the re-hybridization of another sample DNA to allow the gene detection chip to be reused, or for processes in which a probe gene is removed and another probe gene is re-immobilized on the pins to allow the gene detection chip to be reused.
It is also possible to fabricate the present detection device by a process in which the heat-shrinkage properties of a thermoplastic resin (for example, PEEK) are utilized to form a tabular member whose pin holes have somewhat enlarged diameters, pins are inserted into the pin holes, the plate-form member is heat-treated to induce heat shrinkage, and the diameters of the pin holes are reduced to cause the pin holes to constrict the pins. Adopting this arrangement facilitates positioning when pins are inserted into the pin holes because the diameters of the pin holes are greater than the outside diameters of the pins, makes it easier to mount the plate-form member on the pins, and allows the pins to be held airtightly in a state in which the pins and the pin holes are tightly joined with each other.
A particularly preferred f
Kobori Shinichi
Miyahara Hiroyoshi
Nakama Kazuhiro
Satoh Shingo
Covington & Burling
Kyocera Corporation
Noguerola Alex
Tung T.
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