Patent Application
20050048219
Many analytical and preparative methods used in biology and medicine are based on attachment of compounds, such as peptide ligands or oligonucleotide probes, to a substrate. Frequently, multiple compounds are attached, each at a predefined location, onto the surface of the substrate. Such attachment can be achieved in a number of different ways, including covalent and non-covalent bonding.
A number of protocols have been developed to covalently attach a compound to a substrate, such as a microscopic glass slide. In one example, an oligonucleotide is synthesized directly on the substrate surface using a photolithographic process. In another example, a nucleic acid, such as a cloned cDNA, a PCR product, or a synthetic oligonucleotide, is deposited onto the substrate in the form of an array. The array can then be used in hybridization assays in order to determine the presence or abundance of particular sequences in a sample.
Before the compounds can be attached to a substrate, the substrate surface must be thoroughly cleaned to remove contaminants, typically by a chemical wash process. Then, the substrate surface is modified with organosilane having a functional group (e.g., aldehydes and amines) to facilitate attachment of the compounds. This can be achieved by a vapor deposition process or a solution coating process.
In one aspect, the invention is directed towards a method of treating a substrate surface by providing a chamber having a substrate, cleaning a surface of the substrate in the chamber with a plasma, and introducing to the chamber a vapor of un-ionized organosilane molecules having functional groups and reacting the un-ionized organosilane molecules having functional groups with molecules on the plasma-cleaned surface via a silane condensation reaction, thereby producing a layer containing functional groups.
Examples of the functional groups include, but are not limited to, amine, aldehyde, epoxy, isocyanide, thiol, mercapto, hydroxyl, carboxyl, vinyl, halocarbon, disulfide, halogen-substituted alkyl, succinimide, methacryl, and acryl. To facilitate the silane condensation reaction, each organosilane molecule preferably contains at least one alkoxyl, hydroxyl, or halo group attached to its silicon atom. Examples of the organosilane molecules having functional groups include, but are not limited to, 3-aminopropyltrimethoxysilane (3-APTMS) and glycidoxypropyltrimethoxysilane (GPTMS). The plasma used in this method can be O2 plasma, air plasma, CO2 plasma, Ar plasma, N2 plasma, hydrogen plasma, helium plasma, water plasma, hydrogen peroxide plasma, or a combination thereof. The surface to be treated may be composed of glass, quartz, ceramic, silicon, metal, gallium arsenide, or polymer.
The cleaning step in the above-described method can be performed by providing a water vapor or a hydrogen peroxide vapor in the chamber and generating a plasma from the vapor under certain conditions, e.g., at 20 to 300° C. and at a chamber pressure of 50 to 1000 mTorr, thereby cleaning a surface of the substrate. The introducing and reacting step can be performed in the chamber at 20 to 300° C., preferably 50° to 120° C. and at a chamber pressure of 50 mTorr to 760 Torr, preferably 0.5 to 5 Torr.
In some embodiments, the method further includes one or more of the following steps: (1) depositing water or hydrogen peroxide on the surface of the substrate before, during, or after the introducing step; (2) cleaning the chamber, e.g., by vacuum, before the introducing and reacting step; and (3) curing the layer formed on the substrate by a baking process. In other embodiments, the method also includes introducing another vapor having organosilane molecules into the chamber. A water vapor or a hydrogen peroxide vapor can be introduced into the chamber before, during, or after the just-mentioned vapor is introduced.
In another aspect, the invention is directed towards an apparatus having a chamber, electrodes to supply power to the chamber for generating a plasma, an inlet to allow a gas suitable for generating the plasma to enter the chamber, a vessel coupled to the chamber for containing an organosilane solution, and a heater to heat the solution. The organosilane solution has a compound suitable for silanizing a surface of a substrate placed in the chamber. A power supply is coupled to the electrodes to supply power to the chamber to generate the plasma in the chamber. A second vessel can also be coupled to the chamber to store water or hydrogen peroxide.
In another aspect, the invention is directed towards an apparatus having a chamber, means for plasma cleaning a surface of a substrate in the chamber, and means for silanizing the cleaned surface in the chamber.
The silanizing means includes one or more vessels for storing oganosilane solutions. For example, a first vessel coupled to the chamber stores a first organosilane solution. A second vessel also coupled to the chamber stores a second organosilane solution. The first and second organosilane solutions can be the same or different. A computer selects organosilane solutions for silanizing the cleaned surface according to a predefined protocol that defines the sequence or combination of the selected organosilane solutions for silanizing the cleaned surface. The apparatus may further include means for depositing water or hydrogen peroxide on the surface of the substrate.
In another aspect, the invention is directed towards an apparatus having a first chamber, a second chamber, and a gate disposed between the first and the second chambers. The gate is movable between a first position where the first chamber is connected to the second chamber and a second position where the first chamber is closed off from the second chamber. The apparatus includes electrodes to supply power suitable for generating a plasma to the first chamber, an inlet to allow a gas to enter the first chamber, the first gas suitable for generating the plasma to clean a substrate in the first chamber. A vessel coupled to the second chamber contains an organosilane solution having a compound suitable for silanizing a surface of the substrate. The apparatus may also include means for moving a substrate from the first support to the second support when the gate is moved to the first position.
In another aspect, the invention is directed towards an apparatus having a chamber, electrodes to supply power to the chamber for generating a plasma, an inlet coupled to the chamber to allow a gas suitable for generating a plasma to enter the chamber, an inlet coupled to the chamber to allow another gas to enter the chamber, the other gas including an organosilane compound, and an outlet coupled to the chamber to allow the gases to exit the chamber.
In one embodiment, the apparatus includes a heater that receives a vessel, the vessel containing an organosilane solution that generates an organosilane gas when the solution is heated by the heater. A mass flow controller is coupled to the first inlet to regulate the gas flowing through the first inlet. A computer controls the power supply, the mass flow controller, and the heater.
The details of one or more embodiments of the invention are set forth in the accompanying drawings and the description below. Other features, objects, and advantages of the invention will be apparent from the description and drawings, and from the claims.
Like reference symbols in the various drawings indicate like elements.
A compact silanization system is provided by using a single chamber for both cleaning and silanization of substrates, such as glass slides. Referring to
A silane condensation reaction occurs between the un-ionized organosilane vapor and a surface of glass slide 104 in camber 102. The silane condensation reaction is well known in the art (see, e.g., B. Arkles, “Silane Coupling Agent Chemistry,” in Silicon Compounds: Register and Review, United Chemical Technologies, Inc., PA, pp. 59-64). In the silane condensation reaction, a silyl group of each organosilane molecule reacts with a functional group of a molecule on a surface of glass slide 104, e.g., hydroxyl. As a result, a new covalent bond is formed between a silicon atom of the silyl group and an atom of the functional group. In other words, the silicon atom of the silane molecule is linked to the surface of a substrate by covalent bonding. At the same time, a bond in the silyl group cleaves to release a water or alcohol molecule. See Scheme 1 below. This reaction is also referred to as “silanization” herein. To facilitate the silane condensation reaction, each organosilane molecule preferably contains at least one hydride, alkoxyl, hydroxyl, or halo group attached to its silicon atom.
After a preset amount of time, a layer of organosilane is deposited on the surface of the glass slides. The silanized glass surface allows attachment of target compounds, e.g., cDNA, PCR products, oligos, and proteins in array assay.
The following is a description of a cleaning process using the silanization system 100. Initially, glass slides 104 are mounted on a slide holder 138 (see
As the plasma gas enters chamber 102, the pressure inside the chamber increases. Mass flow controller 130 is adjusted to maintain a constant flow of plasma gas into the chamber when a preset pressure is reached. The preset pressure can be 30 mTorr to 2 Torr, preferably 100 to 500 mTorr. Then power supply 108 is turned on to provide plasma power to chamber 102 through electrodes 106. The power supply can be 10 to 2000 watts, preferably 150 to 500 watts. The frequency of power supply 108 can be from 0 (DC) to 10 GHz (microwave). The plasma gas is energized into a plasma that reacts with the surface of glass slides 104 and removes contaminants thereon. Power supply 108 is turned on for a period of 0.1 to 120 minutes, preferably 5 to 30 minutes. After the power supply is turned off, valve 132 is opened to allow the plasma gas to be removed from the chamber. When the pressure inside chamber 102 drops to the baseline pressure, valve 132 is closed.
The following describes a silanization process using the silanization system 100. After the glass slides are thoroughly cleaned by the plasma, oven 128 is adjusted to a temperature sufficient to vaporize the organosilane solution 118 in vessel 116, and the temperature in chamber 102 is adjusted to a level sufficient to facilitate silane condensation reaction. The temperature of the chamber may be controlled by the heat generated by heater 128, or by a separate heater (not shown). Solution 118 contains organosilane having functional groups, such as amines, aldehydes, epoxy, isocyanide, thiols, hydroxyl, carboxyl, vinyl, or halocarbons (e.g., fluorocarbons). The oven temperature can be room temperature to 300° C., preferably room temperature to 150° C. In one example where aminosilane is used, the oven temperature is maintained at 80 to 90° C. In another example where epoxysilane is used, the oven temperature is adjusted to about 150° C. The chamber temperature can be room temperature to 300° C., preferably 50 to 120° C. Valve 120 is opened to allow the vapor from solution 118 to enter chamber 102 through inlet 114. When the chamber pressure reaches a preset pressure of 50 mTorr to 760 Torr, preferably 0.5 to 5 Torr, valve 120 is closed. After a preset time of 6 seconds to 20 hours, preferably 15 to 60 minutes, a layer of organosilane is deposited on the glass slides 104. Valve 132 is then open, and vacuum pump 112 removes gas from the chamber. Valve 132 is closed when the chamber pressure drops to the baseline pressure.
The following describes a curing process used after the glass slides have been silanized. The curing process can be conducted in vacuum or with ambient gas to distribute heat more evenly within the chamber. Any gas that does not react with the organosilane layer can be used to distribute heat. Preferably, N2, Ar, or other inert gases may be used. As an example, mass flow controller 130 is adjusted to allow nitrogen gas to enter chamber 102 through inlet 110 until chamber pressure reaches a preset value of 0 to 760 Torr, preferably 10 to 50 Torr. Chamber 102 is maintained at a preset temperature of 50 to 500° C., preferably 100 to 200° C., in order to bake the glass slides 104. The baking process dries the slides and “cures” the slides by enhancing the uniformity of the organosilane layer over the slides. Baking also allows the organosilane layer to couple more securely to the slides. The baking process is performed for a period of 0.1 minutes to 20 hours, preferably 15 to 60 minutes. A longer baking period is needed when a lower temperature is used, and vice versa. Then valve 132 is opened, and vacuum pump 112 pumps the gases out of chamber 102. When the chamber pressure lowers to the baseline pressure, valve 132 is closed. A vent valve (not shown) of chamber 102 is opened to allow nitrogen or room air to enter the chamber. The silanized glass slides are then removed from chamber 102.
The silanized glass slides are “activated” in the sense that each contains an organosilane layer that includes organosilane molecules with functional groups that interacts, covalently or non-covalently, with target compounds. Examples of the functional groups are amine, aldehyde, epoxy, isocyanide, thiol, mercapto, hydroxyl, carboxyl, vinyl, disulfide, halogen-substituted alkyl, succinimide, acryl, methacryl, and halocarbon (e.g., fluorocarbon). Note that the functional groups of the organosilane layer may be of the same type (e.g., they are all amines), or they may be of different types (e.g. amines plus hydroxyls). A glass slide may contain an organosilane layer having one of the above functional group, or a mixture of the above functional groups. Examples of target compounds are organic molecules DNA, oligos, and proteins. The silanized glass slides can be sealed in packages for later use, or be further processed to produce DNA microarrays or other types of biochips.
An advantage of using silanization system 100 is that glass slides can be conveniently cleaned and silanized in a laboratory at a low cost. The glass slides can be silanized shortly before target compounds are attached to the slides, thereby ensuring the freshness of the silanized slides. In comparison, silanized glass slides purchased from outside vendors have much shorter lifetime since they have already been on the shelf for several days or months. Thus, microarrays or biochips produced from slides that are cleaned and silanized by silanization system 100 may have a longer lifetime in the laboratory.
Another advantage of using silanization system 100 is that a single chamber 102 can be used for the plasma cleaning, water deposition (described below), silanization, and curing of the glass slides 104. By eliminating the need for moving the glass slides from one chamber to another when performing different processing steps, the likelihood that the slides will come into contact with dust or other contaminants is reduced. This ensures the quality of the silanized glass slides.
Treatment of the glass slides 104 may also include deposition of water or hydrogen peroxide before, during, or after organosilane compounds are deposited on the glass slides 104. A vessel 122 containing water 124 (or hydrogen peroxide) is coupled to inlet 114 through valve 126. The steps for cleaning the glass slides using a plasma is the same as described previously. When the plasma gas is pumped out of chamber 102, valve 132 is closed, and then valve 126 is opened so that a water vapor enters chamber 102 through inlet 114. The temperature of chamber 102 is maintained at a preset value between room temperature to 300° C., preferably room temperature to 100° C. As the water vapor enters chamber 102, the chamber pressure increases. When the pressure increases to a preset value between 50 mTorr to 760 Torr, preferably 0.5 Torr to 5 Torr, valve 126 is closed. Water acts as a catalyst to promote polymerization of organosilanes and allows the organosilane compounds to have a better coupling reaction with the glass slides. After 30 to 60 minutes, valve 132 is opened and vacuum pump 112 pumps the water vapor out of chamber 102. When the chamber pressure is reduced to the baseline pressure, valve 132 is closed. Afterwards, vapor deposition of organosilane compound (or compounds) and baking (or curing) of the silanized glass slides are conducted in the same manner as described previously.
Additional vessels (not shown) may be used to contain different types of organosilane solutions. More than one type of organosilanes may be introduced into chamber 102 at the same time. Different types (or different combinations) of organosilanes may also be introduced into chamber 102 sequentially, one after another.
An advantage of silanization system 100 is that the cleaning, water deposition, silanization, and curing steps are performed in the same chamber, so the whole process can be easily automated. Referring to
Referring to
Referring to
First chamber 136 and second chamber 138 are separated by a gate 140 that can move between an open position and a closed position. When gate 140 is moved to the closed position, first chamber 136 is sealed off from second chamber 138 so that different processes can operate in the chambers at the same time. A set of substrates 142 may be plasma cleaned in first chamber 136 while another set of substrates 144 are silanized in second chamber 138. When gate 140 is moved to the open position, first chamber 136 is connected to second chamber 138, and substrates can be moved from the first chamber to the second chamber. A robotic arm (not shown) may be used to move the substrates from the first chamber to the second chamber.
An advantage of using silanization system 400 is that much time is saved by plasma cleaning and silanizing different sets of substrates simultaneously. In addition, although first and second chambers are connected when gate 140 is moved to the open position, first and second chambers are sealed off from the room environment so that the substrates will not be contaminated by room air before the substrates are properly silanized.
Without further elaboration, it is believed that one skilled in the art, based on the description herein, can utilize the present invention to its fullest extent. The publications cited herein are hereby incorporated by reference in their entirety.
Table 1 shows water contact angles measured from of a set of twelve glass slides (or silicon wafers) treated under various conditions. Each value shown in the table is obtained from measurements of 3 slides (or wafers) with 5 measurements per slide (or wafer). The first set of measurements were made on glass slides cleaned by O2 plasma at 70° C. for 20 minutes. The O2 pressure during plasma cleaning was 200 mTorr, and the power was 250 watts. The water contact angles measured from the glass slides were 6.4±0.8 degrees before plasma cleaning, and were 4.5±0.1 degrees after cleaning. After plasma cleaning, a water vapor deposition was conducted at 1 Torr for 30 minutes. Then vapor deposition of 3-APTMS was conducted at 2 Torr for 60 minutes. The water contact angles of the glass slides were 52.5±2.2 degrees after the vapor deposition.
The second set of measurements were made on glass slides cleaned by H2O plasma at 70° C. for 20 minutes. The H2O vapor pressure during plasma cleaning was 200 mTorr, and the plasma power was 250 watts. The water contact angles measured from the glass slides were 6.0 ±0.9 degrees before plasma cleaning, and were 4.0±0.3 degrees after cleaning. After plasma cleaning, water vapor deposition was conducted at 1 Torr for 30 minutes. Then vapor deposition of 3-APTMS was conducted at 2 Torr for 60 minutes. The water contact angles of the glass slides were 42.83±4.0 degrees after vapor deposition.
The third set of measurements were made on glass slides cleaned by plasma generated from room air at 70° C. for 20 minutes. The air pressure during plasma cleaning was 200 mTorr, and the plasma power was 250 watts. The water contact angles measured from the glass slides were 6.3±0.4 degrees before plasma cleaning, and were 4.8±0.6 degrees after cleaning. After plasma cleaning, water vapor deposition was conducted at 1 Torr for 30 minutes. Then vapor deposition of 3-APTMS was conducted at 2 Torr for 60 minutes. The water contact angles of the glass slides were 51.6±1.7 degrees after vapor deposition.
The fourth set of measurements were made on glass slides cleaned by O2 plasma at 70° C. for about 20 minutes. The air pressure during plasma cleaning was 200 mTorr, and the plasma power was 250 watts. The water contact angles measured from the glass slides were 5.5±0.7 degrees before plasma cleaning, and were 4.1±0.5 degrees after cleaning. For this measurement, water vapor deposition was not used. After plasma cleaning, the vapor deposition of 3-APTMS was conducted at 2 Torr for about 60 minutes. The water contact angles of the glass slides were 52.5±2.2 degrees after vapor deposition.
The fifth set of measurements were made on glass slides cleaned by O2 plasma at 70° C. for 20 minutes. The air pressure during plasma cleaning was 200 mTorr, and the plasma power was 250 watts. The water contact angles measured from the glass slides were 6.6±0.6 degrees before plasma cleaning, and were 5.6±0.8 degrees after cleaning. After plasma cleaning, water vapor deposition was conducted at 1 Torr for 30 minutes. Then vapor deposition of GPTMS was conducted at 450 mTorr for 60 minutes. The water contact angles of the glass slides were 54.2±1.5 degrees after vapor deposition.
The sixth set of measurements were made on glass slides cleaned by H2O plasma at 70° C. for 20 minutes under air pressure of 200 mTorr with 250 Watts of plasma power. The water contact angles measured from the glass slides were 4.4±0.7 degrees after cleaning. After plasma cleaning, water vapor deposition was conducted at 1 Torr for 30 minutes. Then vapor deposition of GPTMS was conducted at 450 mTorr for 60 minutes. The water contact angles of the glass slides were 54.4±0.9 degrees after vapor deposition.
The seventh set of measurements were made on glass slides cleaned by air plasma at 70° C. for 20 minutes under air pressure of 200 mTorr with 250 Watts of plasma power. The water contact angles measured from the glass slides were 5.7±0.7 degrees after cleaning. After plasma cleaning, water vapor deposition was conducted at 1 Torr for 30 minutes. Then vapor deposition of GPTMS was conducted at 450 mTorr for 60 minutes. The water contact angles of the glass slides were 56.3±1.9 degrees after vapor deposition.
The eighth set of measurements were made on silicon wafers cleaned by O2 plasma at 70° C. for 20 minutes under air pressure of 200 mTorr with 250 Watts of plasma power. The water contact angles after cleaning were less than 4 degrees. After plasma cleaning, water vapor deposition was conducted at 1 Torr for 30 minutes. Then vapor deposition of 3-APTMS was conducted at 450 mTorr for 60 minutes. The water contact angles of the silicon wafers were 57.6±0.1 degrees after vapor deposition.
The ninth set of measurements were made on silicon wafers cleaned by H2O plasma at 70° C. for 20 minutes under air pressure of 200 mTorr with 250 Watts of plasma power. The water contact angles measured from the silicon wafers were less than 4 degrees after cleaning. After plasma cleaning, water vapor deposition was conducted at 1 Torr for 30 minutes. Then vapor deposition of GPTMS was conducted at 450 mTorr for 60 minutes. The water contact angles of the silicon wafers were 58.6±0.1 degrees after vapor deposition.
A number of embodiments of the invention have been described. Nevertheless, it will be understood that various modifications may be made without departing from the spirit and scope of the invention. For example, the chamber 102 can be cube or cylindrically shaped, and can have varying sizes. The electrodes 106 may be square or round shaped, internal or external to chamber 102, and either inductive or capacitive in the coupling of plasma energy to the chamber. Devices other than an oven may be used to heat the organosilane solutions and water. For example, a heating coil or heating pad may be used. Vessels 116 and 122 are shown coupled to the chamber through inlet 114. They may also be coupled to the chamber through separate inlets. Likewise, additional vessels containing organosilane solutions may be coupled to the chamber through inlet 114 or other inlets. For system 400, various means can be used to move the substrates from the first chamber to the second chamber. For example, a rotatable plate may be used to rotate the substrates from the first chamber to the second chamber. A slidable plate may also be used to slide the substrates from one chamber to another chamber.
Different types of organosilanes additional to the ones mentioned may be used to silanize the cleaned glass slides. Substrates may be composed of materials other than glass, such as quartz, ceramic, silicon, metal, or polymer, and may include additional materials. Substrates may be of various shapes and may have various layers as long as it has a surface that allows a silane condensation reaction to occur. The substrate may be part of a larger device. The temperature and pressure conditions may be different from the ones described may be used as long as plasma cleaning and silanization can occur. The silanizing step may be performed with or without cleaning the chamber. In the steps where water deposition is used, hydrogen peroxide deposition may also be used.
Accordingly, other embodiments are within the scope of the following claims.
This application is a continuation-in-part of, and claims priority to, U.S. application Ser. No. 10/113,076, filed Apr. 1, 2002.
| Number | Date | Country | |
|---|---|---|---|
| Parent | 10113076 | Apr 2002 | US |
| Child | 10885204 | Jul 2004 | US |
