JP2003035711A - Probe carrier as well as method and apparatus for manufacturing the same - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method of manufacturing a probe carrier capable of dealing with various probe pitches without replacing an ink jet-type liquid discharge head (a block) for different probe pitches, to provide an apparatus used for the method and to provide the probe carrier manufactured by the method. SOLUTION: While a liquid discharge head comprising a plurality of discharge nozzles is being scanned relatively on the carrier in a direction not at right angles to its scanning direction, a probe solution is discharged onto the carrier from the discharge nozzles, and a probe is given.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method of manufacturing a probe carrier in which a probe is carried on a carrier by using an inkjet method, a probe carrier manufacturing apparatus used in the manufacturing method, and the probe carrier. In particular, a liquid ejection head having a plurality of ejection nozzles in a direction substantially orthogonal to the scanning direction ejects the probe solution while scanning the carrier on the carrier relatively to the carrier. The present invention relates to a method for manufacturing a reliable probe carrier, a probe carrier manufacturing apparatus used in the manufacturing method, and the probe carrier.

[0002]

2. Description of the Related Art When analyzing the base sequence of a gene DNA, or simultaneously performing highly reliable gene diagnosis on many items, a DNA having a target base sequence is selected using plural kinds of probes. Is required. DNA microchips have been attracting attention as a means for providing a plurality of types of probes used for this sorting operation. Also,
Even in high-throughput screening of drugs and combinatorial chemistry, many solutions of target proteins and drugs (eg, 96, 384, 1
It is necessary to arrange (536 kinds) and perform an orderly screening. Controls a method for arranging multiple kinds of drugs for that purpose, an automated screening technique in that state, a dedicated device, a series of screening operations,
Also, software and the like for statistically processing the results have been developed.

These parallel screening operations basically consist of a large number of known probes as a means for selecting substances to be evaluated, so-called probe.
By using an array, it is possible to detect the presence or absence of an action or reaction on the probe under the same conditions. In general, what kind of action and reaction to use for the probe is determined in advance. Therefore, the probe species mounted in one probe array may be, for example, a group of DNA probes having different base sequences. It is one type of substance when roughly classified. That is, the substance used for the group of probes is, for example, DNA, protein, or a synthesized chemical substance (drug). In many cases, a probe array composed of a plurality of types of probes is often used. However, depending on the nature of the screening work, as probes, DNA having the same base sequence, protein having the same amino acid sequence, and the same chemical It is also possible to use an array form in which a large number of substances are arranged. These are mainly used for drug screening and the like.

In a probe array consisting of a plurality of types of probes, specifically, a group of DNAs having different base sequences, a group of proteins having different amino acid sequences, or a group of different chemical substances is used. In many cases, a plurality of types of constituents are arranged in an array on a carrier according to a predetermined arrangement order.
Among them, the DNA probe array is used for analysis of the base sequence of gene DNA and, at the same time, for highly reliable gene diagnosis of many items.

One of the problems in a probe array consisting of a plurality of types of probes forming this group is to use as many types of probes as possible, for example, D having various types of base sequences.
The NA probe is placed on one carrier. In other words, how densely the probes can be arranged in an array.

[0006] As one method of immobilizing plural kinds of probes in an array on a carrier, US Pat. No. 5,424,424 has been used.
No. 186, a method for producing DNA probes having different base sequences in an array form by sequential extension reaction of DNA on a carrier using a photodegradable protecting group and photolithography. You can Using this approach, for example, 1 cm ² per 1000
It is also possible to prepare a DNA probe array carrying zero or more kinds of DNAs having different sequences. In this method, when synthesizing DNA by a sequential extension reaction, a photolithography process is performed for each of four types of bases (A, T, C, G) by using a dedicated photomask, and a predetermined array is prepared. By selectively extending one of the bases at a position, a plurality of types of DNA having a desired base sequence are synthesized on the carrier with a predetermined sequence. Therefore, the longer the chain length of DNA, the higher the cost required for the preparation and the longer time required. In addition, since the efficiency of nucleotide synthesis in each extension step is not 100%, the ratio of DNA in which the designed nucleotide sequence has a defect is not small. Furthermore, in the case of using a photodegradable protective group in the synthesis, the synthesis efficiency is lower than in the case of using an ordinary acid-decomposable protective group,
In the finally obtained array, there is also a problem that the proportion of DNA according to the designed nucleotide sequence becomes small.

Further, since the product directly synthesized on the carrier is used as it is, it is, of course, impossible to remove the DNA having the defective base sequence from the designed DNA according to the designed base sequence by purification fractionation. . Other,
In the finally obtained array, there is a problem that the base sequence of DNA synthesized on the carrier cannot be confirmed. This is because if a given base was hardly extended at a certain extension stage due to an error in the process, and it was a completely defective product, the screening using this defective product probe array resulted in an incorrect result. However, it means that there is no way to prevent it. The inability to confirm this nucleotide sequence is the biggest and essential problem in this method.

As a method different from the above-mentioned method, DNA for a probe is previously synthesized and purified, and in some cases, after confirming the base length thereof, each DNA is applied onto a carrier by a device such as a microdispenser. And probe
Techniques for preparing arrays have also been proposed. PCT publication WO95 / 355O5 describes a method of applying DNA onto a membrane using a capillary. When this method is applied, it is possible in principle to prepare about 1000 DNA arrays per cm ² . Basically, one capillary-type dispensing device for each probe is used to apply the probe solution to a predetermined position on the carrier, and this operation is repeated,
This is a method of preparing an array. There is no problem if you prepare a dedicated capillary for each probe, but if you try to do the same work with a small number of capillaries, to prevent cross-contamination, the capillaries should be sufficient when replacing the probe type. Need to be washed. Further, it is necessary to control the applied position each time. Therefore, it cannot be said to be a method suitable for preparing an array in which a large number of types of probes are arranged at high density. In addition, since the probe solution is applied to the carrier by tapping the tip of the capillary onto the carrier, reproducibility and reliability cannot be said to be perfect.

Another method is to use DN on a carrier.
When performing solid phase synthesis of A, a method has also been proposed in which a solution of a substance required for synthesis is applied onto a carrier by an inkjet method at each extension step. For example, European Patent Publication EP
No. 0 703 825 B1 discloses nucleotide monomers utilized in solid phase synthesis of DNA, and
A method for solid phase synthesis of a plurality of DNA species each having a predetermined nucleotide sequence by applying activators from different piezo jet nozzles is described. The application (application) by the ink jet method is more reliable than the application (application) of the solution using the capillary, such as the reproducibility of the applied amount, and the structure of the nozzle can be miniaturized. It has characteristics suitable for increasing the density of probe arrays. However, since this method also basically applies the sequential elongation reaction of DNA on a carrier, the above-mentioned US Pat.
The biggest problem in the method described in Japanese Patent No. 424,186, that is, the problem that the base sequence of DNA synthesized on the carrier cannot be confirmed still remains. Although the complexity of performing a photolithography process using a dedicated mask for each extension step is eliminated, the point that a predetermined probe is fixed at each point, which is an essential requirement for the probe array. , With some problems. The EP
No. 0,703,825 B1 describes only a method of using a plurality of piezo jet nozzles formed independently, and when using this small number of nozzles, a method using the above-mentioned capillary is used. Similarly, it is not always suitable for high density probe array preparation.

Further, Japanese Patent Application Laid-Open No. 11-187900 discloses a method in which a liquid containing a probe is attached as droplets to a solid phase by a thermal ink jet head to form a spot containing the probe on the solid phase. But
Since the inkjet head used is a head for a general printer, it cannot be said that the structure is optimal for preparing a probe array. This point will be described in detail below.

The conventional ink jet head is a method developed for printing characters and images. Therefore, the solution used is one color (black) ink for monochrome (generally black) printing, and generally for color printing,
Ink of three primary colors, namely, yellow (Y), cyan (C), and magenta (M). In the case of color printing, black or Y, M, C,
The dark and light inks may be used, but at most 10 or more kinds of inks are not used.

Further, since a large amount of ink is used for printing on paper, a conventional ink jet printing head has a tank (reservoir) for filling ink having a sufficient capacity, and ink for nozzles. A flow path for guiding and a nozzle for ejecting ink are provided.

On the other hand, as described above, it is desired that the ink jet head for preparing the probe array ejects as many kinds of liquids as possible.
A head having a plurality of nozzles is usually used in the manufacture of a probe array carried on a probe carrier, but a head having a same number of one-to-one probe solution reservoirs as the plurality of nozzles is desirable.

Further, in the liquid jet head nozzle of the ink jet system used for manufacturing the probe carrier, since it is necessary to jet the probe solution containing one kind of probe from one nozzle, the nozzle corresponding to at least the probe pitch. Need to pitch. Therefore, in order to manufacture a high-density probe array,
In measurable probe amount and probe pitch,
By narrowing the nozzle pitch interval of the liquid ejection head as much as possible, it becomes possible to arrange many kinds of probes with high density. As a result, the detection efficiency of diagnosis and the like can be improved, and it is also desirable because a large amount of sample is not prepared.

Further, it is desirable that the probe array carried on the probe carrier has a desired amount fixed at a predetermined position. In order to prevent errors during diagnosis,
It is desirable that only necessary probes are fixed at predetermined places on the probe carrier.

Therefore, in the manufacture of the probe carrier, the nozzle pitch interval of the liquid ejection head is made as narrow as possible within the measurable probe amount and the probe pitch to achieve high density, and stable ejection is performed, and the reliability is high. It is desired to manufacture the probe carrier of

[0017]

In the past, when a probe carrier was manufactured by using an ink jet method, basically, a multi-liquid ejection head having a plurality of nozzles at a constant interval (pitch) was used and the probe was used. Probe array by scanning in the main scanning direction using nozzles matching the pitch to apply the probe to the carrier, then moving the head or carrier in the sub scanning direction, and then repeating ejection in the main scanning direction. Are manufactured. The liquid ejection head used at that time is manufactured by ejecting the inkjet probe solution while scanning the liquid ejection head having a plurality of ejection nozzles in the direction orthogonal to the scanning direction with respect to the carrier.

In such a conventional method of manufacturing a probe carrier, when the nozzle pitch interval of the liquid ejection head block is simply narrowed as described above in order to increase the density of the probe array, The probe solution adhering to the vicinity of the nozzle is liable to cause harmful effects, and it becomes more difficult to remove the adhering droplets. Such attachment of the probe solution inevitably occurs due to attachment of a mist-like probe solution (sometimes called a satellite) generated during ejection. When the attached probe solution reaches the nozzle ejection port edge, it affects the ejection direction of the newly ejected probe solution, causing variations in the desired ejection position on the carrier or landing at different probe ejection positions. Different probes may coexist, and the probe solution adhering to the vicinity of the nozzle may gradually grow and may be mixed with different probe solutions in the vicinity due to vibration or the like. Therefore, in the conventional method, there is a limit in increasing the density of the probe array by narrowing the nozzle pitch interval of the liquid ejection head block. Further, even when using a means for removing the probe-forming solution adsorbed on the ejection surface with a wiper as means for eliminating the mixture of these probe solutions, the solution is excessively absorbed, diffused, and sometimes mixed with each other. Many problems, especially mixed solutions of different probe forming solutions, have not been solved.

On the other hand, in the case of manufacturing a probe array having an arbitrary probe pitch (array arrangement) or in order to cope with a probe carrier having an arbitrary shape, a large number of liquid ejection heads corresponding to them are prepared, High cost because it needs to be replaced. Usually, in order to efficiently manufacture a high-density array, a liquid ejection head block in which a plurality of liquid ejection heads are integrated is used. Therefore, if a large number of these liquid ejection head blocks are prepared and replaced, it is further disadvantageous in terms of cost. . Therefore, there is a problem that it is difficult to secure the productivity of a low-cost, high-density, highly reliable probe carrier.

Therefore, an object of the present invention is to cope with an arbitrary probe pitch without exchanging the liquid ejecting head (block) of the ink jet system for each different probe pitch, and moreover, to enable stable ejection and to improve the probe array. (EN) A method for producing a densified, low-cost, highly reliable densified probe carrier, a probe carrier production apparatus used in the production method, and the probe carrier.

[0021]

In order to solve the above problems, in a method for manufacturing a probe carrier according to an embodiment of the present invention, a liquid ejection head having a plurality of ejection nozzles in a direction not orthogonal to a scanning direction is a carrier. A probe in which a plurality of types of probes capable of specifically binding to a target substance are arranged on the carrier by ejecting a probe solution onto the carrier from the discharge nozzle while relatively scanning the top by an inkjet method. A method of manufacturing a carrier, wherein a probe pitch interval in a direction orthogonal to the scanning direction is a, the ejection nozzle pitch interval of the liquid ejection head is b, and the probe pitch direction and the nozzle pitch direction are formed. When the angle is θ, the liquid ejection head is moved by the angle θ that satisfies | cos θ | = a / b so that a becomes smaller than b. It is preferable to scan tilted with respect to the direction perpendicular to the direction.

In the method of manufacturing a probe carrier according to the present invention, it is preferable that the liquid ejection head scans relative to the carrier by moving the carrier.

Further, in the method of manufacturing a probe carrier according to the present invention, it is preferable that the liquid ejection head scans relative to the carrier by moving the liquid ejection head.

In the probe carrier manufacturing method of the present invention, it is preferable that the liquid discharge head scans relative to the carrier by moving the carrier and the liquid discharge head.

Further, in the method for manufacturing a probe carrier of the present invention, a plurality of liquid ejection heads are used, and the plurality of liquid ejection heads are collectively tilted by the angle θ to perform scanning so that the probe solution is applied to the carrier. It is preferable to discharge.

In the method of manufacturing a probe carrier according to the present invention, the liquid ejection head is a head for ejecting the probe solution by utilizing thermal energy, and is for generating thermal energy to be applied to the probe solution. It is preferable to have a thermal energy generator.

Further, in the method for manufacturing a probe carrier according to the present invention, it has a mechanism for removing droplets of the probe solution adhering to the vicinity of the discharge nozzle by wiping in a direction substantially perpendicular to the arrangement direction of the discharge nozzles. Is preferred.

In the probe carrier manufacturing method of the present invention, it is preferable that the amount of the probe solution discharged from the discharge nozzle is 0.1 picoliter to 100 picoliter.

In the method for producing a probe carrier of the present invention, the area occupied by each of the probe solutions applied from the discharge nozzle is 0.01 μm ^{2 to} 40,000 μ.
It is preferably m ² .

In the method for producing a probe carrier of the present invention, the probe is DNA, RNA, cDNA (complementary DNA), PNA, oligonucleotide, polynucleotide, other nucleic acid, oligopeptide, polypeptide, protein, enzyme. , Substrate for enzyme, antibody, epitope for antibody, antigen, hormone,
Hormone receptor, ligand, ligand receptor,
It is preferably selected from the group consisting of oligosaccharides, polysaccharides, and combinations thereof.

In a probe carrier manufacturing apparatus according to another embodiment of the present invention, a liquid discharge head having a plurality of discharge nozzles in a direction not orthogonal to the scanning direction scans the carrier relatively while moving the liquid discharge head from the discharge nozzles. By ejecting the probe solution onto the carrier by an inkjet method,
An apparatus for manufacturing a probe carrier, in which a plurality of types of probes capable of specifically binding to a target substance are arranged on the carrier, wherein the probe pitch interval in the direction orthogonal to the scanning direction is a, and the liquid ejection head Where b is the discharge nozzle pitch interval and θ is an angle formed by the probe pitch direction and the nozzle pitch direction, an angle θ that satisfies | cos θ | = a / b is set so that a becomes smaller than b. The probe carrier manufacturing apparatus is characterized in that the liquid ejection head is inclined with respect to a direction orthogonal to the scanning direction for scanning.

In the probe carrier manufacturing apparatus of the present invention, it is preferable that the liquid ejection head scans relative to the carrier by moving the carrier.

In the probe carrier manufacturing apparatus of the present invention, it is preferable that the liquid ejection head scans relative to the carrier by moving the liquid ejection head.

In the probe carrier manufacturing apparatus of the present invention, it is preferable that the liquid discharge head scans relative to the carrier by moving the carrier and the liquid discharge head.

In the probe carrier manufacturing apparatus of the present invention, a plurality of liquid discharge heads are used, and the plurality of liquid discharge heads are collectively tilted by the angle θ to perform scanning so that the probe solution can be transferred to the carrier. It is preferable to discharge.

In the probe carrier manufacturing apparatus of the present invention, the liquid ejection head is a head for ejecting the probe solution by utilizing thermal energy, and is for generating thermal energy to be applied to the probe solution. It is preferable to have a thermal energy generator.

In addition, the probe carrier manufacturing apparatus of the present invention has a mechanism for removing droplets of the probe solution adhering to the vicinity of the discharge nozzle by wiping in a direction substantially perpendicular to the arrangement direction of the discharge nozzles. Is preferred.

In the probe carrier manufacturing apparatus of the present invention, the amount of the probe solution discharged from the discharge nozzle is preferably 0.1 picoliter to 100 picoliter.

In the probe carrier manufacturing apparatus of the present invention, the area occupied by each of the probe solutions applied from the discharge nozzle is 0.01 μm ^{2 to} 40,000 μ.
It is preferably m ² .

In the probe carrier manufacturing apparatus of the present invention, the probe is DNA, RNA, cDNA (complementary DNA), PNA, oligonucleotide, polynucleotide, other nucleic acid, oligopeptide, polypeptide, protein, enzyme. , Substrate for enzyme, antibody, epitope for antibody, antigen, hormone,
Hormone receptor, ligand, ligand receptor,
It is preferably selected from the group consisting of oligosaccharides, polysaccharides, and combinations thereof.

Further, a probe carrier according to another embodiment of the present invention is a probe carrier manufactured by the above method for manufacturing a probe carrier.

[0042]

BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, the method for producing the probe carrier of the present invention as described above, the production apparatus that can be used for the method, and the probe carrier produced by the method will be described in more detail.

(Probe Solution) In the present invention, a plurality of types of probes fixed on the carrier surface as independent spots, for example, dot-shaped spots are called “probe carriers”, and many of the probe spots are planar. Those arranged in a matrix, that is, arranged in a two-dimensional array form are called "probe array". This probe carrier contains
It includes a test plate or chip generally called a DNA macro array, a DNA chip, or a probe array.

In the present specification, the probe immobilized on the carrier can specifically bind to a specific labeling substance. In addition, this probe includes an oligonucleotide that can be recognized by a specific target, a polynucleotide,
Alternatively, other polymers are included. The term "probe" refers to a therapy of molecules having a probe function, such as individual polynucleotide molecules, and a population of molecules having the same probe function, such as polynucleotides of the same sequence surface-immobilized in dispersed locations, for a period of time. Also included are molecules called ligands. Probes and targets are often used interchangeably, and probes are ligand-
An anti-ligand (sometimes called a receptor) that is capable of binding to or becoming capable of binding to a target as part of a pair. The probes and targets in the present invention may include bases as found in nature, or analogs thereof.

As an example of the probe supported on the carrier, a part of an oligonucleotide having a base sequence capable of hybridizing with a target nucleic acid has a binding part to the carrier via a linker. And a structure having a structure in which it is connected to the surface of the carrier at the bonding portion with. In the case of such a configuration, the position of the binding part with the carrier in the molecule of the oligonucleotide is not particularly limited as long as it does not impair the desired hybridization reaction.

The probe employed in the probe array to which the method of the present invention is applied is appropriately selected according to the purpose of use, but it is prepared in order to preferably carry out the method of the present invention. In the two-dimensional probe array, the probes are DNA, RNA, cDNA (complementary DNA), PNA, oligonucleotide,
At least selected from polynucleotides, other nucleic acids, oligopeptides, polypeptides, proteins, enzymes, substrates for enzymes, antibodies, epitopes for antibodies, antigens, hormones, hormone receptors, ligands, ligand receptors, oligosaccharides, and polysaccharides It is preferable to have one kind.

The probe preferably has a structure capable of binding to the functional group introduced on the surface of the carrier. After discharging the probe solution and applying the probe solution onto the carrier, the probe is used to bind to the carrier. It is desirable to combine them. By forming such a bond, the probe is firmly bound to the carrier, and the spot of the probe attached on the carrier is stopped at a more limited position, so that contamination with adjacent spots is more reliable. It is possible to prevent it.

The structure of such a probe capable of binding to the surface of the carrier includes amino group, sulfhydryl group, carboxyl group, hydroxyl group, acid halide (haloformyl group; -COX), halide (-X), It has a structure formed by introducing an organic functional group such as aziridine, maleimide group, succinimide, isothiocyanate, sulfonyl chloride (-SO ₂ Cl), aldehyde (formyl group; -CHO), hydrazine and iodoacetamide. Preferably.

On the other hand, the "carrier" is a carrier that carries the above-mentioned probe, and is not particularly limited as long as it holds the probe and does not interfere with the detection of the target substance. can give. In addition, a silicon substrate, a metal substrate, a resin substrate, or the like, or each substrate subjected to an appropriate surface treatment may be used. When a glass substrate is used as a carrier, various methods such as a cleaning method and a surface treatment are known, and the carrier itself is advantageous because it is easily available. As a method of introducing a functional group into the surface of the glass substrate, for example, there is a method of introducing a hydroxyl group, a carboxyl group or the like by various surface treatments and using the functional group as it is. Further, the glass substrate may be treated with a silane coupling agent having various functional groups bonded thereto to utilize the functional groups. As a functional group of a commercially available silane coupling agent, thiol (SH) is used.
Group, an amino group, an isocyanate group, a chloride group, an epoxy group, etc., and a functional group involved in binding to a carrier of a probe or a marker substance, which can be bonded to the functional group of the silane coupling agent, is appropriately selected and used. can do. Since the treatment method with a silane coupling agent is generally well known, a detailed description will not be given, but the silane coupling agent having the above functional group is commercially available from Shin-Etsu Chemical Co., Ltd., Nippon Unicar Co., Ltd. So just use them.

On the surface of the carrier, it is desirable to carry out in advance a structure for reacting with various organic functional groups introduced into the probe to form a covalent bond and a treatment for introducing an organic functional group. The functional group formed on the surface of the carrier is preferably a maleimide group or amino group. Various methods can be used as a method for introducing these functional groups onto the carrier surface. For example, for introducing an epoxy group onto the carrier surface, for example, polyglycidyl methacrylate having an epoxy group is applied to a carrier surface made of a resin. Alternatively, a method in which a silane coupling agent having an epoxy group is applied to the surface of a glass carrier and reacted with glass can be used.

A functional group introduced at the end of the probe,
As a preferred combination with a functional group introduced on the surface of a carrier capable of binding to the functional group, a combination of thiol (-SH) (terminal of nucleic acid probe) and maleimide group (terminal of carrier), amino group (terminal of nucleic acid probe) and epoxy A combination with a group (carrier surface) and the like can be mentioned. In particular, when a functional group is introduced into each so that a covalent bond is formed between the probe and the carrier surface, the carrier surface does not have wells in advance, that is, it is substantially flat and has surface characteristics (for water). Adjacent spots will not be connected even if a carrier having a uniform wettability) is used. As a result, a probe array in which nucleic acid probes are arranged with high precision and high density can be manufactured very efficiently and at low cost.

This does not exclude the use of a carrier having a well on the surface of the present invention.
For example, when a light-impermeable matrix pattern (hereinafter referred to as “black matrix”) is formed in advance between the wells to which the probe solution is applied, the hybridization of the probe and the target substance on the carrier is performed. It is possible to further improve the detection accuracy (SN ratio) in the case of optically detecting the fluorescence (for example, detecting fluorescence). In addition, when a matrix whose surface has a low affinity for the probe solution is provided between the adjacent wells, the probe solution can be smoothly transferred to a desired well even if some displacement occurs in applying the probe solution to the well. Can be given. It is desirable to use a carrier having wells on the surface for the purpose of utilizing such effects.

(Characteristics of Discharge Liquid) When a probe solution containing a probe is applied onto a carrier as described above by an ink jet method, droplets do not unnecessarily spread on the carrier and remain at a predetermined position. It is preferable to prepare a liquid composition. The liquid composition preferably does not impair the original performance of the probe and does not impair the reactivity with the functional group on the surface of the carrier introduced into the probe.

When the inkjet method is used, since the droplets applied to the surface of the carrier are minute, in order to prevent evaporation and drying after application, the carrier is placed in a reaction container such as a moisturizing container during the reaction. It is a preferable method to store the above, but it is also effective to include a moisturizer in the liquid to be ejected. In particular, in the thermal jet method, the presence of the moisturizing component and the surface tension adjusting component is important because the temperature is increased during the ejection. As such a marker substance, or as a solvent that imparts the probe to the carrier surface,
5-10 wt% urea, 5-10 wt% glycerin,
Those containing 5 to 10 wt% of thiodiglycol and 1 wt% of acetylene alcohol can be preferably used.

Considering the ink jet discharge characteristics of the liquid, the stability of the probe in the liquid and the thermal jet (Bubble Jet (registered trademark)) discharge, the probe solution has, for example, 2 mer to 5000 me in the liquid.
r, especially a 2mer to 60mer nucleic acid probe,
It is preferably contained at a concentration of 0.05 to 500 μM, particularly 2 to 50 μM.

As a liquid property of the probe solution, from the viewpoint of ejection property from a thermal jet head (bubble jet head), for example, its viscosity is 1 to 15 cp.
s and the surface tension is preferably 30 dyn / cm or more. The viscosity is 1 to 5 cps and the surface tension is 30 to 50 dyn / c.
When m is set, the landing position on the carrier becomes extremely accurate, and it is particularly preferably used.

A structure such as a well is provided on the surface of the carrier so as to prevent the liquid applied on the carrier by the liquid discharge head from spreading on the carrier and mixing with adjacent spots. In the case where the above is provided, the viscosity and surface tension of the liquid, and the base length and concentration of the nucleic acid probe can be used even if they are out of the above ranges.

(Discharge Mode) The amount of the probe solution discharged from one nozzle at a time is appropriately selected according to the areal density of each probe spot on the probe carrier, but the viscosity of the probe solution, the probe solution It is selected according to the dot size and shape of the array to be formed, taking into consideration various factors such as the affinity of the carrier and the reactivity between the probe and the carrier. In this case, the amount of discharged liquid per time is 0.1 to
100 pl (picoliter), preferably 0.1-50
It can be selected from the range of pl. In order to increase the density of the probe, it is preferable that the amount discharged at one time is as small as possible, but if the amount discharged is extremely small, it will not reach the carrier due to air resistance. The discharge amount per discharge is preferably 0.1 to 10 pl, more preferably 0.1.
It is preferably up to 5 pl, and it is preferable to design and adjust the nozzle diameter and the like according to the liquid amount.

The area occupied by the probe solution applied to one well is 0.01 (for example, 0.1 μm × 0.1 μm).
m) μm ^{2 to} 40,000 (for example, 200 μm × 200 μm
m) μm ² , which is generally determined by the size of the array itself and the density of the array matrix. Therefore, the size and volume of one well can be set by the area occupied by the probe solution. The depth of the well depends on the method of manufacturing the well, but is preferably selected in the range of 0.5 μm to 100 μm. The volume is determined by the area and depth of these wells. As a method for producing such a well, for example, the method described in JP-A-11-099000 can be used as it is.

In order to set the density of the nucleic acid probe on the carrier to the above value (for example, 10,000 or more spots per inch, the upper limit is about 1 × 10 ⁶ spots), the spot diameter of each nucleic acid probe is increased. Is, for example, 20 to 100 μm
It is preferable that the spots are in the order of magnitude and that the spots are independent of each other. Such spots are determined by the characteristics of the liquid ejected from the bubble jet head, the surface characteristics of the carrier to which the liquid adheres, and the like.

The distance between the liquid discharge head and the carrier, more precisely, the distance between the nozzle discharge surface and the carrier surface, is usually 1.
It is ejected close to 2 to 1.5 mm, but 0.1 to 0.5 m for stable ejection and high density of the array.
It is preferably m. When the distance between the nozzle ejection surface and the carrier surface is less than 0.1 mm, the probe solution reaches the carrier surface before completely leaving the nozzle ejection port,
When the nozzle is moved for the next ejection, the droplet is pulled and twisted by the nozzle, which is not preferable.

(Liquid Ejecting Device) The probe manufacturing apparatus used in the method of the present invention has a liquid ejecting means used for ejecting the probe solution by the ink jet method. The liquid ejecting mechanism accommodates the probe solution. A liquid storage part (reservoir) for storing, a discharge port provided corresponding to each of the liquid storage parts, a liquid path for common communication between the liquid storage part and the discharge port, and a corresponding discharge port. Provided with a discharge energy generating means capable of discharging an independent probe solution by each of the discharge ports, and a liquid discharge part having a number corresponding to at least the plurality of types of probe solutions, While moving relatively to the carrier having a concave portion corresponding to the arrangement position of the probe on the surface, It is preferred that based on the distribution by ejecting each solution of said plurality of probes can be applied on a carrier.

In the method of the present invention, the liquid ejecting head having a plurality of ejecting nozzles in a direction substantially orthogonal to the scanning direction (a direction not orthogonal to each other) ejects the probe solution while scanning the carrier relative to the carrier. Is a method of manufacturing a probe carrier in which a plurality of types of probes are arranged on the carrier by setting the probe pitch interval in the direction orthogonal to the scanning direction to a and the ejection nozzle pitch interval of the liquid ejection head to Let b be the angle formed by the probe pitch direction and the nozzle pitch direction be θ, then scan the liquid ejection head by the angle θ that satisfies | cos θ | = a / b so that a becomes smaller than b. The scanning is performed with an inclination with respect to the direction orthogonal to the direction.

For a more detailed description, description will be made with reference to FIGS. 1 (a) and 1 (b). FIGS. 1A and 1B are views showing the present invention and a conventional inkjet scanning method, wherein 1 is a liquid ejection head, 2 is a liquid ejection head nozzle, 3 is a carrier, and 4 is a probe spot. The arrow indicates the scanning direction. Here, a represents a probe pitch interval in a direction orthogonal to the scanning direction, b represents an ejection nozzle pitch interval, and θ represents an angle formed by the probe pitch direction and the nozzle pitch direction.

FIG. 1A shows a conventional technique in which the liquid ejection head is relatively scanned so that the nozzle alignment direction of the liquid ejection head and the scanning direction are orthogonal to each other, and the probe solution is spotted on the carrier. And the case of | cos θ | = 1.

On the other hand, FIG. 1B shows, as a method of the present invention, the liquid ejection head is tilted and relatively scanned so that the nozzle alignment direction of the liquid ejection head and the scanning direction are not orthogonal to each other, and the probe solution is applied. It is a figure spotted on the carrier,
This is the case where cos θ | is less than 1.

In FIGS. 1A and 1B, when the ejection nozzle pitch interval b is the same value, the angle θ is adjusted according to the pitch a so that a = b│cos θ│ by the method of the present invention. The probe pitch interval a in the direction orthogonal to the scanning direction can be further shortened by adjusting, and as a result, the density can be increased. on the other hand,
When the probe pitch interval a in the direction orthogonal to the scanning direction has the same value in FIGS. 1A and 1B, the ejection nozzle pitch interval b can be made longer in the method of the present invention, so that the nozzles adhere to the vicinity of the nozzle. Since it is possible to further prevent a mixed solution of different probe solutions due to the probe solution to be used or to remove the probe solution by wiping more easily, it is possible to manufacture a highly reliable probe carrier that enables stable ejection.

Further, if the angle | θ | formed by the probe pitch direction and the nozzle pitch direction is larger than 0, the density can be increased, while considering the overlap of the probe pitch, the nozzle pitch, etc., the head and the probe can be considered. When the reservoir is an integral type, it is preferably 85 ° or less, and when the head and the probe reservoir are independent, it is preferably 89.5 ° or less.

In the method of the present invention, a plurality of liquid ejection heads are used, and the plurality of liquid ejection heads are collectively set to the angle θ.
The probe solution may be discharged onto the carrier by tilting and scanning. The plurality of liquid ejection heads are often formed as an integral type called a liquid ejection head block. In the liquid ejection head block, the nozzle portion may be integrated and each probe may be provided with a separate probe solution, or the nozzle may be composed of one or a plurality of units and fixed in a head block manifold shape. In the liquid ejection head block, the mounting angles of a plurality of heads can be changed at the same time. That is, by providing a mechanism for simultaneously rotating a plurality of heads having the same nozzle pitch, it becomes possible to efficiently and efficiently match the probe pitch and the pitch of the nozzles used in the liquid ejection head.

In the method of the present invention, the liquid ejection head (or liquid ejection head block) may be moved relative to the carrier by moving the carrier during scanning, or the liquid ejection head (or liquid). Discharge head block)
The liquid ejection head (or the liquid ejection head block) may be moved relative to the carrier by moving the head, or the liquid ejection head (or the liquid ejection head block) may be moved by moving both the carrier and the liquid ejection head. The head (or liquid ejection head block) may scan relative to the carrier. However, from the viewpoint of stable ejection, it is preferable to move the carrier rather than moving the liquid ejection head (or liquid ejection head block) because stable scanning can be performed. It is most preferable because scanning can be performed more stably by moving both. In order to move the carrier, a stage holding the fixed carrier may be moved.

The liquid ejection head used in the method of the present invention is a head for ejecting the probe solution by utilizing thermal energy, and a thermal energy generator for generating thermal energy applied to the probe solution. It is preferably provided. Inkjet methods that can be used in the manufacture of probe carriers include thermal jets that eject liquid by generating bubbles using thermal energy generated from electrothermal converters such as heaters and laser light. Method (bubble jet method)
Then, there is a piezo inkjet system in which the liquid is ejected by the deformation of the device caused by applying a voltage to the piezo device, and any of them can be used in the method of the present invention. Of these, the head for the thermal jet method has a simpler structure than the head for the piezo inkjet method, and is suitable for downsizing the head and increasing the number of nozzles. Method,
Since the binding reaction to the carrier is completed in a short time and the secondary structure of DNA is also eliminated by heat, the efficiency of the subsequent hybridization reaction can be increased, and the thermal jet method is also used. The inkjet method is more preferably used for the present invention.

Regarding its typical structure and principle, see, for example, US Pat. Nos. 4,723,129 and 4740.
It is preferable to use the basic principle in the field of ink recording disclosed in Japanese Patent No. 796. This method can be applied to both the so-called on-demand type and the continuous type. In particular, in the case of the on-demand type, it is arranged corresponding to the sheet or liquid path holding the liquid (ink). The electrothermal converter being subjected to a rapid temperature rise corresponding to the recorded information and exceeding the nucleate boiling.
By applying two drive signals, heat energy is generated in the electrothermal converter, film boiling is caused on the heat acting surface of the recording head (ink jet head), and as a result, the drive signals are corresponded one-to-one. This is effective because bubbles can be formed in the liquid (ink). By the growth and contraction of the bubbles, liquid (ink) is ejected through the ejection opening,
Form at least one drop. It is more preferable to make this drive signal into a pulse shape because bubbles can be immediately and appropriately grown and contracted, so that liquid (ink) ejection with excellent responsiveness can be achieved. As the pulse-shaped drive signal, U.S. Pat. Nos. 4,463,359 and 4,345 are used.
Those as described in the '262 patent are suitable. If the conditions described in US Pat. No. 4,313,124 of the invention relating to the rate of temperature rise on the heat acting surface are adopted, more excellent recording can be performed.

As the constitution of the recording head, in addition to the combination constitution of the discharge port, the liquid passage, and the electrothermal converter (the linear liquid passage or the right-angled liquid passage) as disclosed in the above-mentioned respective specifications, U.S. Pat. No. 4,558,333 and U.S. Pat. No. 44, which disclose a configuration in which a heat acting portion is arranged in a bending region.
The structure using the specification of No. 59600 is also included in the present invention. In addition, Japanese Unexamined Patent Publication No. 59-123670 discloses a configuration in which a common slit is used as a discharge portion of the electrothermal converter for a plurality of electrothermal converters, and an opening for absorbing a pressure wave of thermal energy is provided. The effect of the present invention is effective even when the configuration corresponding to the ejection portion is disclosed in JP-A-59-138461. That is, regardless of the form of the recording head (liquid ejection head), according to the present invention, spotting of the probe can be performed reliably and efficiently.

Furthermore, the present invention can be effectively applied to a full-line type liquid ejection head having a length corresponding to the maximum width of a carrier that can be spotted by the liquid ejection device. Such a liquid ejection head may have a configuration that fills its length by a combination of a plurality of heads or a configuration as one head that is integrally formed.

In addition, even in the case of the serial type, when it is attached to the head fixed to the apparatus main body or the apparatus main body, it becomes possible to electrically connect to the apparatus main body and apply the probe solution from the apparatus main body. The present invention is also effective when a replaceable chip type liquid discharge head or a cartridge type head in which a solution reservoir is integrally provided in the liquid discharge head itself is used.

Further, as the structure of the liquid ejecting apparatus, it is preferable to add a head ejection recovering means, a preliminary auxiliary means, etc. because the effect of the present invention can be further stabilized. Specific examples thereof include cleaning means for the liquid ejection head, pressurizing or suctioning means, preheating means for heating using an electrothermal converter or a heating element other than this, or a combination thereof, Preliminary ejection means for performing ejection other than spotting can be mentioned.

The most effective ones for the above solutions are
The film boiling method described above is executed.

The nozzle diameter is designed according to the amount of probe solution discharged from one nozzle at a time, as described later.
Although it is preferably adjusted, it is usually selected from the range of 5 to 70 μm.

Further, the nozzle pitch interval is preferably as short as possible in order to increase the density of the probe array, but from the viewpoint of preventing mixed liquid due to mist, the nozzle pitch interval is preferably large and is 21 μm or more. Is preferred.

In the method of the present invention, it is preferable to have a mechanism for removing the droplets of the probe solution adhering to the vicinity of the discharge nozzle of the liquid discharge head (or liquid discharge head block) by wiping, but in particular, It is preferable to have a mechanism for removing by wiping in a direction substantially perpendicular to the arrangement direction of the discharge nozzles. By removing the droplets of the probe solution adhering to the vicinity of the discharge nozzle, the vicinity of the discharge nozzle is cleaned,
It is possible to obtain ejection stability and liquid mixture prevention effect. In particular, when removing by wiping in a direction substantially perpendicular to the arrangement direction of the discharge nozzles, since the wiping is performed slightly obliquely to the nozzle arrangement direction, or the same wiping material is used, During use, the solution is absorbed more and more, diffuses and does not mix,
It is possible to prevent the generation of a mixed solution of different probe solutions. The liquid ejection head is housed in the head recovery mechanism during the manufacturing process after the start and end of the manufacturing process of the DNA chip array, and the wiping can be performed substantially orthogonal to the nozzle arrangement direction. In this method, it is desirable that the removing means be scraped with an elastomer member, wiped with a moisture absorption / wetting member, or blown with a gas near the nozzle outlet.

The size and capacity of the solution reservoir are appropriately selected depending on the amount of the probe solution discharged from the nozzle at one time, the number of arrays to be manufactured, and the like. When the liquid ejection head is integrally formed by processing a carrier such as a silicon substrate, the nozzle diameter and the amount of the probe solution ejected at one time are naturally within a certain range and face the nozzle opening. In some cases, the size formed on the surface of the carrier is sufficient. On the other hand, the amount of probe solution discharged from the nozzle at once is relatively large, and
When the number of arrays required to be prepared is large, a method in which the probe solution is added to the solution reservoir to cover the amount of solution required when preparing the total number of arrays is considered. Alternatively, the structure may be such that it is attached to a solution reservoir arranged on one surface of the carrier and further has a solution storage portion for increasing the volume. At that time, the addition of the probe solution is carried out via the solution containing portion for increasing the volume, and the shape itself of the solution containing portion can be set so that the probe solution can be easily applied if necessary. For each probe used in this array, a pair of solution reservoirs and nozzles are arranged in an array corresponding to each other. Therefore, the number of solution reservoirs and nozzles forming a pair is not particularly limited, and is selected according to the probe type of the array required. Although the dot diameter, the number of dots, the coating density, and the array shape are naturally limited depending on the manufacturing method of the liquid ejection head, the total number of paired solution reservoirs and nozzles is basically the probe type. Determined by

(Liquid Ejecting Head) In the above, a concrete means for realizing the liquid ejecting head so that the nozzle arrangement direction of the liquid ejecting head and the scanning direction are not orthogonal will be described in more detail. Hereinafter, a preferred embodiment of the present invention will be described in detail with reference to the accompanying drawings. Before that, an outline of the present embodiment will be described.

The present embodiment relates to an apparatus for manufacturing a probe carrier by an ink jet method, which uses a head mount using a plurality of multi-nozzle type liquid ejection heads having a plurality of nozzles.

The head mount has a mechanism for simultaneously changing the mounting angles of a plurality of heads.

When the probe is applied to the probe carrier by the ink jet method, basically, a fixed interval (pitch)
With a multi-head having a plurality of nozzles, the nozzles that match the probe pitch are used to scan and eject in the main scanning direction, then the head or carrier is moved in the sub-scanning direction, and then the main scanning direction. The scanning and discharging of are repeated.

In the case of the multi-nozzle liquid ejection head of this embodiment, a mechanism for simultaneously rotating a plurality of heads having the same nozzle pitch is provided to efficiently match the probe pitch and the nozzle pitch of the liquid ejection head at the same time. It becomes possible.

The specific structure of the probe carrier manufacturing apparatus of one embodiment will be described below.

FIG. 2 is a perspective view showing the internal structure of the head mount 55, and FIG. 3 is a plan view of FIG. 2 seen from above.

2 and 3, reference numerals 204a, 204
Reference numerals b and 204c each denote a multi-nozzle liquid ejection head, and 205 denotes a nozzle (since the nozzle is located on the lower surface of the liquid ejection head, it is not visible in FIG. 3, but is shown by a solid line for convenience of description). The nozzles are arranged at the same pitch in the longitudinal direction of the head. Liquid discharge head 20
4a, 204b, and 204c have holders 20 at one end thereof.
8a, 208b, and 208c, respectively, and these holders are attached to the rotary shaft 2 fixed to the head mount 55.
Head mount 5 centered on 06a, 206b, 206c
5 is rotatably supported in the horizontal plane. Further, the liquid ejection heads 204a, 204b, 204c are
The other ends are supported by holders 210a, 210b, 210c, and these holders are supported by the slide member 214 so as to be rotatable in the horizontal plane about the rotation shafts 212a, 212b, 212c. Slide member 21
6 is supported so as to be movable in the X and Y directions with respect to the head mount 55, and is urged by a spring 216 in the direction of arrow A. Spring 216 of head mount 55
A fine movement screw 218 is provided at a position on the opposite side to the slide member 216 is moved in the X direction by rotating the fine movement screw 218. This makes 3
The two liquid ejection heads 204a, 204b, 204c can be simultaneously tilted by an arbitrary angle θ (with respect to the Y axis) with respect to the position shown by the broken line in FIG. 3, and the tilt with respect to the scanning direction is adjusted. In addition, the holders 210a, 210b, 2
The compression springs 220a, 220b, 220c are provided in the 10c, and the liquid ejection heads 204a, 204b,
204c is biased rightward in the figure.

When the head mount 55 is set in the apparatus so that the main scanning direction X and the straight line connecting the rotary shafts 206a, 206b, 206c of the respective heads are in the same direction,
It is convenient for adjustment.

The head rotating shaft 206 is used for the actual discharge holding.
A plurality of heads are simultaneously rotated about a, 206b, and 206c, and the head angle θ is adjusted so that the desired nozzle pitch and the probe pitch are matched. At this time, if the probe pitch is a and the nozzle pitch is b, an angle θ that satisfies a = b | cos θ |
Only tilt the head. Next, the fine adjustment screws 222a, 22
2b and 222c to adjust the nozzle position to R, G, and B
Align with the position of each probe pattern of.

FIG. 4 is a block diagram of the controller of the probe carrier manufacturing apparatus 90. 59 is the controller 5
A teaching pendant, which is an input / output unit of 8, a display unit 62 for displaying information on the progress of manufacturing and the presence / absence of an abnormality in the head, and 60 an operating unit (keyboard) for instructing the operation of the probe carrier manufacturing apparatus 90 Is. Reference numeral 58 is a controller for controlling the overall operation of the probe carrier manufacturing apparatus 90, 65 is an interface for exchanging data with the teaching pendant 59, 66 is a CPU for controlling the probe carrier manufacturing apparatus 90, and 67 is a CPU 6
R storing a control program for operating 6
OM, 68 is a RAM for storing production information and the like, 70 is a discharge control unit for controlling discharge of the probe to the probe carrier, 7
Reference numeral 1 denotes a stage controller that controls the operation of the XYθ stage of the probe carrier manufacturing apparatus 90, and 90 denotes a color filter manufacturing apparatus that is connected to the controller 58 and operates according to the instructions.

[0093]

EXAMPLES The present invention will be described in more detail with reference to the following examples. Although the examples shown here are examples of preferred embodiments of the present invention, the present invention is not limited to these examples.

Example 1 (Evaluation of Color Mixing Using Fluorescent Dye) The effect of the droplet removing function was confirmed as follows.

25.4 mm x 25.4 mm x 0.5 mm
After ultrasonically cleaning the fused quartz substrate of No. 2 in 1% ultrasonic detergent GP-III (Branson) for 20 minutes, ultrasonic cleaning with tap water and running water cleaning were appropriately performed. Next, 1N at 80 ℃
It was immersed in NaCl for 20 minutes, washed with running water (tap water), washed with ultrapure water ultrasonic waves, washed with running water (ultra pure water), and then dried by blowing nitrogen gas.

Next, a solvent for ejecting the fluorescent dye Rhodamine B with an ink jet head, that is, 7.5 wt% glycerin, 7.5 wt% urea, 7.5 wt% thiodiglycol, a general formula (IV):

[0097]

[Chemical 1]

An aqueous solution containing 1 wt% of acetylene alcohol (for example, trade name: acetylenol EH Kawaken Fine Chemical Co., Ltd.) represented by was dissolved at a concentration of 10 μM. This dye solution was injected into every other liquid reservoir formed by being connected to each nozzle of the inkjet head schematically shown in FIG. The fluorescent dye aminoFITC (concentration 10 μM), which was similarly dissolved, was injected into the reservoir in which the rhodamine B solution was not injected. From each nozzle of this inkjet head, the solutions of the above-mentioned two kinds of dyes were discharged and supplied onto the above-mentioned cleaned glass substrate. At this time, since the probe pitch interval in the direction orthogonal to the scanning direction is 100 μm and the ejection nozzle pitch interval of the inkjet is 575 μm, the angle formed by the probe pitch direction and the nozzle pitch direction is 80 °.
And Each reservoir and nozzle were washed with the above solvent and each dye solution in advance, and the liquid removal by vacuum suction was appropriately repeated. The amount of one droplet is 10 pl, and in this case, the area occupied by the droplets (dots) ejected on the substrate is about 50 μm, and the interval between the dots is about 100 μm. The dye solution was discharged once from the head, wiped by a wiper blade in a direction substantially perpendicular to the direction of arrangement of the discharge nozzles, and was discharged again. This method was repeated 500 times in total.

Nikon fluorescence microscope ECLIPSE E80
0 (Nikon Corporation) 20x objective lens (plan apochromat) and fluorescent filter (B-2A: Rhodamine B)
, B-2E / C: for amino FITC, both are Nikon), and the state of each of the supplied dye aqueous solutions was observed by fluorescence at a magnification of 200 times. Fluorescence of other dyes was not observed, and it was possible to supply an aqueous solution with no color mixture. Further, each dot was arranged in an orderly manner, and no stain or stain derived from mist was observed.

Example 2 (Evaluation of Color Mixing by Hybridization) The glass substrate was washed in the same manner as in Example 1. Then, vacuum distillation was purified by the following formula _{(I): (CH 3 O} ) 3 SiC 3 H 6 NHC 2 H 4 NH 2 (I) aminosilane coupling agent (KBM-603: Compound I-Etsu Chemical stock (Manufactured by the company) was stirred at room temperature for 1 hour to hydrolyze the methoxy group portion. Next, the above substrate was immediately immersed in the silane coupling agent aqueous solution after cleaning, and at room temperature for 1 hour. After that, it was washed with running water (ultra pure water), blown with nitrogen gas to be dried, and then heat-fixed in an oven at 120 ° C. for 1 hour.

After cooling, the following formula (II):

[0102]

[Chemical 2]

The substrate was immersed in a 0.3% solution of N- (6-maleimidocaproxy) succinimide (EMCS; compound II) (ethanol: dimethylsulfoxide = 1: 1) at room temperature for 2 hours, and EMCS was used as aminosilane. It was reacted with the amino group of the coupling agent. After completion of the reaction, the mixture was washed once with ethanol: dimethyl sulfoxide = 1: 1 and three times with ethanol, and dried by blowing nitrogen gas. 5'-ATGAACCGGAGGGCCCATC-3 '3'-TACTTGGCCTCCGGGT-5'5'-AAAAAAAAAAAAAAAAAAAAAAAAA-3'3'-TTTTTTTTTTTTTTTTTTTT'

Has a base sequence of, which is a base sequence complementary to each of the above base sequences, and
A mercapto group (S which is capable of reacting with the finally purified maleimide group on the surface of the substrate through a linker at the 5'end (S
An oligonucleotide having H group: also referred to as sulfhydryl group (compounds III and IV: Bex Co., Ltd.) was used as an oligonucleotide probe used for verification of this example. Formula (III, IV):

[0105] _{3'-TACTTGGCCTCCGGGTAG-OP (O 2} ) O- (CH 2) 6 - 5 ' Compound III

[0106] _{3'-TTTTTTTTTTTTTTTTTTTTTTTTT-OP (O 2} ) O - (CH 2) 6 -5 ' Compound IV

Compounds III and IV were dissolved in the same solvent for ejection as in Example 1 so that the absorbance was 1.0,
In the same manner as in Example 1, the ink was supplied from the inkjet head on every other glass substrate. The reservoirs and nozzles were washed with the solution of the solvent and the oligonucleotide in advance and the liquid was removed by vacuum suction as appropriate. The oligonucleotide solution is then discharged onto the above substrate by the same discharge method as in Example 1, and the droplet amount, dot size, dot interval, discharge pattern, discharge frequency, and wiping method are the same as in Example 1. is there. Further, the above solvent has a high moisturizing property and can prevent the oligonucleotide solution from drying in the reservoir and on the substrate.

The substrates thus ejected with the solutions of compounds III and IV were reacted in a moisturizing chamber having a humidity of 100% at room temperature for 1 hour, and then washed in running water (ultra pure water) for about 30 seconds. Next, the above substrate is 50 m containing BSA (bovine serum albumin Sigma-Aldrich Japan) at a concentration of 2%.
The sample was immersed in M phosphate buffer (pH = 7.0, containing 1M NaCl) for 1 hour, washed appropriately with the above buffer, and stored in this buffer.

(Hybridization) Model Target D
Compounds V and VI (purchased from Beck's Co., Ltd.) having the sequence of Example 1 as NA and having tetramethylrhodamine bound to the 5'end were used for hybridization.

As the model target DNA, the formula (V, VI):
(Only V is shown, VI is A25 for DNA part)

[0111]

[Chemical 3]

A DNA molecule of the above, having the sequence of the above,
Compounds V and VI (purchased from Bex Co., Ltd.) in which tetramethylrhodamine was bound to the 5'end as a fluorescent label were used to carry out a hybridization reaction with the prepared probe on the substrate.

This hybridization reaction was carried out by using a phosphate buffer solution (10 mM phosphate buffer solution pH = 7.0, 50) containing two substrates, each containing Compounds V and VI at a concentration of 5 nM.
2 ml (containing mM NaCl) was used and performed separately in the hybrid pack. The substrate was sealed in a hybrid pack together with the model target DNA solution, heated to 70 ° C. in a thermostat, then cooled to 50 ° C., and left in that state for 10 hours. Next, the substrate was taken out from the hybrid pack and washed with a hybridization buffer for the purpose of removing unreacted target DNA. After washing, the substrate was placed on a slide glass in a state of being covered with a buffer solution, covered with a cover glass, and the fluorescence from the fluorescent label was observed. The fluorescence microscope used for this observation was an ECLIPSE E800 (Nikon Corporation) with a 20 × objective lens (plan apochromat) and a fluorescence filter (Y-2E / C) set. Fluorescence was observed only from the portion of each substrate where the probe complementary to the target DNA was to be present. The dots were arranged in an orderly manner, and no stain or stain derived from mist was observed.

[0114]

As described above, while the liquid ejection head having a plurality of ejection nozzles in the direction not orthogonal to the scanning direction relatively scans the carrier, the probe solution is ejected onto the carrier. By manufacturing the probe array by using the probe array, it is possible to cope with various probe pitches without replacing the liquid ejection head (block) for each different probe pitch. That is, by scanning the liquid ejection head while inclining with respect to the direction orthogonal to the scanning direction, the probe pitch becomes short at a constant nozzle interval, and thus a higher density probe array can be manufactured, When the probe pitch is constant, the nozzle pitch can be made longer, so that the ejection stability can be improved. In this case, it is easy to remove the droplets adhering to the vicinity of the nozzle by wiping. You can also Therefore, it is possible to obtain a low-cost, stable, high-density probe carrier that can stably discharge the probe solution.

[0115]

[Sequence list]                                 SEQUENCE LISTING <110> CANON INC. <120> Probe carrier, Method and Apparatus for producing Probe carrier <130> 4692002 <150> JP 2001/94400 <151> 2001-03-28 <160> 4 <210> 1 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Designed oligonucleotide to be hybridized with the designed oligon ucleotide "gatgggcctccggttcat" <400> 1   atgaaccgga ggcccatc 18 <210> 2 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> Designed oligonucleotide used as a probe to be stabilized on a car rier <400> 2   gatgggcctc cggttcat 18 <210> 3 <211> 25 <212> DNA <213> Artificial Sequence <220> <223> Designed oligonucleotide to be hybridized with the designed oligon ucleotide "ttttttttttttttttttttttttt" <400> 3   aaaaaaaaaa aaaaaaaaaa aaaaa 25 <210> 4 <211> 25 <212> DNA <213> Artificial Sequence <220> <223> Designed oligonucleotide used as a probe to be stabilized on a sur face of a carrier <400> 4   tttttttttt tttttttttt ttttt 25

[Brief description of drawings]

FIG. 1 is a diagram showing an inkjet scanning method according to the present invention.

FIG. 2 is a perspective view showing an internal configuration of a head mount.

FIG. 3 is a plan view of FIG. 3 viewed from above.

FIG. 4 is a diagram showing a configuration of a control unit that controls the operation of the probe carrier manufacturing apparatus.

[Explanation of symbols]

11 Carrier 12 probes 13 Liquid ejection head unit 14 nozzles 15 scanning direction 204 Liquid ejection head 205 nozzle 206 rotation axis 208 holder 210 holder 212 rotation axis 214 Slide member 216 spring 218 Fine movement screw 220 Main compression spring 55 head mount 58 Controller 59 Teaching pendant 60 Control 62 display 65 Interface 66 CPU 67 ROM 68 RAM 70 Discharge control unit 71 Stage controller 90 Probe carrier manufacturing device

Front page continuation (51) Int.Cl. ⁷ Identification code FI theme code (reference) C12N 15/09 ZNA B41J 3/04 103B G01N 33/566 102H 37/00 102 C12N 15/00 ZNAF (72) Inventor Okamoto Shoji 3-30-2 Shimomaruko, Ota-ku, Tokyo Canon Inc. F-term (reference) 2C056 FA03 FB01 HA07 JB04 2C057 AG12 AH20 AJ10 BA03 BA13 4B024 AA11 AA20 CA09 HA19 HA20

Claims (21)

[Claims] 1. A liquid ejection head having a plurality of ejection nozzles in a direction that is not orthogonal to the scanning direction relatively ejects a probe solution onto the carrier from the ejection nozzles while performing relative scanning on the carrier. A method for producing a probe carrier in which a plurality of types of probes capable of specifically binding to a target substance are arranged on the carrier, wherein a probe pitch interval in a direction orthogonal to the scanning direction is a
And the discharge nozzle pitch interval of the liquid discharge head is b and the angle formed by the probe pitch direction and the nozzle pitch direction is θ, | cos θ | = a / b so that a becomes smaller than b. A method for manufacturing a probe carrier, which comprises scanning the liquid discharge head with an angle θ satisfying the above condition while being inclined with respect to a direction orthogonal to the scanning direction. 2. The method of manufacturing a probe carrier according to claim 1, wherein the liquid ejection head scans relative to the carrier by moving the carrier. 3. The method for producing a probe carrier according to claim 1, wherein the liquid ejection head scans relative to the carrier by moving the liquid ejection head. 4. The probe carrier according to claim 1, wherein the liquid ejecting head scans relative to the carrier by moving the carrier and the liquid ejecting head. Manufacturing method. 5. The probe solution is ejected onto the carrier by using a plurality of liquid ejection heads and tilting the plurality of liquid ejection heads at a time by the angle θ to scan the carrier. 5. The method for producing the probe carrier according to any one of items 1 to 4. 6. The liquid ejection head is a head for ejecting the probe solution using thermal energy,
The method for producing a probe carrier according to claim 1, further comprising a thermal energy generator for generating thermal energy to be applied to the probe solution. 7. The mechanism according to claim 1, further comprising a mechanism for removing droplets of the probe solution adhering to the vicinity of the discharge nozzle by wiping in a direction substantially perpendicular to the arrangement direction of the discharge nozzles. The method for producing the probe carrier according to any one of items. 8. The amount of the probe solution ejected from the ejection nozzle is 0.1 picoliter to 100 picoliter.
Item 6. A method for producing a probe carrier according to item. 9. The area occupied by each of the probe solutions applied from the discharge nozzle is 0.01 μm ^{2 to} 400.
The method for producing a probe carrier according to claim 1, wherein the probe carrier has a diameter of 00 μm ² . 10. The probe is DNA, RNA, or cDNA.
NA (Complementary DNA), PNA, oligonucleotide, polynucleotide, other nucleic acid, oligopeptide, polypeptide, protein, enzyme, substrate for enzyme, antibody, epitope for antibody, antigen, hormone, hormone receptor, ligand, ligand receptor , Oligosaccharides, polysaccharides, and combinations thereof.
The method for producing the probe carrier according to any one of 1. 11. A liquid ejection head having a plurality of ejection nozzles in a direction not orthogonal to a scanning direction relatively ejects a probe solution onto the carrier from the ejection nozzles by an inkjet method while relatively scanning the carrier. ,
An apparatus for manufacturing a probe carrier, in which a plurality of types of probes capable of specifically binding to a target substance are arranged on the carrier, wherein a probe pitch interval in a direction orthogonal to the scanning direction is a
And the discharge nozzle pitch interval of the liquid discharge head is b and the angle formed by the probe pitch direction and the nozzle pitch direction is θ, | cos θ | = a / b so that a becomes smaller than b. An apparatus for manufacturing a probe carrier, wherein the liquid ejection head is inclined by an angle θ satisfying the above condition with respect to a direction orthogonal to the scanning direction for scanning. 12. The probe carrier manufacturing apparatus according to claim 11, wherein the liquid ejection head scans relative to the carrier by moving the carrier. 13. The probe carrier manufacturing apparatus according to claim 11, wherein the liquid discharge head scans relative to the carrier by moving the liquid discharge head. 14. The probe carrier according to claim 11, wherein the liquid ejecting head scans relative to the carrier by moving the carrier and the liquid ejecting head. Manufacturing equipment. 15. The probe solution is ejected onto the carrier by using a plurality of liquid ejection heads and scanning the plurality of liquid ejection heads at a time by inclining by the angle θ. 15. The probe carrier manufacturing apparatus according to any one of 1 to 14. 16. The liquid ejection head is a head for ejecting the probe solution using thermal energy, and includes a thermal energy generator for generating thermal energy to be applied to the probe solution. The probe carrier manufacturing apparatus according to claim 11, wherein the probe carrier manufacturing apparatus is a probe carrier manufacturing apparatus. 17. The mechanism according to claim 11, further comprising a mechanism for removing droplets of the probe solution adhering to the vicinity of the discharge nozzle by wiping in a direction substantially perpendicular to the arrangement direction of the discharge nozzles. An apparatus for manufacturing a probe carrier according to any one of claims. 18. The probe carrier according to claim 11, wherein the amount of the probe solution ejected from the ejection nozzle is 0.1 picoliter to 100 picoliter. Manufacturing equipment. 19. The area occupied by each of the probe solutions applied from the discharge nozzle is 0.01 μm ² to 40.
It is 000 μm ^2.
9. The apparatus for producing a probe carrier according to any one of items 8. 20. The probe is DNA, RNA, or cDNA.
NA (Complementary DNA), PNA, oligonucleotide, polynucleotide, other nucleic acid, oligopeptide, polypeptide, protein, enzyme, substrate for enzyme, antibody, epitope for antibody, antigen, hormone, hormone receptor, ligand, ligand receptor 20. The probe carrier manufacturing apparatus according to claim 11, wherein the probe carrier is selected from the group consisting of :, oligosaccharides, polysaccharides, and combinations thereof. 21. A probe carrier manufactured by the method according to any one of claims 1 to 10.