Note: Descriptions are shown in the official language in which they were submitted.
CA 02523896 2005-10-19
Nan~c~ntact Printing
Back ound
1n recent years, there has been considerable effort aimed at understanding new
phenomena in the nanoscale, a diversity of new nanostructured materials have
been
fabricated and characterized. New devices with intriguing properties we just
begim~ing to
be engineered. The expectations for a new generation of cheap and innovative
tools that
will change our lives are very high. The combination of a new set of expected
and
unexpected properties together with a whole new family of materials and
fabrication
methods will enable devices that we could not even have conceived just ten
years ago.
Coulomb blockade in metal nanoparticles as well as in semiconductor quantum
dots,
narrow band fluorescence emission from semiconductor nanoparticles, quantized
ballistic
conduction in nanowires and nanoiubes: these are just a few new
materials/phenomena
that will have an impact on the way we design optical and electroiucs devices.
For a
I5 review of nanodevices and fabrication techniques, see Bashir, Superluttice
and
Microstructures (2001), ~9(1);l-16; Xia, et al., Cheraa. Rev. (1999), 99;1823-
1848; and
Gonsaives, et al., Advanced Materials (2001), 13( 10):703-714, the entire
teachings of
which are incorporated herein by reference.
Nanoscience development, similarly to many other branches of science, but
probably in a more extreme way, relies on state-of the-art techniques for the
imaging and
fabrication of iIs tools. Undoubtedly, the development ofthe transmission
electron
microscope (TEM) and scanning tunneling microscope (STM) gave birth to the
whole
field of nanoscience. While the development of electron-beam (e-beam)
lithography,
being the first tool that could build structure and devices in the nanometer
scale, made the
field of nanotechnology a reality.
The first stages of nanoscience and mainly of nanoteclmology have been
dominated by the development and characterization of new materials and devices
based
on inorganic semiconductors and metals. One of the main reasons for this is
that e-beam
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CA 02523896 2005-10-19
lithography is a technique to pattern inorganic materials on an inorganic
substrate. A
significant advancement in recent years has been the development of novel
highly
versatile nanolithographies based on scanning probe microscopes (SPM). Using
various
types of SPMs a wide variety of organic and inorganic substrates can now be
patterned
either by inducing localized chemical modifications or by forming self
assembled
monolayers (SAMs). For example, Mirkin and coworkers have developed an atomic
force microscope (AFM)-based technique (Dip Pen Nanolithography, DPN) in which
a
SAM can be generated by controlled transfer of molecules from the microscope
tip to a
substrate, v~~ith resolution below 5 nm (see Lee, et al., Science (2002),
295:1702-1705;
Demers, et al., Ar~gew. Chem. Int. Ed. (2001), 40(16):3069-3071; Hong, et al.,
Scfence
(1999), 286:523-525; Piner, et al., Science (1999), 283:66i-663; Deniers, et
al., Arzgew.
Clzelaz. Int. Ed (2001), 40(16):3071-3073; Deniers, et al., Science (2002),
2.96:1836-1838,
U.S. Patent Application Publication Nos. 2002/0063212, 2003/0049381,
200310068446,
and 2003/0157254., the entire teachings of which are incorporated herein by
reference).
The development of such techniques represents a major breakthrough, as now it
is
possible to build devices based not only on inorganic but also on organic and
biomaterials. Organic based nanomaterials are likely to offer a number of
interesting
properties that can be effectively modulated an the nanoscale. Furthermore,
the typical
disadvantages of orgazuc materials are less important in nanodevices; far
example, there
is less need for good mechanical properties or high thermal stability. Thanks
both to these
novel fabrication techniques and to the elucidation of basic concepts in
surface and supra-
moIecular chenvstry, navel devices are currently well under development.
Using orgazue and inorgazuc based nano-lithography techniques many different
nano-devices (e.g nano-transistors, nana-sensors and nano-waveguides) are
presently
being fabricated. However, in order to predict how great an impaci
nanotechnology will
have, one must estimate the speed of fabrication for complex devices.
Unfomznately, all
nano-lithographies have in common the same drawback: tlzey are extrenzely
slow, and it
has been postulated that device fabrication time (and reproducibility) will be
the main
limiting factor in nanotechnology. In particular, the problem of how to scale
up
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CA 02523896 2005-10-19
production has not been solved. Addressing the problem of production scale-up
is critical
if we hope to see the enormous amount of knowledge that we are now acquiring
translated into transistors, sensors, antennas, lenses and drug delivery
systems to use in
everyday life.
It would be desirable for nanotechnology to have an equivalent of micro-
contact
printing: this stamping technique engineered by ~Yhitesides and coworkers (see
U.S.
Patent Nos. 5,512,131, 5,900,160, 6,048,623, 6,180,239, 6,322,979, 6,518,168,
the entire
teachings ofwhich are incorporated herein by reference) has revolutionized the
way
people design micro-devices and has had an enormous impact in allowing non-
chemist to
build devices as complex as bio-MEMS. Unfortunately, micro-contact printing
has
serious resolution linutations, so its application in nanotechnology is
limited.
The only research efforts to directly address this problem is that by Chvu and
coworkers at Princeton. In a recent Hewlett Packard press release
("breakthrough in
nal~o-electronics"), their patents and patent applications on nano-imprinting
(U.S. Patent
Nos. 5,772,905 and 6,309,580, and U.S. Patent Application Publication Nos.
200210167117, 2003/0034329, 200310080471, and 200310080472, the entire
teachings of
which are incorporated herein by reference) were considered one of the
fundamental
steps towards the realization of nano-transistors. The method is based on a
hard mold
(i.e., a mold made of an inorganic material) that is stamped an a soft polymer
film
overcoating a silicon wafer. The printed substrates typically consist of
metallic wires or
semiconductor materials (see C,'hou, et al., Nature (2002), 417:835-837; and
Austin, et
al., J. Yac. Sci, Technol. ~ (2002), 20(2):665-667, the entire teachings of
wlueh are
incorporated herein by reference). As with many other fundamental aspects of
nanotechnology, the fabrication methods for inorganic materials are preceding
those for
organic materials. In fact, the main lunitatiom to nano-imprint is that it
needs a "hwd"
mold and that it is tailor-made to print a shape on a sificon wafer. It is
difficult to
envision how such a method could be adapted for soft molds (i.e., a mold made
of an
organic material) and/or how it could be used to transfer the high degree of
complexity
and information that an organic (particularly a bio-organic) substrate can
carry.
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CA 02523896 2005-10-19
A major drawback of existing nanolithography techniques for fabricating
nanoscale devices is that features of the device must be fabricated in a
series of steps.
Thus, these techniques are limited to relatively simple devices since the
fabrication of
devices having many features would take a prohibitive amount of time. The only
major
effort to address this problem is the fabrication of multi-tip arrays for SPMs
(Zhang, et
al., Nanoteclznology (2002), 13:212, the entire teachings of which are
incorporated herein
by reference). While such approaches will mainly enable the parallel
fabrication of a
perhaps tens or hundreds of nano-devices, it would be desirable to develop a
nanoscale
stamping technique that could complement such parallel device production, and
move it
toward mass-production by developing a method that can produce many features
in a
parallel manner on a device in a single processing step.
Snmmary of the Invention
'fhe method of the invention allows multiple features of a nanoscale devices
to be
fabricated at the same time. Iu one aspect of the invention, the method
involves forming
a complement image of a master. The master used in tlus embodiment of the
method of
the invention comprises a first set of molecules bound to a first substrate to
form a
pattern. The master is contacted with a second set of molecules that
assembling via
attractive forces or via bond formation on the first set of molecules, Each
molecule in the
second set of molecules comprises a reactive functional group and a
recognition
component that is attracted to or binds to at least a portion ofone or more of
the fwst set
of molecules. The reactive functional goup of the second set of molecules is
then
contacted with a surface of a second substrate. The surface of the second
substrate reacts
with the reactive functional group of the second set of molecules to form a
bond between
the second set of molecules and the second substrate. The attractive force
bet~.veen the
first sei of molecules and the second set of molecules is then broken, and the
second set
of molecules bound to the second substrate forms a complement image of the
master.
Once the master has been separated from the complement image by breaking the
bonds
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CA 02523896 2005-10-19
between the first and tl2e SeCOnd Set of molecules, the master can be reused
one or more
times to forth additional complement images.
In another aspect of the invention, the method involves forming a reproduction
of.
a master, or a portion thereof The master used in this embodiment of the
method of the
invention comprises a first set of molecules bound to a first substrate to
form a pattern. A
second set of molecules is assembled on the first set of molecules via bond
formation.
The second set of molecules comprises a reactive functional group and a
recognition
component that binds to at least a portion of one or more molecules from the
first set of
molecules. The reactive functional group of the second set of molecules is
then contacted
with a surface of a second substrate. The reactive functional group reacts
with the
surface of the second substrate to form a bond between the second set of
molecules aid
the second substrate. The bonds between the first set of molecules and the
second set of
molecules are then broken, and the second set of molecules bound to the second
substrate
fours a complement image of the master. A third set of molecules is then
assembled via
1S bond formation on the second set of molecules of the complement image. Each
molecule
in the third set of molecules comprises a reactive functional group, and a
recognition
component that binds to the second set of molecules. The reactive functional
group of
the third set of molecules is then contacted with a surface of a third
substrate. The
surface of the third substrate reacts with the reactive functional group of
the third set of
molecules to form a bond between the third set of molecules and the third
substrate. The
bonds between the second set of molecules and the third set of molecules are
then broken,
and the third set of molecules bound to the third substrate form a
reproduction of the
pattern, or portion thereof, of the master. Once the complement image has been
separated from the reproduction, the complement image can be reused one or
more times
to form additional reproductions.
In another aspect, the invention relates to a composition comprising a master
having a pattern of a first set of molecules bound to a first substrate; and a
complement
image comprising a pattern of a second set of molecules bound to a second
substrate via a
reactive functional group on each molecule of the second set of molecules. In
this aspect
Page 5
CA 02523896 2005-10-19
of the invention, each molecule in the second set of molecules has a
recognition
component that binds to at least a portion of a molecule from the first set of
molecule.
In another aspect, the invention relates to a composition, comprising a first
pattern
of a first set of molecules bound to a first substrate; and a second pattern
of a second set
of molecules bound to a second substrate via a reactive functional group on
each
molecule of the second set of molecules. In this aspect of the invention, each
molecule of
the second set of molecules comprises a recognition component that binds to at
least one
molecule in the fzxst set ofmoleeules, and the second pattern is a complement
image of
the first pattern.
I 0 Tn another aspect, the invention relates to a composition, comprising a
first pattern
of a first set of molecules bound to a first substrate; and a second
substrate, wherein the
second substrate comprises a degraded portion and an undegraded portion, and
the
undegraded portion is a complement image of the fZrst pattern.
In another aspect, the invention relates to a composition, comprising a first
pattern
I S of a first set of molecules bound to a first substrate; and a second
substrate having a
patterned layer of a material deposited thereon, wherein the patterned layer
of deposited
material on the second substrate is a complement image of the &rst pattern.
In another aspect, the invention relates to a kit for printing a molecular
pattern on
a substrate. The kit comprises a master comprising a pattern of a first set of
molecules
20 bound to a substrate; and a second set of molecules. The second set of
molecules
comprises a reactive functional group; and a recognition component that binds
to the fZrst
set of molecules.
In another aspect, the invention relates to a molecular printer for generating
a
complement image of a master, wherein the master has a first set of molecules
bound to a
25 first substrate. The molecular printer comprises a device for delivering a
solution of a
second set of molecules to a surface of the master, and a device for
contacting the second
set of molecules with a second substrate. In this embodiment, the second set
of
molecules comprises a reactive functional group; and a recognition component
that binds
to the first set of molecules.
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CA 02523896 2005-10-19
The method of the invention complements all the chemically oriented
nanolithography techniques that have been developed in recent years and which
generally
require complex instrumentation. For example, it has already been shown that
DNA
testing arrays can be fabricated using Dip Pen Nanolithography. Once the
master for
these devices is built, the method of the invention, that does not require
complex
instrumentation and materials, can be used to print a large number of cheap
and
extremely sensitive devices for the detection of, for example, biohazards. By
way of
example, Iet's assume that such a device covers an area of 3 mm~. From a
single mold
built on a silicon wafer (~ 106 mmz), one million sensors for a specific bio-
molecule
(such as anthrax) could be fabricated in approximately 3 hours using the
method of the
invention. Because the transfer process is self assembly based, all the steps,
besides the
fabrication of the master, can be done in parallel over very Iarge areas and
on multiple
substrates.
The amount of information stored in a molecule, such as a DNA strand, can be
I S enormous. The method of the invention has the possibility of transferring
this
information in a massively parallel way (i.e., in one or only a few step
instead of. many
steps). Thus devices that are now built using rnulti-step techniques could be
fabricated in
a single step. This opportunity will redirect research and device manufacture
towards
increasing complexity in .fabricated substrates. As a simple example, if a
master were
fabricated on a lnunz substrate having a series of nano and microfluidic
channels (e.g.,
50) having 50 different types of DNA. strands defining the walls of the
channels, in one
single printing step with the method of the invention, one could fabricate on
a 1 rrnnz
substrate a complement image of the series of nano and microfluidic channels,
each with
the wall functionalized in a different way: a real lab on a chip. This is not
possible with
any current fabrication method, which would require 50 consecutive steps.
A unique feature of the method of the invention is the ability to copy, and
thus
replicate, the master itself using the parallel method of the invention
instead of the
methods of the prior art in which printed items cannot act as a master. This
is a major
advantage over a.ny existing methods. In fact, typically for large production
Iines many
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CA 02523896 2005-10-19
masters are needed. This, combined with the wearing of existing molds, means
that a
canstant production of masters is required. Tn the method of the invention,
once a master
is produced, reproductions of the master can be produced from it, and these
new master
will then be used to print the final devices. Reproducibility should be
improved and, more
importantly the instruments for the primary master fabrication which produce
features in
a serial fashion will have to be used only to fabricate the primary master.
The method of the invention is revolutionary not only because it can be used
to
print organic SAMs, but because the method can be used to transfer multiple
types of
information (e.g., chemical + shape) and to reproduce a master in a parallet
fashion.
Brief Descr~tion of the Drawing
The izivention is described with reference to a particular embodiment shown in
the
figures. The embodiment in the figures is Shawn by way of example and is not
meant to
be limiting in any way.
IS Figs. lA-D are a schematic representation of one embodiment of the method
of
the invention for producing a complement image.
Fig. 2 is a schematic representation of a first set of molecules bound to a
second
set of molecules.
Figs. 3A and 3B are AFM images of a master having a monolayer of nucleic acid
molecules bound to the surface of a substrate.
Fig. 3C is an AFM image of a complement image of the master spawn i11 Fig. 3A.
Fig. 3D is an AFM image of a complement image of the master shov~nl in Fig.
3B.
Fig. 4A is an AFM image of a master having nucleic acids bound to a substrate
in
a grid pattern.
Fig. 4B is an AFM image of a complement image of the master shown in Fig. 4A.
Page 8
CA 02523896 2005-10-19
Definitions
A "master," as used herein, is a substrate that has a first set of molecules
bound to
a surface of the substrate in a random or non-random pattern, Preferably, the
first set of
molecules are bound to the master in a non-random pattern. The first set of
molecules
,5 may include one or more different molecules. The information encoded in the
pattern
may be from the position of each of the molecules on the surface of the
substrate andlor
the chemical nature of the molecule (e.g., a molecule from the first set of
molecules
having a particular nucleic acid sequence will bind specifically to a nucleic
acid molecule
having a complementary sequence).
A "complement image of a master," as used herein, is an image on a substrate
that
is a mirror image, when the pattern on the master is asynmetiical, or a copy,
when the
pattern on the master is symmetrical, of the spatial andlor chemical
information encoded
in the master, or a portion thereof. In one embodiment, the complement image
is formed
by binding a second set of molecules to a second substrate. For example, if
the first set of
I S molecules bound to the master are nucleic acid molecules that form a non-
centrosymmetric pattern, a complement image of the master will be a mirror
image of the
master foamed on a second substrate with a second set of molecules that are
nucleic acids
that have a sequence that is complementary to at least a portion of a nucleic
acid
sequence from the first set of molecules. Typically, the chemical information
transferred
to the complement image is not identical to the inforn~ation on the master but
is enough
information to allow at least a portion of the information from the master to
be
reproduced. ror example, when the first and second sets of molecules are
nucleic acid
molecules, at least three or more consecutive bases of a molecule from the
first set of
molecules must be cornplementaty three or more consecutive bases from the
second set
of molecules. A complement image can be funned from a portion of the pattern
on the
master by selecting molecules for the second set of molecules that only bind
to a portion
of the molecules of the first set of molecules that are bound to the master.
When the
second set of molecules binds only to a portion of the first set of molecule,
the height
profile of the complement image may have t~.~o or more levels, In addition, a
complement
Page 9
CA 02523896 2005-10-19
image may encode only a mirror image of the spatial infotrnation encoded in
the master
or may encode both the chemical and spatial information encoded in the master.
For
example, if the first set of molecules bound to the master are nucleic acid
molecules that
form an asymmetric pattern, a complement image of the master will be a mirror
image of
the master formed an a second substrate with a second set of molecules that
are nucleic
acids that have a sequence that is complementary to at least a portion of a
nucleic acid
sequence from the first set of molecules. In this example, both spatial and
chemical
information are transferred from the master to the complement image.
Furthermore, only
a portion of the chemical infotrnation may be transferred to the complement
image. For
example, when the first set of molecules on the master are nucleic acid
molecules, the
second set of molecules that form the complement image may be nucleic acid
sequences
that are complementary to only a portion of a nucleic acid sequence on the
master (i.e., is
not complementary to the whole sequence).
A "reproduction of a master," as used herein, is copy of the spatial andlor
chemical information encoded in a pattern of a master. The reproduction may be
a copy
of only a portion of the pattern of the master or may be a copy of the entire
pattern of the
master. 1n addition, a reproduction of a master may copy only the spatial
information of
the toaster or may copy both the spatial and chemical information encoded in
the master.
In addition, a reproduction of a master may reproduce only part of the
chemical
information.
"Chetnieal information encoded in a molecule" refers to the ability of the
molecule to bind specifically to another molecule or to a specific type of
molecule,
typically, in a specific conformation. For example, a particular nucleic acid
sequence
binds specifically to a complementary sequence; or protein A binds
specifically to
imntunoglobulins.
The term "pattern," as used herein, refers to the spatial location of each
molecule
in a set of molecules bound to a substrate, and the chemical structure of each
molecule in
the set of molecules.
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CA 02523896 2005-10-19
The term "reactive functional group," as used herein, is a group that can
react to
form a bond with a surface of a substrate. Examples of reactive functional
groups include
thiol groups or a protected thiol group, which can bind to surfaces made of
gold, silver,
copper, cadmium, zinc, palladium, platinum, mercury, lead, iron, chromium,
manganese,
tungsten, or any alloys thereof. Protected thiol groups can be deprotected
before they can
bind to the substrate surface. Methods of protecting and deprotecting thiol
groups can be
found in Greene and Wuts, "Protective Groups in Organic Synthesis", John Wiley
&
Sons (I991), the entire teachings of which are incorporated into this
application by
reference. Another example of reactive functional groups is a silane or a
chlorosilane,
which can bind to a surface of doped or undoped silicon. .Another example of a
reactive
functional group is a carboxylic acid, which can bind to a sw:face that is an
oxide, such as
silica, alumina, quartz, or glass. Another example of reactive functional
groups are
nitrites and isonitriles, which can bind to a surface of platinum, palladium
or auy alloy
thereof. Another example of a reactive functional group is a hydroxamic acid,
which can
bind to a copper surface.
When a set of molecules binds to the surface of a substrate, the molecules
will
fold or staelc against one another such that a portion of the molecule will be
exposed on
the surface of the substrate. The exposed functionality may be hydrophobic,
hydrophilic,
or an amphipathic functionality. Tn addition, the exposed functionality rnay
be a
fi~nctionality that selectively binds various biological or other chemical
species such as
proteins, antibodies, antigens, sugars and other carbohydrates, and the like.
The exposed
functionality may comprise a member of any specific or non-specific binding
pair, such
as either member of the following non-limiting list: antibodylantigen,
antibodylhapten,
enzymelsubstrate, enzyme/inhibitor, enzymelcofactor, binding
protein/substrate, carrier
prateinlsubstrate, lectinlcaibohydrate, receptorlhorrnone, receptorleffector,
complementary strands of nucleic acid, repressorlinducer, or the like.
Examples of
exposed functionalities include -OH, -CONH-, -CONHCO-, -NH2, -IVIi-, -COON,
-COOR, -CSNH-, -IV'02 , -SOz, -SH, -RCOR-, -RCSR-, -RSR, -ROR-, -PO4 3, -
OS03'2,
-S03-, -COO , -SOO , -RSOR-, -CONR2, -(OCHZCHZ)~OH (where n=1-~0, preferably I-
Page 11
CA 02523896 2005-10-19
8), -CH3, -P03H , -2-imidazole, -N(CH3)Z, -N(R)2, -P03Hz, -CN, -(CFZ)"CF3
(where n=1-
20, preferably I-$), and an olefin. R is hydrogen, a hydrocarbon, a
halogenated
hydrocarbon, a protein, an enzyme, a carbohydrates, a lectin, a hormone, a
receptor, an
antigen, an antibody, or a hapten.
The term "silane," as used herein, refers to a functional group having the
following structural formula:
R2
Si--ORz
IIORZ
R2 in the above structural formula, for each occurrence, is independently
selected from
the group consisting of-H, an alkyl, an aryl, an alkenyl, an alkynyl, and an
arylalkyl.
The term "chlorosilane," as used herein, refers to a functional group having
the
following stnictural formula:
Rs
i
Rs
Rs
R6 in the above structural formula, for each occurrence, is independly
selected from -Cl
or-0R2, provided that at least one of Rb is -Cl. Preferably, each Rb is --Cl.
The term "spacer," as used herein, refers to a divalent group that connects
two
components of a molecule. Prefers-ed spacers include allcylene, a
heteroalkylene, a
heterocycloalkylene, an aIkenylene, an allcynylene, an arylene, a
heteroarylene, an
arylalkylene, and a heteroarylalkylene, whexein the alkylene, heteroalkylene,
heterocycloalkylene, alkenylene, alkynylene, arylene, heteroarylene,
arylalkylene, or
heteroarylallcylene may be substituted or unsubstituted.
Page 12-
CA 02523896 2005-10-19
The tenn "alkyl," as used herein, means a straight chained or branched C~-Czo
hydrocarbon or a cyclic Cs-Cao hY~oc~bon that is completely saturated. Alkyl
groups
may be substituted or unsubstituted.
The term "alkylene" refers to an alkyl group that has at least two points of
S attachment to at least two moieties (e.g., methylene, ethylene,
isopropylene, etc.).
Alkylene groups may be substituted or unsubstituted.
"AIkenyl groups" are straight chained or branched Cz-Czo hydrocarbon or a
cyclic
C3-CZO hydrocarbon that have one or more double bonds. Alkenyl groups may be
substituted or unsubstituted.
An "alkenylene" refers to an all~enyl group that has two points of attachment
to at
least two moieties. Allcenylene groups may be substituted or unsubstatuted.
"Alkynyl groups" are straight chained or branched CZ-Czo hydrocarbon or a
cyclic
C3-Czo hydrocarbon that have one ox more triple bonds. Aiky~iyl groups may be
substituted or unsubstituted.
An "alkynylene" refers to an alkynyl group that has two points of attachment
to at
least two moieties. AlkynyIene groups may be substituted or unsubstituted.
An "heteroalkyIene" refers to a group having the formula -X-((aIkylene)-X}q-,
wherein X is -0-, -NR~-, or-S-; and q is an integer farm 1 to 10. R.~ is a
hydrogen, alkyl,
aryl, arylalkyl, alkenyl, alkynyl, heteroaryl, heteroarylallcyl, or
heterocycloalkyl.
Heteroalkylene groups may be substituted or unsubstituted,
The term "aryl," as used herein, either alone or as pait of another moiety
(e.g.,
arylalkyl, etc.), refers to carbocyclic aromatic groups such as phenyl. Aryl
groups also
include fused polycyclic aromatic ring systems in which a carbocyclic aromatic
ring is
fused to another car6ocyclic aromatic ring (e.g., 1-naphthyl, 2-naphthyl, 1-
anthracyl, 2-
25 anthracyl, etc.) or in which a carbocylic aromatic ring is fused to one or
more carbocyelie
non-aromatic rings (e.g., tetrahydronaphthylene, indan, etc.). The point of
attachment of
an arylene fused to a earboeyclie, non-aromatic ring may be on either the
aromatic, non-
womatic ring. Aryl groups may be substituted or unsubstihited.
An "arylene" refers to an aryl group that has at least two points of
attachment to at
Page 13
CA 02523896 2005-10-19
Least two moieties (e.g., phenylene, etc.). Arylene groups may be substituted
or
unsubstituted.
An "arylalkyl" group refers to an aryl group that is attached to another
moiety via
an alkylene linker. Arylalkyl groups may be substituted or unsubstituted. When
an
arylalkylene is substituted, the substituents may be on either the aromatic
ring or the
alkylene portion of the arylallcyl.
An "arylalkylene group," as used herein, refers to an arylaLkyl group that has
at
least two points of attachment to at Least fivo moieties. The second points of
attachment
can be on either the aromatic ring or the alkylene. An arylallcylene may be
substituted or
unsubstituted. When an arylalkylene is substituted, the substifuents may be on
either the
aromatic ring or the alkylene portion of the arylalkylene.
The term "heteraaryl," as used herein, means an aromatic heterocycle which
contains 1, 2, 3 or 4 heteroatoms selected from nitrogen, sulfur ar oxygen. A
heteroaryl
may be fused to one or two rings, such as a cycloalkyl, a heterocycloalkyl, an
aryl, or a
hetexoaryl. The point of attachment of a heteroaryl to a molecule may be on
the
heteroaryl, cycloallcyl, heterocycloalkyl or aryl ring, and the heteroaryl
group maybe
attached thrnugh carbon or a heteroatom. Examples of heteroaryl groups include
imidazolyl, furyl, pyrrolyI, thienyl, oxazolyl, thia.zolyl, isoxazolyl,
thiadiazolyl,
oxadiazolyl, pyridinyl, pyrimidyI, pyrazinyl, pyridazinyl, quinolyl,
isoquniolyl, indazolyl,
benzoxazolyl, benzofuryl, benzothiazolyl, uzdolizinyl, imidazopyridinyl,
p~~razolyl,
ti-iazolyl, isotluazolyl, oxazolyl, tetrazolyl, benzimidazolyl, benzoxaaolyl,
benzothiazolyl,
benzothiadiazolyl, benzoxadiazolyl, indolyl, tetrahydroindolyl, azaindolyl,
imidazopyridyl, qunizaolinyl, purinyl, pyrrolo[2,3]pyrimidyl,
pyrazolo[3,4]pyrimidyl or
benzo(b)thienyl each of which is optionally substituted. Heteroaryl groups may
be
substituted or unsubstituted.
A "heteroarylene" refers to an heteroaryl group that has of least two points
of
attachment to at least two moieties. Heteroarylene groups may be substituted
or
unsubstituted.
A "heteroarylalkyl group" refers to an heteroaryl group that is attached to
another
Page 14
CA 02523896 2005-10-19
moiety via an alkylene linker. Heteroarylalkyl groups may be substituted or
unsubstituted. When a heteroarylalkylene is substituted, the substituents may
be on either
the aromatic ring or the alkylene portion of the heteroarylalkyl.
Heteroarylalkyl groups
may be substituted ar unsubstituted.
A "heteroarylalkylene" refers to an heteroarylalkyl group that has at least
two
points of attachment to at least two moieties. HeteroaryIalkylene groups may
be
substituted or unsubstituted.
A "heterocycloalkyl" refers to a non-aromatic ring which contains one or more,
far example., one to four, oxygen, nifxogen or sulfur (e.g., morpholine,
piperidine,
piperazine, pyrrolidine, and thiomvrpholine). Heterocycloalkyl groups may be
substituted or unsubstituted.
A "heterocycloalkylene" refers to a heterocycloalkyl that has at least two
points of
attachment to at least two moieties. Heterocycloalkylene groups may be
substituted or
unsubstituted.
Suitable substituents for an alkyl, an alkylene, an alkenyl, an alkenylene, an
alIcynyl, an alkynylene, a heteroallcyl, a heteroalkylene, a heterocycloalkyl,
a
heterocycloallcylene group, an aryl, an arylene group, an arylalkyl, an
arylalkylene, a
heteroaryl, a heteroarylene, a heteroarylalkyl, and a heteroaryallcylene
groups include my
substituent that is stable under the reaction conditions used in the method of
the
2~ invention. Examples of substituents include an aryl (e.g., phenyl), an
arylalkyl (e.g.,
bencyl), nifro, cyano, halo (e.g., fluorine, chlorine and bromine), alkyl
(e.g., methyl,
ethyl, isopropyl, cyclohexyl, etc.) haloalkyl (e.g., trifluoromethyl), alkoxy
(e.g., methoxy,
ethoxy, etc.), hydroxy, -NR3R,, -NR3C(O)R5, -C(0)NR3Rd, -C(O)R3, -C(O)ORS,
-OC(O)R5, wherein R3 and R,~ for each occurrence are, independently, -H, an
allcyl, an
aryl, or an arylalkyl; and RS for each occurrence is, independently, an alkyl,
an aryl, or an
arylalkyl.
Alkyl, alkylene, heterocycloalkylene groups, and any saturated portion of
alkenyl,
alkenylene, alk}myl, alkynylene groups, may also be substituted with = 0 and
=S.
When a heteroalkylene, a heterocycloalkyl, a heterocycloalkylene, a
heteroaryl,
Page 15
CA 02523896 2005-10-19
or a heteroarylene group contain a nitrogen atom, it may be substituted or
unsubstituted.
When a nitrogen atom in the aromatic ring of a heteroaryl or a heteroarylene
group has a
substituent, the nitrogen may be a quaternary nitrogen.
The term "nucleic acids," or "oligonucleotides," as used herein, refers to a
polymer of nucleotides. Typically, a nucleic acid comprises at least three
nucleotides.
The polymer may include natural nucleosides (i.e., adenosine, thymidine,
guanosine,
cytidine, uridine, deoxyadenosine, deoxythymidine, deoxyguanosine, and
deoxycytidine)
ar modified nucleosides. Examples of modified nucleotides include base
modified
nucleoside (e.g., aracytidine, inosine, isoguanosine, nebularine,
pseudouridine, 2,6-
diaminopurine, 2-aminopurine, 2-thiothymidine, 3-deaza-S-azacytidine, 2'-
deoxyuridinc,
~-nitorpyrtole, 4-methylindole, 4-thiouridine, 4-thiothymidine, 2-
aminoadenosine, 2-
thiothymidine, 2-thiouridine, S-bromocytidine, 5-iodouridine, inosine, 6-
azauridine, 6-
chloropurine, 7-deazaadenosine, 7-deazaguanosine, 8-azaadenosine, 8-
azidoadenosine,
benzimidazole, M1-methyladenosine, pyrrolo-pyrimidine, 2-amino-6-chloropurine,
3-
methyl adenosine, S-propynylcytidine, S-propynyluridine, 5-bromouridine,
5-fluarouridine, 5-methylcytidine, 7-deazaadenosine, 7-deazaguanosine, 8-
oxoadenosine,
8-oxoguanosine, 0(6)-methylguanine, and 2-thiocytidine), chemically or
biologically
modified bases (e.g., methylated bases), modified sugars (e.g., 2'-
fluororibose, 2'-
aminoribose, 2'-azidoribose, 2'-0-methylribose, L-enantiomeric nucleosides
arabinose,
and hexose), modified phosphate groups (e.g., phosphorothioates and S'
-N-phosphoramidite linkages), and combinations thereof. Natural and modified
nucleotide monomers for the chemical synthesis ofnucleie acids are
cammercially
available.
The term "peptide nucleic acid (PNA)," as used herein, refers to a polymer
that
has a peptide backbone in which a natural or nvn-natural nucleic acid base is
attached to
each amino acid residue. Peptide nucleic acids are described in Hanvey, et
al., Science
(1992), 258:1481-1485, the entire teachings of which are incorporated by
reference. A
PNA can bind specif cally to a nucleic acid or another PNA that has a
complementary
sequence of at least three consecutive bases, preferably six consecutive
bases, to the
Page 16
CA 02523896 2005-10-19
sequence of the PNA.
The term "attractive force," as used herein, is a force that draws two or more
molecules together. Examples of attractive forces include attraction of a
molecule having
a net positive charge to a molecule having a net negative charge, dipole-
dipole attraction,
and magnetic attraction.
Unless specified as a covalent bond, the term "hind" or "bound" includes bath
covalent and non-covalent associations, such as hydrogen bonds, ionic bonds,
covalent
bonds, and van der Waals bonds.
The Perm "recognition component," as used herein, is a component of a molecule
that can bind specifically to another molecule.
"specific binding," as used herein, is when a recognition component of a
molecule binds one or more other molecule or complex, with specificity
sufficient to
differentiate between the molecule or complex and other components or
contaminants of
a sample. Molecules that include recognition coraponents and their targets are
conventional and are not described here in detail. Techniques for preparing
and utilizing
such systems are well known in the art and are exemplified in the publication
of Tijssen,
P., "Laboratory Techniques in Biochemistry and Molecular Biology practice and
Theories of Enzyme Immunoassays" (198g), eds. F3urdon and Knippenberg, l\ew
York:Elsevier, the entire teachings of which are incorporated herein.
Preferred
recognition components and their targets include nucleic acid/complementary
nucleic
acid, antigenlantibody, antigenlantibody fragment, avidin/biotin,
streptavidinlbiotin,
protein A/Ig, lectinlcarbohydrate and aptamer/target.
As used herein, "aptamer" refers to a non-naturally occurring nucleic acid
that
binds selectively to a Target. The nucleic acid that fonns the aptamer may be
composed of
ZS naturally occurring nucleosides, modifed nucleosides, naturally occurring
nucleosides
with hydrocarbon linkers (e.g., an allcylene) or a polyether linker (e.g., a
PEG linker)
inserted between one or more nucleosides, modified nucleosides with
hydrocarbon or
PEG linkers inserted between one or more nucleosides, or a combination of
thereof. In
one embodiment, nucleotides or modified nucleotides of the nucleic acid ligand
can be
Page 17'
CA 02523896 2005-10-19
replaced with a hydrocarbon linker or a polyether linker provided that the
binding affinity
and selectivity of the nucleic acid ligand is not substantially reduced by the
substitution
(e.g., the dissociation constant of the apta~ner far the target should not be
greater than
about 1 x I 0's Ivl]. The target molecule of a aptamer is a three dimensional
chemical
structure that binds to the aptamer. However, the aptamer is not simply a
linear
complementary sequence of a nucleic acid target but may include regions that
bind via
complementary Watson-Crick base pairing interrupted by other structures such
as hairpin
loops). Targets of aptamers include peptide, polypeptide, carbohydrate and
nucleic acid
molecules,
IO
Detailed Description
The method of the invention involves stamping of molecular patterns andlor
devices based on the reversible self assembly of molecules, particularly
organic
molecules. This method is suitable for the stamping of almost any
nmofabricated device,
inorganic andlor organic, and can be used to transferring a Iarge amount of
information
from one substrate to another. The working principle of this technique is
completely
different from any present nanofabrication technique.
In the method ofthe invention, a master, that includes a substrate having a
frst set
of molecules bound to at least one surface in a pattern, is used to induce the
assembly of a
second set of molecules via reversible supra-molecular chemistry (e.g.,
hydrogen 'bonds,
ionic bands, covalent bonds, van der Waals bonds, or a combination thereof;
then, with
the use of substantially irreversible surface chemistry, the second set of
molecules are
attached to a surface of a substrate and subsequently the reversible bonds
between the
fwst set of molecules and the second set of molecules are broken. The term
"substantially
irreversible," as used herein, means that the second set of molecules are
attached to the
surface of the substrate by bonds that are stable to conditions that will
break the bonds
between the first and the second set of molecules. The method of the invention
uses
supra-molecular bonds as a means far shape-transfer, this avoids the need for
mechanical
contacts, and thus constitutes a major departure from nmo-imprinting developed
by Chou
Page 18
CA 02523896 2005-10-19
and co-workers, This method is tailor-made to transfer organic patterns
reliably. The use
of organic molecules allows a great number of variations and enables the
transfer of
multiple surface features at the same time.
Referring to Fig. 1, in one embodiment, the method of forming a complement
image of a master involves providing a master 10 that comprises a first set of
molecules
12 bound to a first substrate I4 to form a pattern. A second set of molecules
15 is
assembled on the first set of molecules via bond formation. The second set of
molecules
comprises a reactive functional group 18 and a recognition component 20 (not
shown in
Fig. 1 ) that binds to the f rst set of molecules I2 (see Fig, 2 which
provides a blowp of
the second set of molecules bound to the first set of molecules). The reactive
functional
group 18 of the second set of molecules 16 is then contacted with a surface of
a second
substrate 22. The reactive functional group reacts with the surface of the
second
substrate to form a bond between the second set of molecules and the second
substrate.
in one embodiment, the remaining exposed surface of the second substrate may
be further
contacted with another group of molecules 24 that each have a reactive
functional groups,
such as an alkane having a thiol substituent, that can bind to the surface in
order to cover
the exposed surface of the second substrate. The bonds between the first set
of molecules
and the second set of molecules are then broken, and the second set of
molecules bound
to the second substrate forms a complement image of the master 26. Once the
master has
been separated from the complement image by breaking the bonds between the
first and
the second set of molecules, the master can be reused one or more tines to
form
additional complement images. In one embodiment, a lateral dimension of at
Least one
feature of the complement image is less than 200 nm.
In one embodiment, the second set of molecules may also include one or more of
the following components: an exposed functionality 28; a covalent bond or a
first spacer
that links the reactive functional group to the recognition component; and a
covalent
bond or a second spacer that links the exposed functionality to the
recognition
component.
Page 19
CA 02523896 2005-10-19
The second set of molecules may include two or more different molecules. Ror
example, two or more molecules of the second set of molecules may have
different
recognition components, such as different nucleic acid sequences; or two or
more
molecules of the second set of molecules may have both different recognition
S components and different exposed functionalities. Typically, one or more
molecules
from the first set of molecules determines where each of the molecules from
the second
set oErnolecules binds.
In one embodiment, the two or more different molecules of the second set of
molecules form a pattern on the second substrate that has a height profile
that comprises
two or more dep#hs. For example, two or mare molecules of the second set of
molecules
may have either a first spacer, a second spacer or bath a first and a second
spacer that
have different lengths. The difference in the length of the spacers can cause
the
molecular image transferred to the second substrate to have two or more
different depths.
In one embodiment, the second set of molecules is assembled on the first set
of
molecules by contacting the master with a solution comprising the second set
of
molecules. In one method of transferring the pattern on a master to a second
substrate,
the master is held in contact with the second substrate by capillary action of
the solution
containing the second set of molecules. A small mechanical force may also be
applied to
hold the two substrates together. The solution containing the second set of
molecules is
then slowly evaporated causing the master and the second substrate to come
closer
together and facilitating binding of the second set of molecules to the second
substrate.
The bonds formed between the first set of molecules and the second set of
molecules may be hydrogen bonds, ioiuc bonds, covalent bonds, van der Waals
bonds, or
a combination thereof. Preferably, the bonds formed between the first set of
molecules
25 and the second set of molecules are hydrogen bonds. In one embodiment, the
bonds
between the first set of molecules and the second set of molecules are broken
by applying
heat. In another embodiment, the bonds between the first set of molecules and
the second
set of molecules are broken by contacting the bonds with a solution having a
high ionic
strength. In yet another embodiment, the bonds between the first set
ofmolecules and the
Page 20
CA 02523896 2005-10-19
second set of molecules are broken by contacting the bonds with a solution
having a high
ionic strength and applying heat. Alternatively, the bonds between the first
set of
molecules and the second set of molecules are broken by contacting them with a
solution
containing an enzyme that breaks the bonds. Typically, the bonds between the
first set of
molecules and the second set of molecules can be broken without breaking most
of the
bonds between the second set of molecules and the second substrate.
The reactive functional group on the second set of molecules is typically a
group
that can bind to the surface of the second substrate. For example, when the
reactive
functional group on the second set of molecules is a thiol group or a
protected thiol
group, the surface of the second substrate may be gold, silver, copper,
cadmium, zinc,
palladium, platinum, mercury, lead, iron, chromium, manganese, tungsten, or
any alloys
thereof. In another example, the reactive functional group on the second set
of molecules
is a silane or a chlorosilme, and the surface of the second substrate is doped
or undoped
silicon. In another example, the reactive functional group on the second set
of molecules
I S is a carboxylic acid, and the surface of the second substrate is an oxide,
such as silica,
alumina, quartz, ar glass. In another example, the reactive functional group
on the
second set of molecules is a nitrite or an isonitrile, and the surface of the
second substrate
is platinum, palladium or any alloy thereof. In another example, the reactive
functional
group on the second set of molecules is a hydroxamic acid, and the surface of
the second
substrate is copper.
In one embodiment, at least some of the molecules of the first set of
molecules
include a recognition component that binds to the one or more molecules of the
second
set of molecules. .Far example, each of the molecules of the first set of
molecules may
include a recognition component that is a nucleic acid sequence. In one
embodiment,
each of the fzrst set of molecules is a nucleic acid sequence and the
recognition
component of the second set of molecules is a nucleic acid sequence,
Preferably, the
nucleic acid recognition component of each of the second set of molecules is
complementary to at least a portion of a nucleic acid sequence of at Ieast one
of the
molecules from the first set of molecules. For example, three or more
consecutive
Page 21
CA 02523896 2005-10-19
nucleic acid bases, preferably six or more nucleic acid bases, of a molecule
from the
second set of molecules is complementary with three or more consecutive
nucleic acid
bases, preferably six or more nucleic acid bases, of a molecule from the first
set of
molecules. When the second set of molecules is assembled on the first set of
molecules,
the second set of molecules will hybridize with molecules from the first set
of molecules
that have a complementary sequence, or a portian thereof, to the nucleic acid
recognition
component of the second set of molecules. 1n this embodiment, typically, the
first set of
molecules bound to the master are contacted with a solution of the second set
of
maIecules under conditions that promote hybridization. Conditions that promote
hybridization are known to those skilled in the art. A general description of
hybridization
conditions are discussed in Ausebel, F. M., et al., Current Protocols in
Molecular
Biology, Greene Publishing Assoc. and 'Wiley-Interscience, 1989, the teachings
of which
are incorporated herein by reference. Factors such as sequence length, base
composition,
percent mismatch between the hybridizing sequences, temperature and ionic
strength
I S influence the stability of nucleic acid hybrids.
W are embodiment, the first set ofmolecules includes tv~ro or mare different
molecules that have recognition companents that are different nucleic acid
sequences. In
this embodiment, the second set of molecules includes molecules that have a
nucleic acid
sequence, or a portion thereof, that is complementary to at least one of the
molecules of
the fu-st set of molecules. In one embodiment, hydrogen bonds between
hybridized
molecules from the first set of molecules and the second set of molecules are
broken by
contacting the hydrogen bonds with an enzyme. For example, an enzyme from the
helicase faznily of enzymes may be use to break the bonds between hybridized
nucleic
acid molecules. Various helicases have been xeported to dehybridize double
stranded
oligonucleotides. l~or example, E. coIi Rep, E. coli DnaB, E. coli UvrD (also
lmown as
Helicase In, E. coli RecBCD, E, coli RecQ, bacteriophage T7 DNA helicase,
human
RECQL series; WRN(RECQ2), BLM(RECQL3), RECQI~, RECQLS, S. Pombe rqhl,
C. elegance To4Al 1.6 (typically, the helicase name is derived from the
organism from
which enzymes comes). Helieases can be divided into hvo types: 1) heliease
that move
Page 22
CA 02523896 2005-10-19
along the nucleic acid strand in the 3' direction, and 2) helicases that more
along the
nucleic acid strand in the 5' direction one. Typically, the particular type of
helicase used
to break the hydrogen bonds between the hybrided nucleic acids are selected by
considering structural hindrance of the particular hybridized nucleic acids.
Cofactors
S which stabilize single stxanded DNA, such as single stranded DNA binding
protein
(SSB), could be added.
Another method of breaking the bonds between two hybridized nucleic acids
would be to use a restriction endonucIease, which recognizes specific base
sequence and
cleaves both strands at a specific location in the nucleic acid sequence.
Examples of
restriction endanucleases include BarnHI, EcoRI, and BstXI. Other methods of
dehybridazition of nucleic acids using enzymes can be found in Lubert Stryer ,
Biochemistry, 4th Edition; Benjamin Lewin, Gene VII; i~risten Moore Picha and
Srnita
S. Patel, "Bacteriophage T7 DNA Helicase Binds dTTF, Fonns Hexamers, and Binds
DNA in the Absence of Mg2+," J, Biol. Chen:, (1998), Vol. 273, Issue 4.2,
27315-27319;
Sheng C'ui, Raffaella K.lima, AIex Ochem, Daniele Arosio, Artuxo Falaschi, and
Alessandro Vindigni, "Characterization of the DNA-unwinding Activity of Human
RECQ1, a Helicase Specifically Stimulated by Human Replication Protein A," J.
Biol.
Chem. (2003), VoI. 278, Issue 3,1424-1432; Umezu, K., and Nakayama, fI.
(1993), J.
Nfol. Biol,. 230:1145-1150; Nakayama, K., Irino, N., and Nakayama, H., lblol.
Gen.
Genet. (I985), 200:266-271; Kusano, K., Berres, M. E., and Engels, W. R.,
Genetics
( I 999),15:1027-1039; Ozsoy, A. Z., Sekelsky, J. J,, and Matson, S. W.,
Nucleic Acids
Res. (2001), 29:2986-299, the entire teachiilgs of these references is
incozporated herein
by reference.
Alternatively, the bonds between the first set of molecules and the second set
of
molecules are broken by applying heat, by contacting the bonds with a solution
having a
high ionic strength, or by contacting the bonds with a solution having a high
ionic
strength and applying heat.
Typically, the nucleic acid sequence of the first and second sets of molecules
are
DNA, RNA, modified nucleic acid sequences or any combinations thereof.
Page 23
CA 02523896 2005-10-19
In an alternative embodiment, a component of each of the fizst set of
molecules is
a peptide nucleic acid {PNA) sequence and the recognition component of the
second set
of molecules is a PNA sequence. Alternatively, a component of each of the
molecules
from the first set of molecules is a peptide nucleic acid (PNA) sequence and
the
S recognition component of the second set of molecules is a nucleic acid
sequence, or vice
versa. PNA molecules hybridize to other PNA molecules and to nucleic acid
sequences
in a manner similar to that of nucleic acid hybridization to othex nucleic
acid. Thus, at
Ieast one or more molecule from the second set of molecules must have at least
three
consecutive bases, preferably six consecutive bases, that are complementary to
three
consecutive bases, preferably six consecutive bases, of a molecule from the
first set of
molecules.
When a set of molecules bind to the surface of a substrate, the molecules will
fold
or stack against one another such that a portion of the molecule will be
exposed on the
surface of the substrate. The exposed functionality may be hydrophobic,
hydroplulic, or
1 S an ainplupathic functionality. In addition, the exposed fiuletionality
rnay be a
functionality that selectively binds various biological or other chemical
species such as
proteins, antibodies, antigens, sugars and other carbohydrates, and the like.
Other
examples of exposed functionalities include -4H, -CONH-, -CONHCO-, -NH2, -NH-,
-COOH, -COOR, -CSNH-, -NOz-, -SO2, -SH, -RCOR-, -RCSR-, -RSR, -ROR-, -P04 3,
-OS03 z, -S03w, -COO, -SOO, -RSOR-, -CONRz, -(OCHzCHz)nOH (where n=1-20,
preferably 1-8), -CH3, -P03H , -2-imidazole, -N(CH3)z, -IV(R)z, -P03H2, -CN, -
(CFz~,CF3
(where n=1-Z0, preferably 1-8), and an olefn, wherein, R is hydrogen, a
hydrocarbon, a
halogenated hydrocarbon, a protein, an enzyme, a carbohydrates, a lectin, a
horncione, a
receptor, an antigen, an antibody, or a hapten.
The exposed functionality may include a protecting group which may be removed
to effect further modification of the complement image or the reproduction of
the master.
For example, a photoremovable protecting group may be used, A wide variety of
positive light-reactive groups are known in the art, for example,
nitroaromatic
compounds such as o-nztrobenzyl derivatives or benzylsulfonyl. Photoremovable
Page 24
CA 02523896 2005-10-19
protective groups are described in, for example, U.S. Pat. No. 5,143,854, the
entire
teachings of which are incorporated herein by reference, as well as an article
by
Patchornik, .IACS, 92:6333 (1970) and Amit et al, JOC, 39:192, (1974), both of
which
are incorporated herein by reference.
In one embodiment, the complement image can be further modified by binding
the exposed functional group of at least one of the second set of molecules to
a metal or a
metal ion. For example, when the exposed functional group is -SH, the metal or
metal
ion can be Au°, Ag°, or Age. AItematively, when the exposed
functional group is
-COOH, the metal or metal ion can be Ag° or Ag+.
1.0 The second set of molecules may have a first spacer, a second spacer, or a
first
and the second spacer. The spacers may be, independently, selected from the
group
consisting of an alkylene, a heteroalkyIene, a heterocycloalkylene, an
alkenylene, an
alkynylenc, m arylene, a heteroarylene, arylalkylene, and a
heteroarylalkylene. The
alkylene, heteroalkylene, heterocycloalkylene, alkenylene, alkynylene,
arylene,
heteroarylene, arylalkylene, and heteroarylalkylene spacers may be substituted
or
unsubstituted. In one embodiment, either the first or the second spacers, or
both the first
and the second spacers are substituted with one or more halogen and/or
hydroxy.
The master may be prepared by any method known to those skilled in the art
(see
Xia, et al., Chem. Rev. (1999), 99;1823-.1848, the entire teachings of which
are
incorporated by reference). Preferably, the method of forming the master is a
nanopatterning method. Isi one embodiment, the master is prepared by forming a
pattern
of one or more metal, metal oxide, or combinations thereof on a surface of a
substrate
using electron beam Iitha~-aphy. The surface of the substrate is then
contacted with a
first set of molecules. In this embodiment, each of the first set of molecules
has a
reactive functional group that foams a bond between the metal or metal oxide
and the
molecules of the first set of molecules, so that the first set of molecules
binds to the
substrate fom~ing a master having a first set of molecules bound to the
substrate to form a
pattern. For example, when the reactive functional group of at least one
molecule from
the first set of molecules is a thiol group or a protected thiol group, at
least a portion of
Page 25
CA 02523896 2005-10-19
the patterned formed may be a metal selected from the group consisting of
gold, silver,
copper, cadmium, zinc, palladium, platinum, mercury, Lead, iron, chromium,
manganese,
tungsten, and any alloys thereof. Iz~. another example, when the reactive
functional group
of at least one molecule from the first set of molecules a silane or a
ehlorosilane, at least a
5 portion of the patterned formed is a metal selected from the group
consisting of doped
and undoped silicon. In another example, when the reactive functional group of
at least
one molecule from the first set of molecules is a carboxylic acid, at Least a
portion of the
patterned formed is an oxide selected from the group consisting of silica,
alumina, quaiatz,
and glass. In another example, when the reactive functional group of at least
one
molecule from the first set of molecules is a nitrite or an isonifirile, at
Least a portion of the
patterned fanned is a metal selected from the group consisting of platinum,
palladium
a~~d alloys thereof. In another example, when the reactive functional group of
at least one
molecule from the first set of molecules is a hydroxamic acid, at least a
portion of the
patterned formed is copper.
15 Alternatively, the master can be prepared using dip pen nanolithography.
Methods of preparing molecularly patterned substrates using dip pen
naiiolithography are
described in Schwartz, Langnauir (2002), 18:4041-4046 and in Piner, et ah,
Science
(1999), 283:661-663, the entire teachings of both references are incorporated
herein by
reference.
20 Alternatively, the master can be prepared using replacement lithography,
nanoshading or nanografting. These methods are described in Sun, et al., JA CS
(2002),
124(1 I):2414-2415; Amxo, et al., Langrnuir (2000),16:3006-3009; Liu, et al.,
Nano
Letters (2002), 2(8):863-867; and Liu, et al., Acc. Cheat. Res. (2000),
33:45'7-466; the
entire teachings of these references are incorporated herein by reference.
25 Another ernbodisnent is a lithographic method in which at least one portion
of the
second substrate surface is free of the second set of molecules. In this
embodiment, the
exposed surface of the second substrate is contacted with a reactant selected
to be
chemically inert to the second set ofrnolecules and to degrade at least the
surface Layer of
the second substrate, thereby degrading the portion of the surface of the
second substrate
Page 26
CA 02523896 2005-10-19
that is free of the second set of molecules. Typically, the reactant is a
reactive ion
etching compound. The second set of rnolecuIes is then removed to uncover a
portion of
the surface of the second substrate.
In another embodiment, at least one portion of the second substrate surface is
free
of the second set of molecules, and a material is deposited on the portion of
the second
substrate surface that is free of the second set of molecules. Examples of
deposited
material include semiconductors, dielectrics, metals, metal oxides, metal
nitrides, metal
carbides, and combinations thereof The second set of molecules is then removed
to
uncover a portion of the surface of the second substrate.
10 Tn one aspect of the invention, the method of forming a complement image of
a
master involves assembling a second set of molecules via attractive forces on
the first set
of molecules. Examples of attractive forces include attraction of a molecule
having a net
positive charge to a molecule having a net negative charge, dipole-dipole
attraction, and
magnetic attraction. Tn one preferred embodiment, the attractive force is a
magnetic
15 force. In one example, when the attractive force is a magnetic force, one
or wore
molecules fxom the first set of molecules and from the second set of molecules
include an
iron or iron oxide component. >i1 this embodiment, the attractive forces
between the first
set of molecules a~~d the second set of molecules can be broken by applying a
magnetic
field.
20 In another aspect of the invention, the method involves forming a
reproduction of
a master, or a portion thereof. The master used in: this embodiment of the
method of the
invention comprises a first set of molecules bound to a first substrate to
form a pattern. A
second set of molecules is assembled on the flr5t set of molecules via bond
fonuation.
The second set of molecules comprises a reactive fitnctional group and a
recognition
25 component that hinds to the first set of molecules. The reactive functional
group of the
second set of molecules is then contacted with a surface of a second
substrate. The
reactive fractional group reacts with the surface of the second substrate to
form a bond
befween the second set of molecules and the second substrate. The bonds
behveen the
first set of molecules and the second set of molecules are then broken, and
the second set
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CA 02523896 2005-10-19
of molecules bound to the second substrate forms a complement image of the
master, A
third set of molecules is then assembled via bond formation on the second set
of
molecules of the complement image. Each molecule in the third set of molecules
comprises a reactive functional group, and a recognition component that binds
to the
S second set of molecules. The reactive functional group of the third set of
molecules is
then contacted with a surface of a thud substrate. The surface of the third
substrate reacts
with the reactive functional group of the third set of molecules to form a
bend between
the third set of molecules and the third substrate. The bonds between the
second set of
molecules and the third set of molecules are then broken, and the third set of
molecules
10 bound to the third subshate form a reproduction of the pattern, or portion
thereof, of the
master. Once the cocnplernent image has been separated form the reproduction,
the
complement image cart be reused one or more times to form additional
reproductions. In
one embodiment, a lateral dimension of at least vne feature of the
reproduction is less
than 200 nm.
15 The method of forming a reproduction is the same as that used to form a
complement image except that tire complement image of the master is used as a
template
(or "master") to transfer the pattern to the tliird substt~tes. Thus, the
embodiments and
examples disclosed above for the second set of molecules and the second
substrate apply
as well to the third set of molecules and the third substrate, respectively.
In addition,
20 examples of conditions for assembling the second set of molecules on the
first set of
molecules and for breaking the bonds beriveen the first and the second set of
molecules
can apply equally as well to conditions for assembling the third set of
molecules on the
second set of molecules and for breaking the bonds between the third and the
second set
ofmolecules.
2S Tn another aspect, the invention relates to a composition comprising a
master
having a pattern of. a first set of molecules bound to a first substrate; and
a complement
image comprising a pattern of a second set of molecules bound to a second
substrate via a
reactive functional group on each molecule of the second set of molecules. In
this aspect
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CA 02523896 2005-10-19
of the invention, each molecule in the second set of molecules has a
recognition
component that binds to at least a portion of a molecule from the first set of
molecule.
Tn another aspect, the invention relates to a composition, comprising a first
pattern
of a fzrst set of molecules bound to a fixst substrate; and a second pattern
of a second set
of molecules bound to a second substrate via a reactive functional group on
each
molecule of the second set of molecules, In this aspect of the invention, each
molecule of
the second set of molecules comprises a recognition component that binds to at
least one
molecule in the first set ofmolecules, and the second pattern is a complement
image of
the first pattern. Alternatively, the invention relates to a composition,
comprising a first
pattern of a f rst set of molecules bound to a first substrate; and a third
pattern of a third
set of molecules bound to a third substrate via a reactive functional group on
each
molecule of the third set of molecules. In this aspect of the invention, each
molecule of
the third set of molecules comprises a recognition component that binds to at
least one
molecule in the second set of molecules, and the third pattern is a
reproduction of the first
pattern, or a portion thereof.
Iu another aspect, the invention relates to a composition, comprising a first
pattern
of a first set of molecules bound to a first substrate; and a second
substrate, wherein the
second substrate comprises a degraded portion and an undegraded portion, and
the
undegraded portion is a complement image of the first pattern.
20 In another aspect, the invention relates to a composition, comprising a
first pattern
of a fast set of molecules bound to a first substrate; and a second substrate
having a
patterned layer of a material deposited thereon, whereat the patterned Iayer
of deposited
material on the second substrate is a complement image of the first pattern.
In another aspect, the invention relates to a kit for printing a molecular
pattern on
25 a substrate. The kit comprise a master comprising a pattern of a first set
of molecules
bound to a substrate; and a second set of molecules. The second set of
molecules
comprises a reactive functional group; and a recognition component that binds
to the first
set of molecules.
Page 29
CA 02523896 2005-10-19
In another aspect, the invention relates to a molecular printer for generating
a
complement image of a master, wherein the master has a first set of molecules
bound to a
first substrate. The molecular printer comprises comprising a device for
delivering a
solution of a second set of molecules to a surface of the master, and a device
for
contacting the second set of molecules with a second substrate. In this
embodiment, the
second set of molecules comprises a reactive functional group; and a
recognition
component that binds to the first set of molecules.
Generally, the apparatus comprises one or more reservoirs that contain the
second
set of molecules, one or more vessels or components far holding a master in
position for
delivery of the solution containing the second set of molecules. In addition,
the apparatus
may include a computer controlled means for transferring in a predetermined
mariner the
solution of the second set of molecules fi~om the reservoirs to the surface a
master. A
clamp that sectues the master to the second substrate may also be included in
the
apparatus of the invention. The temperature of the solution of the second set
of
1 ~ molecules and the vessel containing the master may also be controlled. The
apparatus
may also include areservoir containing a solution for breaking the bonds
between the
first and the second molecules, such as a solution having a high ionic
strength or a
solution containuig an enzyme that will break the bands, at~d a means fox
delivering the
solution. In addition, after the second subshate has been bound to the second
set of
molecules, a heating element may be used to heat a solution in contact with
the bound
first and second sets of molecules to break the bonds. The computer controlled
means for
transferring solutions and controlling temperature can be implemented by a
variety of
general purpose laboratory robots, such as that disclosed by Harrison et al,
Bioteclzniques,14: 88-97 (1993}; Fujita et al, Iiiotechraiques, 9: 584-591
(1990); Wada et
al, Rev. Sci. Irzstruan., S4: 1569=1572 (1983), the entire teachings of these
references are
incorporated herein by reference. Such laboratory robots are also available
commercially,
e.g. Applied Biosystems model 800 Catalyst {Faster City, Calif.}. In one
embodiment,
the apparatus also includes a device for separating the second substrate from
the master
Page 30
CA 02523896 2005-10-19
after the bonds between the first set of molecules and the second set of
molecules have
been broken.
These and other aspects of the present invention will be further appreciated
upon
consideration of the following Examples, which are intended to illustrate
certain
S particular embodiments of the invention but are not intended to limit its
scope, as defined
by the Claims.
EXAMPLES
Example 1: Preparation of a Complement Imo a of a DNA Monolayer
A. Preparation of DNA solutions
All glassware was cleaned with a solution of 75% I-IaS04 and 25%H202 before
use, All water used was ultrapure water (18MSZIcm).
The primary DNA, 5'-!5-ThiolMC6-DIACG CAA CTT CGG GCT CTT - 3', .
were purchased from Tntegrated DNA Technologies, Inc. (ZDT), Coraville, IA.
All DNA
strands were used as received from the manufaturer. The primary DNA was
dissolved in
water at the concentration of lp,g/mL and divided into smaller aliquots of 50
p.L, and
stored at -20°C. When a portion of this solution was used, an aliquot
was reduced by
ZO pIacing it in a 40 mM buffer solution (0.17 M sodium phosphate, pH 8 )
having
dithiothreitol {DTT) for 16 hr. The oligonucleotides were separated from the
by-products
of the DTT reaction using size exclusion chromatography (NAP 10 column from
Pharmacia Biotech) following the manufactures instructions. 10 mM sodium
phosphate
buffer (pH b.8) was used to equilibrate the column and to elute the
oligoriucleotides. The
concentration of the resulting DNA solution was calculated from die absorbance
of the
solution at 2G0 nm. Iu the case of primary DNA (i.e., the DNA used to form the
master),
1M potassium phosphate buffer solution (pH 3.8) was added to the DNA solution
to
increase the ionic strength of the solution. 'fhe final concentration of DNA
was 4-5 p,l4~I.
Page :l I
CA 02523896 2005-10-19
u~ the case of secondary DNA solution (i.e., DNA used to form the complement
image), 1M NaC1 in TE buffer (IOmM Tris buffer pH 7.2 and ImM EDTA) was added
to
increase the ionic strength of the solution. The secondary DNA used was
purchased from
Integrated DNA Technologies, lnc. (IDT), Coraville, IA and had the following
structure
S 5'-lSThiolMC6-DIAAG AGC CCG AAG TTG CGT - 3'.
B. Preparation of a Master having a DNA Monolayer
Clem and atomically flat gold on mica was used as a substrate, This substrate
was placed in the primary DNA solution prepared above for 5 days to allow the
DNA to
I O bind to the surface of the substrate. The substrate was rinsed with 1M
potassium
phosphate buffer 2 tunes and with water 5 times. The substrate was exposed to
1 mM
spacer thiol, 6-rnercapto-'1-hexanol, aqueous solution for 2 hr to minimize
nonspecific
adsorption of single-stranded DNA, then rinsed with water 5 times.
15 C. Preparation of Complement Jma~e
The master prepared in step B was dipped into the secondary DNA solution for 2
hours to allow the complementary DNA to hybridize to the DNA bound to the
master.
The substrate was rinsed with 1M NaCI in TE buffer 2 times aid with water 5
times.
A second clean gold on mica substrate was placed in contact with the master so
2Q that the two gold surfaces were facing each other and had a small amount of
water in
between them. A small mechanical force was applied to push the two substrates
together.
As water between two substrates was evaporating, the spacing betaveen the
surfaces
decreased due to increasing capillary attraction forces. Consequently, thiol
groups of
secondary DNA approached the second substrate and bound to it. After about 5
hr, the
25 substrates were dipped into 1 M NaCI in TE buffer solution (70°C)
for 20 min. The
substrates (i.e., the master and the complement image) spontaneously separated
and were
rinsed with IM NaCI in TE buffer 2 times and with water S times, then air-
dried. Both
the master (see Figs. 3A and 3B) and the complement image (see Figs. 3C and
3D) were
imaged using AFM tapping mode.
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CA 02523896 2005-10-19
D. Results
The coverage of the first substrate surface with DNA was complete. The
thorough coverage made AFM imaging difficult due to strong interaction between
monolayer and a tip. The layer transferred to the second substrate also had
complete
coverage.
Example 2: Pattern Transfer of Gold Grid
An AFM calibration gold grid was dipped in 4 ~I solufion of the primary DNA
molecules described in Example 1 for 5 days to generate a patterned master.
The master
was exposed to I nlltrl 6-mercapto-1-hexanol aqueous solution for 2 hr to
minimize
nonspecific adsorption of single-stranded DNA, then rinsed with water 5 times
and air-
dried. The master was then exposed to a 6 ~M solution of the secondary DNA
described
in Example 1 for 2 hours so that hybridization occurred. A second substrate of
gold on
mica was placed on the master so that the two gold surfaces were facing each
other and
had a small amount of water in between them. A small mechanical force was
applied to
force the two substrates together. After about S hr, the substrates were
dipped into 1M
NaCI in TE buffer solution (70°C} for 20 min. The two substrates (i.e.,
master and the
complement image} spontaneously separated and were rinsed with 1M NaCI in TE
buffer
20 2 times and with water S times, then air-dried. Both the master and the
complement
image were imaged using AFM tapping mode (see Figs. 4A and 4B, respectively).
Example 3: Fabrication of a DIVA Chin
A master is prepared using Dip Pen Nanolithography, as described in Demer, et
25 al., Anoew. CI1BM2. ant. Ed. (200I), 40:30713073, the entire teachings of
which are
incorporated herein by reference. To prepare the master, a surface of a gold
on mica
substrate is contacted with a I mM solution of 1-octadecanethiol (ODT) in
ethanol for 5
min. to cover the exposed gold surface with ODT molecules. The substrate is
then
unmersed in a 1 mM solution of 1,16-mercaptohexadecanoic acid (Ivl~iA} and the
tip of
page 33
CA 02523896 2005-10-19
an atomic force microscope is used to displace ODT molecules bound to the
surface by
contacting the surface with a force of about 0.5 nN making a 100 nm dot . The
MHA in
solution binds to the exposed gold surface of the dot. The carboxylic acid
groups of the
MHA are activated with a 10 mglmL solution of 1-ethyl-3-(3-
5 dimethylaminoprappyl)carbodiimide hydrochloride (EDAC) in 0. s M
morpholine/ethanesulfonic acid at pH 4.5, and then rinsed With a solution of
O.1M
sodium boratelboric acid buffer, pH 9.5. A ZS pM solution of a DNA modified
with a I-
n-hexyl amine group in the borate buffer is placed on the surface of the
substrate. The
anune groups of the DNA bind to the activated MHA molecules farming a DNA dot
10 having a I00 nm diameter. The procedure of forming an MT~A dot and binding
a DNA
molecule to it is repeated many more times wish different amuse modified DNA
molecules to form a master having a DNA array with featuxe of about s 00 nm.
The master is used to print a complement image array of DNA sequences on a
second substrate in which each DNA sequence is complementaxy to one of the DNA
15 molecules on the master and is located at a position on the second
substrate that is a
mirror image of its complementary sequence on the master. The complement image
array is prepared by modifying a set of DNA molecules that includes all of the
DNA
molecules that are complementary to the DNA molecules on the master with a
hexyl thiol
linker. The thiol modified DNA molecules are placed in a phosphate buffer
having a pH
20 of 6.8 and 1 M NaCI. The master is immersed in the solution containing the
ihiol
modified DNA molecules for 2 hrs, then master is removed from the solution and
rinsed
with 1 M NaCI in TE buffer once and with water five times.
A second clean gold on mica substrate is placed in contact with the master so
that
the two gold surfaces are facing each other and have a small amount of water
in between
25 them. A Sinall mechanical force is applied to push the two substrates
together. As water
between the two substrates evaporates, the spacing between the surfaces
decreases due to
increasing capillary attraction forces. Consequently, thiol groups of the
thiol modified
DNA molecules approach the second substrate and bind to it. After about 5 hrs,
the
substrates are dipped into 1 M NaCI in TE buffer solution (70°C) for 20
min. The
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CA 02523896 2005-10-19
substrates spontaneously separate and are rinsed with 1 M NaCI in TE buffer 2
times and
with water five times, then allowed to air dry. The master can be used to
prepare one or
more additional complement images.
Example 4; Preparation of a Complement Image of a DNA Arr
A DNA chip is purchased and used as a primary master. The DNA chip has a 12
x 12 squarE array in which each square is 300 run x 300 nrn and has a
different DNA
sequence attached to a substrate for a total of 144 different DNA sequences.
The 300 nm
x 300 nm squares are spaced 100 nm apart along the x- and y-axis of the
surface of the
substrate.
The master is used to print a 12 x 12 complement image array of DNA sequences
on a second substrate in which each DNA sequence is complementary to one of
the DNA
molecules on the master and is located at a position on the second substrate
that is a
mirror unage of its complementary sequence on the master. A set of DNA
molecules that
includes all of the DNA molecules that are complementary to the DNA molecules
on the
master (i.e., 144. different complementary DNA sequences) are modified with a
hexyl
thiol linker. The thiol modified DNA molecules are placed in a phosphate
buffer having
a pH of 6.8 and I M NaCI. The master is immersed in the solution containing
the thiol
modified DNA molecules for 2 hrs, then master is removed from the solution and
rinsed
with 1 M NaCI in TE buffer once and with water Five times.
A clean gold on mica substrate is placed in contact with the master so that
the
gold surface of the new substrate is facing the 12 x 12 array of DNA
molecules. A small
amount of water is in between the two surfaces. A small mechanical force is
applied to
push the two substrates together. As water between the two substrates
evaporates, the
spacing between the surfaces decreases due to increasing capillary attraction
forces.
Consequently, thiol groups of the thiol modified DNA molecules approach the
second
substrate and bind to it. After about 5 hrs, the substrates are dipped into 1
M NaCI in TE
buffer solution (70°C) far 20 nun. The substrates spontaneously
separate and are rinsed
with lIvl NaCI in TE buffer 2 times and with water five times, then allowed to
air dry.
Page 3J
CA 02523896 2005-10-19
The complement image has a 12 x 12 array of DNA molecules that are
complernelltary to
the DNA molecules on the master. The master can be used to prepare one or more
additional complement image arrays following the same procedure.
In addition, the primary master can be replicated one or more times by
following
the procedure, as described above, except that the complement image is used in
place of
the master and a third set of 144 DNA molecules having the same sequences as
the DNA
molecules on the primary master and modified with a beryl thiol linker is
assembled on
the complement image. A third substrate of gold on mica is then bmught in
contact with
the complement image as described above for the primary master and the second
substrate. The third set of DNA bound to the third substrate aid separated
from the
complement image is a replica of the primary master.
Other embodiments of the invention will be apparent to those skilled in the
art
from a consideration of the specification or practice of the invention
disclosed herein. It
is intended that the specification and examples be considered as exemplary
only, with the
true scope and spirit of the invention being indicated by the following
claims.
Page 36
