U.S. patent application number 11/287808 was filed with the patent office on 2006-06-29 for manufacture and use of non-standard size microarray slides.
Invention is credited to John S. Hinchcliffe, Mel N. Kronick, Matthew J. Marton, Dianne Rees, Bernett E. Saunders, Carol T. Schembri.
Application Number | 20060141507 11/287808 |
Document ID | / |
Family ID | 36612116 |
Filed Date | 2006-06-29 |
United States Patent
Application |
20060141507 |
Kind Code |
A1 |
Kronick; Mel N. ; et
al. |
June 29, 2006 |
Manufacture and use of non-standard size microarray slides
Abstract
Methods and devices are disclosed for microarray analysis. In
one embodiment a method is disclosed for processing a non-standard
size slide having an array of chemical compounds attached to a
surface of the slide. A sample is exposed to the surface of the
non-standard size slide wherein components in the sample bind to
the chemical compounds on the surface of the slide. The sample and
the slide are incubated under conditions for carrying out the
binding reactions, and the surface of the non-standard size slide
is examined for the results of the binding reactions. Prior to the
exposing step or the incubating step or the examining step, the
non-standard size slide is placed into a slide holder comprising a
slide-holding section a slide-holding section adapted to dispose
the non-standard size slide to a processing instrument in a manner
similar to that for a standard size slide. The non-standard size
slide may also include an identifier such as a bar code.
Inventors: |
Kronick; Mel N.; (Palo Alto,
CA) ; Saunders; Bernett E.; (Portland, OR) ;
Marton; Matthew J.; (Seattle, WA) ; Hinchcliffe; John
S.; (Seattle, WA) ; Rees; Dianne; (Sunnyvale,
CA) ; Schembri; Carol T.; (San Mateo, CA) |
Correspondence
Address: |
AGILENT TECHNOLOGIES, INC.;INTELLECTUAL PROPERTY ADMINISTRATION, LEGAL
DEPT.
P.O. BOX 7599
M/S DL429
LOVELAND
CO
80537-0599
US
|
Family ID: |
36612116 |
Appl. No.: |
11/287808 |
Filed: |
November 28, 2005 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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11030391 |
Jan 5, 2005 |
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11287808 |
Nov 28, 2005 |
|
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09671966 |
Sep 27, 2000 |
6864097 |
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11030391 |
Jan 5, 2005 |
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Current U.S.
Class: |
435/6.11 ;
435/287.2; 435/7.1; 702/19 |
Current CPC
Class: |
Y10T 436/25 20150115;
B01J 2219/00612 20130101; G02B 21/34 20130101; B01J 2219/00662
20130101; B01J 2219/00637 20130101; B01J 2219/0061 20130101; B01J
2219/00659 20130101; Y10T 436/2575 20150115; B01L 9/52 20130101;
B01J 2219/00547 20130101; B01J 2219/00626 20130101; B01J 2219/00531
20130101; B01L 2300/0819 20130101; B01L 2300/0822 20130101; B01L
3/508 20130101; B01L 2300/0636 20130101; B01J 2219/00608
20130101 |
Class at
Publication: |
435/006 ;
435/007.1; 702/019; 435/287.2 |
International
Class: |
C12Q 1/68 20060101
C12Q001/68; G01N 33/53 20060101 G01N033/53; G06F 19/00 20060101
G06F019/00; C12M 1/34 20060101 C12M001/34 |
Claims
1. A method for processing a non-standard size slide having an
array of chemical compounds attached to a surface thereof, said
method comprising: (a) exposing a sample to said surface of said
non-standard size slide wherein said chemical compounds, if
present, bind to components in said sample, (b) incubating said
sample under conditions for carrying out said binding reactions,
and (c) examining said surface of said non-standard size slide for
the results of said binding reactions, wherein prior to step (a) or
step (b) or step (c) said non-standard size slide is placed into a
slide holder comprising a slide-holding section adapted to dispose
said non-standard size slide to a processing instrument in a manner
similar to that for a standard size slide.
2. A method according to claim 1 wherein said non-standard size
slide has dimensions in the plane of the slide of about one inch by
about 1 to 2.5 inches.
3. A method according to claim 1 wherein said array of said
non-standard size slides has about 100 to about 25,000 features
comprising said chemical compounds.
4. A method according to claim 1 wherein said chemical compounds
are biopolymers.
5. A method according to claim 1 wherein said chemical compounds
are polynucleotides or polypeptides.
6. A method according to claim 1 wherein said slide holding section
is adjustable to receive a non-standard size slide and dispose said
non-standard size slide to a processing instrument in a manner
similar to that for a standard size slide.
7. A method for preparing a plurality of non-standard size slides,
each slide having an array of chemical compounds attached to a
surface thereof, said method comprising: (a) forming a plurality of
arrays on a surface of a sheet of slide material, and (b) dividing
said sheet into a plurality of non-standard size slides wherein
each slide has a single array of chemical compounds on a surface
thereof and each slide has at least one dimension that is less than
that of a standard microscope slide,
8. A method according to claim 7 wherein said dimension is
length.
9. A method according to claim 7 wherein said non-standard size
slides have dimensions in the plane of the slides of about one inch
by about 1 to 2.5 inches.
10. A method according to claim 7 wherein said sheet has dimensions
of about 6 inches by about 6 inches.
11. A method for processing a non-standard size slide prepared by
the method of claim 7, said method comprising: (a) exposing a
sample to said surface of said non-standard size slide wherein said
chemical compounds bind to components in said sample, (b)
incubating said sample under conditions for carrying out said
binding reactions, (c) placing said non-standard size slide into a
slide holder comprising a slide-holding section adapted to dispose
said non-standard size slide to a processing instrument in a manner
similar to that for a standard size slide, and (d) examining said
surface of said non-standard size slide for the results of said
binding reactions.
12. A method according to claim 11 wherein said array of said
non-standard size slides has about 100 to about 25,000 features
comprising said chemical compounds.
13. A method according to claim 11 wherein said chemical compounds
are biopolymers.
14. A method according to claim 11 wherein said chemical compounds
are polynucleotides or polypeptides.
15. A method comprising forwarding data representing a result
obtained from a method according to claim 14.
16. A method according to claim 15 wherein the data is transmitted
to a remote location.
17. A method comprising receiving data representing a result
obtained from a method according to claim 14.
18. A method according to claim 11 wherein said slide holding
section is adjustable to receive a non-standard size slide and
dispose said non-standard size slide to a processing instrument in
a manner similar to that for a standard size slide.
19. A device comprising (a) a non-standard size slide having at
least one dimension that is less than that of a standard microscope
slide, said non-standard size slide having a single array of
chemical compounds on a surface thereof, and (b) a holder for said
non-standard size slide wherein said holder comprises a
slide-holding section adapted to dispose said non-standard size
slide to a processing instrument in a manner similar to that for a
standard size slide.
20. A device according to claim 19 wherein said at least one
dimension is length.
21. A device according to claim 19 wherein said non-standard size
slide has dimensions in the plane of the slide of about one inch by
about 2 inches.
22. A device according to claim 19 wherein said array of said
non-standard size slide has about 100 to about 25,000 features
comprising said chemical compounds.
23. A device according to claim 19 wherein said chemical compounds
are biopolymers.
24. A device according to claim 19 wherein said chemical compounds
are polynucleotides or polypeptides.
25. A device according to claim 19 wherein said non-standard size
slide comprises an identifier.
26. A device according to claim 25 wherein said identifier is a bar
code on said surface.
27. A device according to claim 19 wherein said slide holding
section is adjustable to receive a non-standard size slide and
dispose said non-standard size slide to a processing instrument in
a manner similar to that for a standard size slide.
28. A device comprising (a) a non-standard size slide having at
least one dimension that is less than that of a standard microscope
slide, said non-standard size slide having a single array of
chemical compounds on a surface thereof, and (b) an identifier on
said non-standard size slide.
29. A device according to claim 28 wherein said at least one
dimension is length.
30. A device according to claim 28 wherein said non-standard size
slide has dimensions in the plane of the slide of about one inch by
about 2 inches.
31. A device according to claim 28 wherein said array of said
non-standard size slide has about 100 to about 25,000 features
comprising said chemical compounds.
32. A device according to claim 28 wherein said chemical compounds
are biopolymers.
33. A device according to claim 28 wherein said chemical compounds
are polynucleotides or polypeptides.
34. A device according to claim 28 wherein said identifier is a bar
code on said surface.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation-in-part of U.S. patent
application Ser. No. 11/030,391 filed Jan. 5, 2005, which is a
divisional of U.S. patent application Ser. No. 09/671,966 filed
Sep. 27, 2000, now U.S. Pat. No. 6,864,097, the disclosures of
which are incorporated herein by reference in their entirety.
BACKGROUND OF THE INVENTION
[0002] This invention relates to the manufacture of supports or
substrates having bound to the surfaces thereof a plurality of
chemical compounds, such as biopolymers. In particular, the
invention relates to the manufacture of microarray slides of
non-standard size and, more particularly, to the manufacture of
microarray slides having a size that is smaller in at least one
dimension than a standard microscopic slide.
[0003] In the field of diagnostics and therapeutics, it is often
useful to attach species to a surface. One important application is
in solid phase chemical synthesis wherein initial derivatization of
a substrate surface enables synthesis of polymers such as
oligonucleotides and peptides on the substrate itself. Support
bound oligomer arrays, particularly oligonucleotide arrays, may be
used in screening studies for determination of binding
affinity.
[0004] Determining the nucleotide sequences and expression levels
of nucleic acids (DNA and RNA) is critical to understanding the
function and control of genes and their relationship, for example,
to disease discovery and disease management. Analysis of genetic
information plays a crucial role in biological experimentation.
This has become especially true with regard to studies directed at
understanding the fundamental genetic and environmental factors
associated with disease and the effects of potential therapeutic
agents on the cell. Such a determination permits the early
detection of infectious organisms such as bacteria, viruses, etc.;
genetic diseases such as sickle cell anemia; and various cancers.
This paradigm shift has lead to an increasing need within the life
science industries for more sensitive, more accurate and
higher-throughput technologies for performing analysis on genetic
material obtained from a variety of biological sources.
[0005] Unique or misexpressed nucleotide sequences in a
polynucleotide can be detected by hybridization with a nucleotide
multimer, or oligonucleotide, probe. Hybridization is based on
complementary base pairing. When complementary single stranded
nucleic acids are incubated together, the complementary base
sequences bind to one another or pair to form double stranded
hybrid molecules. These techniques rely upon the inherent ability
of nucleic acids to form duplexes via hydrogen bonding according to
Watson-Crick base-pairing rules. The ability of single stranded
deoxyribonucleic acid (ssDNA) or ribonucleic acid (RNA) to form a
hydrogen bonded structure with a complementary nucleic acid
sequence has been employed as an analytical tool in molecular
biology research. An oligonucleotide probe employed in the
detection is selected with a nucleotide sequence complementary,
usually exactly complementary, to the nucleotide sequence in the
target nucleic acid. Following hybridization of the probe with the
target nucleic acid, any oligonucleotide probe/nucleic acid hybrids
that have formed are typically separated from unhybridized probe.
The amount of oligonucleotide probe in either of the two separated
media is then tested to provide a qualitative or quantitative
measurement of the amount of target nucleic acid originally
present.
[0006] Direct detection of labeled target nucleic acid hybridized
to surface-bound polynucleotide probes is particularly advantageous
if the surface contains a mosaic of different probes that are
individually localized to discrete, and often known, areas of the
surface. Such ordered arrays containing a large number of
oligonucleotide probes have been developed as tools for high
throughput analyses of genotype and gene expression.
Oligonucleotides synthesized on a solid support recognize uniquely
complementary nucleic acids by hybridization, and arrays can be
designed to define specific target sequences, analyze gene
expression patterns or identify specific allelic variations. The
arrays may be used for conducting cell study, diagnosing disease,
identifying gene expression, monitoring drug response,
determination of viral load, identifying genetic polymorphisms,
analyzing gene expression patterns or identifying specific allelic
variations, and the like.
[0007] In one approach, cell matter is lysed, to release its DNA as
fragments, which are then separated out by electrophoresis or other
means, and then tagged with a fluorescent or other label. The
resulting DNA mix is exposed to an array of oligonucleotide probes,
whereupon selective binding to matching probe sites takes place.
The array is then washed and interrogated to determine the extent
of hybridization reactions. In one approach the array is imaged so
as to reveal for analysis and interpretation the sites where
binding has occurred. Arrays of different chemical compounds or
moieties or probe species provide methods of highly parallel
detection, and hence improved speed and efficiency, in assays.
Assuming that the different sequence polynucleotides were correctly
deposited in accordance with the predetermined configuration, then
the observed binding is indicative of the presence and/or
concentration of one or more polynucleotide components of the
sample.
[0008] The arrays may be microarrays created on the surface of a
support by in situ synthesis of biopolymers such as
polynucleotides, polypeptides, polysaccharides, etc., and
combinations thereof, or by deposition of molecules such as
oligonucleotides, cDNA and so forth. In general, arrays are
synthesized on a surface of a support or substrate by one of any
number of synthetic techniques that are known in the art. In one
approach, for example, the support may be one on which a single
array of chemical compounds is synthesized. Alternatively, multiple
arrays of chemical compounds may be synthesized on the support,
which is then diced, i.e., cut, into individual assay devices,
which are supports that each comprises a single array, or in some
instances multiple arrays, on a surface of the support.
[0009] The dimensions of a standard microscope slide are generally
about one inch (about 25 mm) in width, about three inches (about 75
mm) in length and about 0.040 inches (about 1 to 1.2 mm) in
thickness. Many apparatus for automating various procedures that
are employed in manipulations involving microarrays on slides are
designed for the standard microscope slide. Such manipulations
include contacting the surface of the slide with various reagents
and washing buffers, providing for specific chemical conditions
during which specific binding (e.g., hybridization) can occur or
during which species-selective washing can occur, examining the
surface for the results of contacting the surface with a sample,
and so forth. Furthermore, many of the above known apparatus are
designed assuming one array of chemical compounds per slide. A
standard microscope slide can carry up to about 20,000 or more
features comprising chemical compounds. In many circumstances much
fewer features are required for analyzing samples for various
analytes or components that might be present in the samples. For
example, many determinations require only about 1500 to about 2500
features. This has given rise to having multiple arrays on the
surface of a standard microscope slide. However, since many of the
known apparatus are designed assuming one array per slide, standard
microscope slides that have more than one array for conservation of
samples and reagents, minimization of sample handling steps, and
the like, usually include barriers for separating the arrays from
one another during the processing and examining stages. The
barriers may be physical such as, for example, ridges, gaskets, and
the like, or chemical such as, for example, coatings that are
hydrophobic, and the like. All of the above contribute to higher
costs of manufacture and use.
[0010] There is, therefore, a need for methods of preparing slides
with a single array of features where the slides are generally
smaller than the standard microscope slide. There is also a need
for methods and devices for using non-standard size slides with
known equipment that is designed for handling a standard microscope
slide in an automated manner. There is also a need for having one
identifier associated with one array on a slide.
SUMMARY OF THE INVENTION
[0011] One embodiment of present invention is a method for
processing a non-standard size slide having an array of chemical
compounds attached to a surface of the slide. A sample is exposed
to the surface of the non-standard size slide wherein components in
the sample bind to the chemical compounds on the surface of the
slide. The sample and the slide are incubated under conditions for
carrying out the binding reactions, and the surface of the
non-standard size slide is examined for the results of the binding
reactions. Prior to the exposing step or the incubating step or the
examining step, the non-standard size slide is placed into a slide
holder comprising a slide-holding section adapted to dispose the
non-standard size slide to a processing instrument in a manner
similar to that for a standard size slide.
[0012] Another embodiment of the invention is a method for
preparing and processing a plurality of non-standard size slides
wherein each slide has an array of chemical compounds attached to a
surface of the slide. A plurality of arrays is formed on a surface
of a sheet of slide material, which is divided into a plurality of
non-standard size slides. Each of the slides has a single array of
chemical compounds on a surface thereof and each slide has at least
one dimension that is less than that of a standard microscope
slide. A sample is exposed to the surface of the non-standard size
slide and the chemical compounds on the surface bind to components
in the sample, which is incubated with the surface under conditions
for carrying out the binding reactions. The non-standard size slide
is placed into a slide holder comprising a slide-holding section
adapted to dispose the non-standard size slide to a processing
instrument in a manner similar to that for a standard size slide.
The surface of the slide is examined for the results of the binding
reactions.
[0013] Another embodiment of the invention is a device comprising a
non-standard size slide having at least one dimension that is less
than that of a standard microscope slide and a holder for the
non-standard size slide wherein the holder comprises a
slide-holding section adapted to dispose the non-standard size
slide to a processing instrument in a manner similar to that for a
standard size slide. The non-standard size slide has a single array
of chemical compounds on a surface of the slide.
[0014] Another embodiment of the present invention is a device
comprising a non-standard size slide having at least one dimension
that is less than that of a standard microscope slide and an
identifier on the non-standard size slide. The non-standard size
slide has a single array of chemical compounds on a surface of the
slide.
BRIEF DESCRIPTION OF THE DRAWINGS
[0015] FIG. 1 is a depiction of a sheet of slide material, which
comprises lines to delineate individual non-standard size
slides.
[0016] FIG. 2 is a depiction of a single non-standard size slide
shown in FIG. 1.
[0017] FIG. 3 is a depiction of a single non-standard size slide
partially inserted into a slide holder in accordance with
embodiments of the present invention that has as slide holding
section adapted to dispose the non-standard size slide to a
processing instrument in a manner similar to that for a standard
size slide.
[0018] FIG. 4 is a depiction of a single non-standard size slide
partially inserted into a slide holder in accordance with
embodiments of the present invention that has as slide holding
section adapted to dispose the non-standard size slide to a
processing instrument in a manner similar to that for a standard
size slide where the slide holding section is adjustable.
DETAILED DESCRIPTION OF THE INVENTION
[0019] In one aspect, the present invention provides methods for
preparing substrates having an array of features bound to at least
one surface of the substrate. The features generally comprise
chemical compounds. The present methods provide array slides that
have a non-standard size as compared to the standard microscope
slide. The chemical compounds are arranged to form an array on each
of a plurality of predetermined sections of a larger sheet of
substrate that is divided to provide single slides, each comprising
a single array of chemical compounds. In some embodiments the
single slides each have an identifier, which is specific for the
one array on its surface. In some embodiments the non-standard size
slide is used in conjunction with a holder, which has a slide
holding section adapted to dispose the non-standard size slide to a
processing instrument in a manner similar to that for a standard
size slide. This permits the present slides to be processed by
known equipment designed for processing of slides having dimensions
of a standard microscope slide.
[0020] Preferred materials for the sheet of material are those that
provide physical support for the chemical compounds that are
deposited on the surface or synthesized on the surface in situ from
subunits. The materials should be of such a composition that they
endure the conditions of a deposition process and/or an in situ
synthesis and of any subsequent treatment or handling or processing
that may be encountered in the use of the particular array.
[0021] Typically, the support material is transparent. By
"transparent" is meant that the support material permits signal
from features on the surface of the support to pass therethrough
without substantial attenuation and also permits any interrogating
radiation to pass therethrough without substantial attenuation. By
"without substantial attenuation" may include, for example, without
a loss of more than 40% or more preferably without a loss of more
than 30%, 20% or 10%, of signal. The interrogating radiation and
signal may for example be visible, ultraviolet or infrared light.
In certain embodiments, such as for example where production of
binding pair arrays for use in research and related applications is
desired, the materials from which the substrate may be fabricated
should ideally exhibit a low level of non-specific binding during
hybridization events. However, it should be noted that the nature
of the transparency of the substrate is somewhat dependent on the
nature of the scanner employed to read the substrate surface. Some
scanners work with opaque or reflective substrates.
[0022] The materials may be naturally occurring or synthetic or
modified naturally occurring. Suitable rigid substrates may include
glass, which term is used to include silica, and include, for
example, glass such as glass available as Bioglass, and suitable
plastics. Should a front array location be used, additional rigid,
non-transparent materials may be considered, such as silicon,
mirrored surfaces, laminates, ceramics, opaque plastics, such as,
for example, polymers such as, e.g., poly(vinyl chloride),
polyacrylamide, polyacrylate, polyethylene, polypropylene,
poly(4-methylbutene), polystyrene, polymethacrylate, poly(ethylene
terephthalate), nylon, poly(vinyl butyrate), etc., either used by
themselves or in conjunction with other materials. The surface of
the support is usually the outer portion of a support.
[0023] The surface of the material onto which the chemical
compounds are deposited or formed may be smooth or substantially
planar, or have irregularities, such as depressions or elevations.
The surface may be modified with one or more different layers of
compounds that serve to modify the properties of the surface in a
desirable manner. Such modification layers, when present, will
generally range in thickness from a monomolecular thickness to
about 1 mm, usually from a monomolecular thickness to about 0.1 mm
and more usually from a monomolecular thickness to about 0.001 mm.
Modification layers of interest include: inorganic and organic
layers such as metals, metal oxides, polymers, small organic
molecules and the like. Polymeric layers of interest include layers
of: peptides, proteins, polynucleic acids or mimetics thereof (for
example, peptide nucleic acids and the like); polysaccharides,
phospholipids, polymethanes, polyesters, polycarbonates, polyureas,
polyamides, polyethylene amines, polyarylene sulfides,
polysiloxanes, polyimides, polyacetates, and the like, where the
polymers may be hetero- or homo-polymeric, and may or may not have
separate functional moieties attached thereto (for example,
conjugated). Various further modifications to the particular
embodiments described above are, of course, possible. Accordingly,
the present invention is not limited to the particular embodiments
described in detail above.
[0024] The material used for an array support or substrate may take
any of a variety of configurations ranging from simple to complex.
Usually, the material is substantially rectangular and relatively
planar such as, for example, a slide. In many embodiments, the
material is shaped generally as a rectangular solid. As mentioned
above, multiple arrays of chemical compounds are synthesized on a
sheet, which is then singulated, such as, e.g., cut by breaking
along score lines, into single array slides. The sheet of material
may be of any convenient size depending on the nature of the
equipment used, production lot size, productive efficiencies,
production throughput demands, and so forth. In some embodiments,
the sheet of material is usually about 5, about 6, about 7, about
8, about 9, about 10, about 11, about 12, about 13 inches in length
and about 5, about 6, about 7, about 8, about 9, about 10, about
11, about 12, about 13 inches in width so that the sheet may be
divided into multiple single array supports having the dimensions
indicated below. The thickness of the sheet may be less than 1 cm,
or even less than 5 mm, 2 mm, 1 mm, or in some embodiments even
less than 0.5 mm or 0.2 mm. The thickness of the support is about
0.01 mm to 5.0 mm, usually from about 0.1 mm to 2 mm and more
usually from about 0.2 to 1. In a specific embodiment by way of
illustration and not limitation, a wafer that is 6.25 inches by 6
inches by 1 mm is employed.
[0025] The individual or single support is the section of the sheet
that is produced by dividing the sheet, usually along predetermined
lines as discussed above. The individual support usually has a
single array of chemical compounds that have been synthesized or
deposited on a surface when the individual support was part of the
sheet. The dimensions of the individual support are determined by
the number of features in the array on the surface of the support,
the intended use of the support, e.g., in conducting assays
involving the chemical compounds on the surface of the support,
ease of manual and automated handling steps, and so forth.
[0026] In general, at least one dimension, for example, length, of
the individual support of the invention is smaller than the
corresponding dimension of a standard microscope slide. A standard
size microscope slide is usually about 3 inches in length and 1
inch in width. Accordingly, as used herein, the phrase
"non-standard size slide" refers to a slide having at least one
dimension such as length or width that is smaller than a
corresponding dimension of a standard microscope slide as defined
above. By the term "smaller than" is meant that the dimension is at
least about 90%, at least about 85%, at least about 80%, at least
about 75%, at least about 70%, at least about 65%, at least about
60%, at least about 55%, at least about 50%, at least about 45%, at
least about 40%, at least about 35%, at least about 30%, at least
about 25%, at least about 20%, at least about 15%, at least about
10%, at least about 5% smaller than the corresponding dimension of
the standard microscope slide. Furthermore, more than one dimension
of the non-standard size slide may be smaller than corresponding
dimensions of a standard microscope slide in which case each
smaller dimension of the non-standard size slide is independently
smaller. For example, by way of illustration and not limitation, a
non-standard size slide may be smaller in length by about 80% and
smaller in width by about 40%.
[0027] Typically, the individual support has a length of less than
about 0.2 inches, less than about 0.3 inches, less than about 0.4
inches, less than about 0.5 inches, less than about 0.6 inches,
less than about 0.7 inches, less than about 0.8 inches, less than
about 0.9 inches, less than about 1.0 inches, less than about 1.1
inches, less than about 1.2 inches, less than about 1.3 inches,
less than about 1.4 inches, less than about 1.5 inches, less than
about 1.6 inches, less than about 1.7 inches, less than about 1.8
inches, less than about 1.9 inches, less than about 2.0 inches,
less than about 2.1 inches, less than about 2.2 inches, less than
about 2.3 inches, less than about 2.4 inches, less than about 2.5
inches, less than about 2.6 inches, less than about 2.7 inches,
less than about 2.8 inches, about 2.9 inches. In some embodiments
the length of the individual support is in the range of about 0.5
to about 2.5 inches, about 0.8 to about 2.2 inches, about 1.0 to
about 2.0 inches.
[0028] The individual support has a width of less than about 0.2
inches, less than about 0.3 inches, less than about 0.4 inches,
less than about 0.5 inches, less than about 0.6 inches, less than
about 0.7 inches, less than about 0.8 inches, less than about 0.9
inches, less than about 1.0 inches, less than about 1.1 inches,
less than about 1.2 inches, less than about 1.3 inches, less than
about 1.4 inches, less than about 1.5 inches, less than about 1.6
inches, less than about 1.7 inches, less than about 1.8 inches,
less than about 1.9 inches, less than about 2.0 inches. In some
embodiments the individual support has a width in the range of
about 0.5 to about 1.5 inches, about 0.8 to about 1.2 inches, about
0.9 to about 1.1 inches.
[0029] Any of a variety of geometries of arrays on a support may be
used. As mentioned above, an individual support usually contains a
single array. Features of the array may be arranged in rectilinear
rows and columns. This is particularly attractive for single arrays
on a support. The configuration of the arrays and their features
may be selected according to manufacturing, handling, and use
considerations.
[0030] Regardless of the geometry of the array on the surface of an
individual support or on the surface of a sheet comprising a
multiple of individual supports, the arrays normally do not
comprise the entire surface of the sheet or of the support. For
sheets of material comprising a multiple of individual supports,
the sheet typically has a border along its longitudinal edges that
is about 0.5 to about 3 mm wide, usually, about 1 to about 2 mm
wide. In many embodiments, the border of the individual supports
obtained from the sheet has the same dimensions as the border for
the sheet.
[0031] In some embodiments one area of the individual support that
is a non-interfeature area or a portion of a border or a
combination thereof comprises an identifier such as, e.g., a bar
code. It is often desirable to have some type of identification on
the array substrate that allows matching a particular array to
layout information, since array layout information in some form is
used to meaningfully interpret the information obtained from
interrogating the array. Unique identifiers and their generation
have been previously described, such as in U.S. Pat. No. 5,812,793,
U.S. Pat. No. 5,404,523, and the references cited therein. Such
unique identifiers (often referred to as "Globally Unique
Identifiers" or "GUIDs", or "Universally Unique Identifiers" or
"UUIDs") can, for example, include a network card identification,
which is specific to that card, along with a time and local counter
number, and other components.
[0032] Use of such unique identifiers in association with array
layouts generated at the same or different locations would
virtually eliminate the possibility of the same identifier being
associated with different array layouts. However, such unique
identifiers typically require 128 bit data string. A string of such
length when written, for example, as a bar code, typically takes up
about 3 to 4 cm, which is not consistent with the size of the
individual supports disclosed herein. However, U.S. Pat. No.
6,180,351 (Cattell) issued Jan. 30, 2001, describes an approach
wherein a second identifier is employed of shorter length than a
corresponding unique identifier, and which is associated in some
manner with the unique identifier. The second identifier is placed
on the individual support in a location not occupied by features of
the array. The disclosure of U.S. Pat. No. 6,180,351 is
incorporated herein by reference in its entirety.
[0033] As mentioned above, in some embodiments the non-standard
size slide is used in conjunction with a holder, which has a slide
holding section that has dimensions comparable with a standard
microscope slide. This permits the present slides to be processed
by known equipment designed for processing of slides having
dimensions of a standard microscope slide. Such processing
instruments include, by way of illustration and not limitation,
array scanners or readers, hybridization chambers, loading devices
for applying samples to arrays, systems for carrying out
hybridization and washing steps, and so forth.
[0034] The non-standard size slide is mounted in a slide holder and
retained thereon in a mounted position without the array contacting
the holder. The slide holder is then inserted into a known
processing instrument that is designed for processing standard
microscope slides or for slides of other dimensions as the case may
be. The slide holder may be of any design as long as the slide
holder has a slide holding section that is adapted to dispose the
non-standard size slide to the processing instrument in a manner
similar to that for a standard size slide. The determining factor
is the dimensions of a device that is accommodated by a particular
known processing instrument. As mentioned above, such instruments
are usually designed for standard microscope slides. However, in
general, the present invention has the flexibility to employ a
slide holder that has a slide holding section with dimensions that
are accommodated by a particular processing instrument, which may
be other than that of a standard microscope slide.
[0035] The phrase "adapted to dispose" is used herein to mean that
the configuration of the slide holding section of the slide holder
comprises various members such as retaining members, adjusting
members, grooves, indentations, wall members, and so forth to
present the non-standard size slide to the processing instrument in
a manner similar to that for a standard size slide or a slide for
which the processing instrument is designed. The non-standard size
slide is disposed to the processing instrument in a manner similar
to that for a standard size slide where the processing instrument
can receive the slide holder with the non-standard size slide
without modification to the processing instrument and that an array
and/or an identifier or the like on the surface of the slide can be
processed by the instrument as if such array and/or identifier or
the like were on the surface of a standard size slide.
[0036] In some embodiments the slide holding section is adapted to
dispose an identifier on a non-standard size slide to the
processing instrument in the same or similar manner as an
identifier on a standard size slide. The processing instrument
comprises hardware and software to perform the function of reading
the identifier and sending information to the appropriate
processing components of the processing instrument to indicate
where an array is found on the surface of the non-standard size
slide and when to start and stop the processing of the array.
Typically, the processing instrument is adapted to process an array
on a slide or some other function by a combination of hardware and
software. This includes the structure of a particular component of
the processing instrument and may also include a microprocessor,
embedded real-time software and I/O interface electronics to
control a sequence of operations and so forth.
[0037] In some embodiments the slide holder has a slide holding
section that is adjustable. In these embodiments the slide holder
comprises various components that permit the slide holding section
to be adjusted to receive and retain a non-standard size slide of
any non-standard size smaller than a standard size slide. Of
course, the adjustable nature of the slide holding section should
permit a non-standard size slide to be disposed to the processing
instrument in a manner similar to that for a standard size slide.
An adjustable slide holding section may be realized using various
components such as sliding wall members, movable retaining members,
and so forth.
[0038] Various embodiments of a slide holder are discussed next by
way of illustration and not limitation. In one embodiment the slide
holder comprises a slide holding section that is generally planar
and has a receiving portion that receives a non-standard size
slide. The slide holder itself may be planar or of some other shape
as long as the slide holding section has a shape and dimensions
that are accommodated by the processing instrument in question. The
receiving portion may be in the form of a cut out portion in the
slide holding section where the dimensions of the cut out portion
are the same as or slightly larger than the dimensions of the
non-standard size slide. The cut out portion may be fully or
partially open. As discussed more fully below, when the latter, the
slide holding portion may include a backer member that limits
access to the cut out portion to a direction other than where the
backing member is situated in the device. In some embodiments the
slide holding section may comprise various movable members so that
a slide-receiving portion of the slide holding section may be
adjusted to accommodate a non-standard size slide of any size
smaller than a standard size slide.
[0039] The receiving portion of the slide holding section is
designed so that the non-standard size slide is placed in the
receiving portion by sliding, direct insertion, and the like. The
sides of the receiving portion usually comprise retaining means for
retaining the slide in the slide holding section. The retaining
means may comprise one or more retaining members. For example, the
sides of the receiving portion may comprise one or more grooves,
retaining buttons, channels, or the like for receiving and/or
seating the non-standard size slide. In one aspect and as mentioned
above, the slide holder comprises an adaptor portion for varying
the size of the cut out portion to adapt to the dimensions of the
non-standard slide (e.g., to provide a receiving portion that is
the same or slightly larger than the dimensions of the non-standard
size slide).
[0040] In the instance where receiving portion comprises a cut out
portion that is only a partial cut out, a part of the slide holding
section remains in the area of the cut out. For example, the cut
out portion may be a recess in the slide holding section designed
so that the non-standard size slide may be seated in the recess and
only one surface of the slide faces outwardly, i.e., away from the
inside of the recess. As above, the sides of the cut out portion
may comprise retaining means or the like as discussed above.
[0041] The slide holder, in one embodiment, may have a body with
side portions and a channel intermediate the side portions. The
channel extends in a direction between ends of the body. In this
case, the backer member may be a bottom surface of the channel. In
such a configuration, front and rear clamp member sets may have
their members disposed about the channel, and one of those sets may
have its members fixed to the body side portions while the other
set is movable to an open position away from the fixed set. For
example, the front clamp member set may be fixed to the body side
portions and the rear clamp member set may be movable. In this
case, the slide is retained in the mounted position by being urged
against the fixed clamp member set. It should be noted that the
terms "front" and "rear" are relative terms based upon the
orientation of the viewer and such terms should not be considered
limiting to the scope of the slide holder discussed herein.
[0042] The slide holder may further comprise a control member set
positioned on an accessible location on the holder, for example at
a position outside the channel. The member set is moved to move the
movable clamp set to the open position. In the case where the
movable clamp set is the rear clamp member set, the control member
may simply be moved rearward to move the rear clamp member set to
the open position.
[0043] In the holder configuration where the body has the channel
as described above, the slide may be mounted on the holder by
sliding the slide in an endways direction of the channel and into
the mounted position in which a leading end of the slide abuts the
closed end of the channel. Clamp member sets positioned about the
channel, may be held in the open position during such a mounting
procedure (for example, by the control member set rearward).
[0044] The slide holder may additionally have two spaced apart
guides extending from the body adjacent respective sides of the
channel. With this configuration the slide may be slid into the
mounted position along the guides. The guides may be dimensioned
such that, when the slide is in the mounted position, a trailing
end of the slide is positioned between the guides. During any
mounting of the slide portions of the slide, portions of the slide
front and rear surfaces may be gripped (such as with a user's
fingers) and the gripped portions used to then slide the slide into
the mounted position. The guides, in such case may be dimensioned
such that the gripped portions are positioned between the guides
when the slide is in the mounted position.
[0045] The slide holder may optionally additionally include means
for ready removal of the slide from the mounted position. In some
embodiments, the removing step includes gripping portions of the
slide front and rear surfaces, which are between the guides, and
using the gripped portions to slide the slide in an endways
direction opposite to that in which the slide was slid during the
slide mounting.
[0046] The present invention, then, provides a holder for a
non-standard sized slide, which holder has any of the features
already described above. In one embodiment, the holder may include
a backer member and the clamp sets as described above. The clamp
members may be positioned such that the holder can receive and
retain a slide having an area of no more than 200 cm.sup.2 (or no
more than 100 cm.sup.2 or even no more than 50 or 40 cm.sup.2). The
holder may also be dimensioned such that the moieties on the rear
side of the mounted slide are spaced from the backer member by
between 0.1 mm to 10 mm (or more preferably between 0.5 and 5 or 3
mm). As mentioned above, the holder itself may have various shapes,
for example, rectangular. In one aspect, the holder will have a
maximum area of a side that is no more than 300 cm.sup.2 (or
preferably no more than 200 cm.sup.2 or 100 cm.sup.2). When a
channel is present in the holder it may, for example, be no wider
than 20 cm (or no wider than 15 cm.sup.2, 10 cm.sup.2, or 5
cm.sup.2).
[0047] In some embodiments, the present invention provides a
transparent non-standard size slide having opposed front and rear
surfaces, the slide carrying moieties such as an array of
biopolymers on a rear surface and an identifier on a front surface.
The identifier may be, for example, a bar code, which is printed on
an opaque label attached to the front side of the slide. In some
embodiments, the array on the surface of the slide is read through
the front surface and the identification code is read from a front
side.
[0048] Alternatively, as mentioned above, in some embodiments, the
holder may be used with an array of the mounted slide located on a
forward facing surface (that is, away from the holder). While the
surface protection benefit is lost in this configuration, the
benefits of ease of handling and physical protection of the slide
are retained. One may wish to read the array on a forward facing
side of the slide to allow for various opaque slides, mirrored
slides or to avoid the issues of thickness variations in
transparent slides (that is, arrays on a backward facing surface of
a slide which are read through the slide from the front side, may
be on different focal planes of the scanner in the case where the
thickness of the transparent slide varies).
[0049] Different embodiments of methods and devices of the present
invention can provide any or more of a number of useful features.
For example, moieties on the slide (such as the exposed array) can
be protected from damage and the slide itself protected from
breakage. Background signals during array reading may be reduced by
the use of a backer member. Further, it may be relatively easy to
use devices of the present invention and extensive manipulations of
the slide may be avoided, while relatively precise positioning of
the slide (and hence the moieties) in a reader may be obtained for
assisting in the reading of the exposed array.
[0050] The surface of the slide facing outwardly may be the surface
comprising the array of features. Alternatively, depending on the
nature of the processing step, the surface of the slide facing
outwardly may be that opposite to the surface with the array, in
which instance the surface comprising the array of features faces
inwardly, i.e., toward the inside of the recess. The recess may be
designed so that there is a space between the wall of the recess
and the surface of the slide. In this way a chamber is formed when
the non-standard size slide is inserted into the recess. The
chamber may be used for carrying out a processing step on the array
of features on the surface of the slide. The slide holder may
comprise one or more inlets and outlets for introducing reagents
into the chamber in this embodiment.
[0051] The slide holder may be manufactured from one or more of a
number of materials. The requirements of the material include
providing physical integrity for the slide, allowing manipulation
of the slide into and out of a processing instrument, chemical
properties including inertness and stability, appearance, and so
forth. Such materials include, for example, materials discussed
above for the manufacture of the slide. In some embodiments the
material from which the slide holder is manufactured is an opaque
plastic, e.g., ABS black plastic, etc., and the like. Where the cut
out portion is only partially cut out, the remaining portion of the
slide holder that faces the slide may be a transparent material as
defined above to allow, for example, for interrogation or
examination of the surface of the slide that faces the remaining
portion. The holder may be made in sections such as, for example,
two, three or more molded sections.
[0052] Embodiments of the invention are next described by way of
illustration and not limitation with reference to the attached
drawings. FIG. 1 depicts sheet 10 of slide material that is
approximately 6 inches by 6 inches. Sheet 10 is divided by score
lines 12 into 18 non-standard size slides 14 of approximately,
usually, exactly, equal dimensions, each slide being about 1 inch
by about 2 inches. It should be noted that a substrate of
approximately the same size as above would be divided into regions
corresponding to 12 standard-size (i.e., 1 inch by 3 inch)
slides.
[0053] An individual slide 14 is depicted in FIG. 2 and has array
16 comprising array features 17 and identifier 18. FIG. 3 depicts
slide 14 partially inserted into slide holding section 20 of slide
holder 22. The dimensions of slide holder 22 permit slide holder 22
to be inserted and retained in a processing instrument designed for
processing standard microscope slides. Accordingly, the outer
dimensions of slide holder 22 are the same as or approximately the
same as the outer dimensions of a slide holder for a standard size
slide. The dimensions of slide holding section 20 are such as to
receive and retain non-standard size slide 14. In this way, slide
holder 22 may be received in a known processing apparatus in many
instances designed for processing standard microscope slides.
[0054] FIG. 4 depicts slide 14 partially inserted into slide
holding section 30 of slide holder 32. Slide holder 32 also
comprises adjustable member 34, which is designed to permit slide
holding section 30 to be adjusted to receive and retain slide 14.
The dimensions of slide holder 32 permit slide holder 32 to be
inserted and retained in a processing instrument designed for
processing standard microscope slides. Accordingly, the outer
dimensions of slide holder 32 are the same as or approximately the
same as the outer dimensions of a slide holder for a standard size
slide. The dimensions of slide holding section 30 may receive and
retain non-standard size slide 14 of any size smaller that a
standard size slide by positioning adjustable member 34 in an
appropriate position. In this way, slide holder 32 may be received
in a known processing apparatus in many instances designed for
processing standard microscope slides and a non-standard size slide
of any non-standard size may be processed in the known processing
apparatus.
[0055] For example, holder 22 with mounted slide 14 may be inserted
into a processing instrument such as, for example, a reader, such
as a laser scanner, which has a suitable mounting means for
receiving and releasably retaining holder 22 in a known position.
In some embodiments the scanner is able to read the location and
intensity of fluorescence at each feature of an array following
exposure to a fluorescently labeled sample (such as a
polynucleotide containing sample). For example, such a scanner may
be similar to the G2565.RTM. DNA microarray scanner available from
Agilent Technologies, Inc., Palo Alto, Calif. The array 16 may then
be read through front side 14a of slide 14. Usually, a scanning
interrogating laser beam (not shown) is directed through a beam
splitter and then through front side 14a and scanned across array
16. Resulting fluorescent signals from the array that have passed
back through slide 14 and out through front side 14a may then be
detected at a detector. Results from the interrogation can be
processed such as by rejecting a reading for a feature that is
below a predetermined threshold and/or forming conclusions based on
the pattern read from the array (such as whether or not a
particular target sequence may have been present in the sample).
The results of the interrogation or processing can be forwarded
(such as by communication) to a remote location if desired, for
further use.
[0056] As mentioned above, the reading may include directing a
light beam through the slide from the front side and onto the array
on the rear side, and detecting a resulting signal from the array,
which has passed from the rear side through the slide and out the
slide front side.
[0057] The holder may further include retaining means such as,
e.g., front and rear clamp sets, which can be moved apart to
receive the slide between the sets. In this case, the slide is
retained in the mounted position by the clamp sets being urged
(such as resiliently, for example by one or more springs) against
portions of the front and rear surfaces, respectively. The clamp
sets may, for example, be urged against the slide front and rear
surfaces of a mounted slide at positions adjacent a periphery of
that slide. Alternatively, the array may be read on the front side
when the slide is positioned in the holder with the array facing
forward (that is, away from the holder).
[0058] Identifier 18 may be read from the front side of slide 14 by
a bar code reader where the identifier is a bar code. The
identifier is usually placed in a location that does not interfere
with the processing of the arrays. Information from the read bar
code can be used to retrieve array layout information, which can be
used in the reading and/or processing of the interrogation results.
After the reading of array 18 is complete, the holder 22 may be
removed from the scanner. A user may now remove slide 14 for
storage or disposal. Removal of slide 14 from holder 22 depends on
the particular design of the slide holding portion of the slide
holder. Such removal may involve, among others, disengaging any
retaining means and removing the slide by prying, sliding, applying
pressure or force, releasing restraints, and the like.
[0059] Any suitable examining approach may be utilized. The nature
of the examining device including a detector for examining the
array for the results of one or more chemical reactions is
dependent on the nature of the chemical reactions including any
label employed for detection, such as fluorescent as mentioned
above, chemiluminescent, calorimetric based on an attached enzyme,
and the like. As mentioned above, the examining device may be a
scanning device involving an imaging system or optical system.
Other known examining devices may be employed. Such devices may
involve the use of other optical techniques (for example, optical
techniques for detecting chemiluminescent or electroluminescent
labels) or electrical techniques (where each feature is provided
with an electrode to detect hybridization at that feature in a
manner disclosed in U.S. Pat. Nos. 6,221,583 and 6,251,685, and
elsewhere). Other examining techniques include visual inspection
techniques, non-light based methods, and so forth.
[0060] Prior to reading an array, the array is contacted with a
sample to be analyzed. In one approach involving polynucleotide
analysis, cell matter is lysed, to release its DNA and RNA as
fragments, which are then separated out by gel filtration, membrane
purification, precipitation, solvent extraction, electrophoresis or
other means, and then tagged with a fluorescent or other label. The
nucleic acid mix is exposed to the array of oligonucleotide probes,
whereupon selective attachment to matching probe sites takes place.
The array is then washed and the result of exposure to the array is
determined. In this particular example, the array is imaged by
scanning the surface of the support so as to reveal for analysis
and interpretation the sites where attachment occurred.
[0061] The signal referred to above may arise from any moiety that
may be incorporated into the sample being analyzed for the purpose
of detection. Often, a label is employed, which may be a member of
a signal producing system. The label is capable of being detected
directly or indirectly. In general, any reporter molecule that is
detectable can be a label. Labels include, for example, (i)
reporter molecules that can be detected directly by virtue of
generating a signal, (ii) specific binding or reacting pair members
that may be detected indirectly by subsequent binding or reacting
to a cognate that contains a reporter molecule, (iii) mass tags
detectable by mass spectrometry, (iv) oligonucleotide primers that
can provide a template for amplification or ligation, (v) specific
labeled nucleotide monomers which are incorporated into the target
samples by enzymatic or chemical incorporation means, and (vi) a
specific polynucleotide sequence or recognition sequence that can
act as a ligand such as for a repressor protein, wherein in the
latter two instances the oligonucleotide primer or repressor
protein will have, or be capable of having, a reporter molecule and
so forth. The reporter molecule can be a catalyst, such as an
enzyme, a polynucleotide coding for a catalyst, promoter, dye,
fluorescent molecule, chemiluminescent molecule, coenzyme, enzyme
substrate, radioactive group, a small organic molecule, amplifiable
polynucleotide sequence, a particle such as latex or carbon
particle, metal sol, crystallite, liposome, cell, etc., which may
or may not be further labeled with a dye, catalyst or other
detectable group, a mass tag that alters the weight of the molecule
to which it is conjugated for mass spectrometry purposes, and the
like.
[0062] The signal may be produced by a signal producing system,
which is a system that generates a signal that relates to the
presence or amount of a target polynucleotide in a medium. The
signal producing system may have one or more components, at least
one component being the label. The signal producing system includes
all of the reagents required to produce a measurable signal. The
signal producing system provides a signal detectable by external
means, by use of electromagnetic radiation, desirably by visual
examination.
[0063] The arrays prepared as described above are particularly
suitable for conducting hybridization reactions. Such reactions are
carried out on an array comprising a plurality of features relating
to the hybridization reactions. The array is exposed to liquid
samples and to other reagents for carrying out the hybridization
reactions. The support surface exposed to the sample is incubated
under conditions suitable for hybridization reactions to occur.
[0064] After the appropriate period of time of contact between the
liquid sample and the array, the contact is discontinued and
various processing steps are performed. Following the processing
step(s), the array is moved to an examining device as discussed
above where the array is interrogated.
[0065] Results from the reading may be raw results (such as
fluorescence intensity readings for each feature in one or more
color channels) or may be processed results such as obtained by
rejecting a reading for a feature that is below a predetermined
threshold and/or forming conclusions based on the pattern read from
the array (such as whether or not a particular target sequence may
have been present in the sample). The results of the reading
(processed or not) may be forwarded (such as by communication) to a
remote location if desired, and received there for further use
(such as further processing).
[0066] When one item is indicated as being "remote" from another,
this means that the two items are at least in different buildings
and may be at least one mile, ten miles, or at least one hundred
miles apart. "Communicating" information references transmitting
the data representing that information as electrical signals over a
suitable communication channel (for example, a private or public
network). "Forwarding" an item refers to any means of getting that
item from one location to the next, whether by physically
transporting that item or otherwise (where that is possible) and
includes, at least in the case of data, physically transporting a
medium carrying the data or communicating the data.
[0067] As mentioned above, in the present invention, the surface of
the individual support normally has only one array. Depending upon
intended use, the array may contain multiple spots or features of
chemical compounds such as, e.g., biopolymers in the form of
polynucleotides or other biopolymer. A typical array on an
individual support may contain more than ten, more than one
hundred, more than five hundred, more than one thousand, more than
fifteen hundred, more than two thousand, more than twenty five
hundred features. In many embodiments the number of features on the
individual supports is in the range of about 100 to about 25,000,
about 200 to about 20,000, about 300 to about 15,000, about 500 to
about 10,000, about 1000 to about 3000, about 1500 to about 2500,
about 1800 to about 2200. The features may occupy an area of less
than 20 cm.sup.2 or even less than 10 cm.sup.2. For example,
features may have widths (that is, diameter, for a round spot) in
the range from a 10 .mu.m to 1.0 cm. In other embodiments each
feature may have a width in the range of 1.0 .mu.m to 1.0 mm,
usually 5.0 .mu.m to 500 .mu.m, and more usually 10 .mu.m to 200
.mu.m. Non-round features may have area ranges equivalent to that
of circular features with the foregoing width (diameter)
ranges.
[0068] In comparison, standard microscope slides may, and usually
do, contain more than one array. The number of features on the
standard microscope slide may be more than 20,000, more than
25,000, more than 30,000, more than 35,000, more than 40,000, more
than 50,000, more than 75,000, or more than 100,000 features.
[0069] Each feature, or element, within the molecular array is
defined to be a small, regularly shaped region of the surface of
the substrate. The features are arranged in a predetermined manner.
Each feature of an array usually carries a predetermined chemical
compound or mixtures thereof. Each feature within the molecular
array may contain a different molecular species, and the molecular
species within a given feature may differ from the molecular
species within the remaining features of the molecular array. Some
or all of the features may be of different compositions. Each array
may contain multiple spots or features separated by spaces or areas
that have no features. Such interfeature areas are usually present
but are not essential. As with the border areas discussed above,
these interfeature areas do not carry any chemical compound such as
polynucleotide (or other biopolymer of a type of which the features
are composed). Interfeature areas typically will be present where
arrays are formed by the conventional in situ process or by
deposition of previously obtained moieties, as described above, by
depositing for each feature at least one droplet of reagent such as
from a pulse jet (for example, an inkjet type head) but may not be
present when, for example, photolithographic array fabrication
processes are used. It will be appreciated though, that the
interfeature areas, when present, could be of various sizes and
configurations.
[0070] As mentioned above, in the present methods individual
supports are prepared having a plurality of chemical compounds in
the form of an array on the surface of such supports. The chemical
compounds may be deposited on the surface of the support as fully
formed moieties. On the other hand, the chemical compounds may be
synthesized in situ in a series of steps such as, for example, the
addition of building blocks, which are chemical components of the
chemical compound. Examples of such building blocks are those found
in the synthesis of polymers. The invention has particular
application to chemical compounds that are biopolymers such as
polynucleotides, for example, oligonucleotides.
[0071] As referred to above, embodiments of the invention have
particular application to supports bearing oligomers or polymers.
The oligomer or polymer is a chemical entity that contains a
plurality of monomers. It is generally accepted that the term
"oligomers" is used to refer to a species of polymers. The terms
"oligomer" and "polymer" may be used interchangeably herein.
Polymers usually comprise at least two monomers. Oligomers
generally comprise about 6 to about 20,000 monomers, preferably,
about 10 to about 10,000, more preferably about 15 to about 4,000
monomers. Examples of polymers include polydeoxyribonucleotides,
polyribonucleotides, other polynucleotides that are C-glycosides of
a purine or pyrimidine base, or other modified polynucleotides,
polypeptides, polysaccharides, and other chemical entities that
contain repeating units of like chemical structure. Exemplary of
oligomers are oligonucleotides and peptides.
[0072] A monomer is a chemical entity that can be covalently linked
to one or more other such entities to form an oligomer or polymer.
Examples of monomers include nucleotides, amino acids, saccharides,
peptoids, and the like and subunits comprising nucleotides, amino
acids, saccharides, peptoids and the like. The subunits may
comprise all of the same component such as, for example, all of the
same nucleotide or amino acid, or the subunit may comprise
different components such as, for example, different nucleotides or
different amino acids. The subunits may comprise about 2 to about
2000, or about 5 to about 200, monomer units. In general, the
monomers have first and second sites (e.g., C-termini and
N-termini, or 5' and 3' sites) suitable for binding of other like
monomers by means of standard chemical reactions (e.g.,
condensation, nucleophilic displacement of a leaving group, or the
like), and a diverse element that distinguishes a particular
monomer from a different monomer of the same type (e.g., an amino
acid side chain, a nucleotide base, etc.). The initial
substrate-bound, or support-bound, monomer is generally used as a
building block in a multi-step synthesis procedure to form a
complete ligand, such as in the synthesis of oligonucleotides,
oligopeptides, oligosaccharides, etc. and the like.
[0073] A biomonomer references a single unit, which can be linked
with the same or other biomonomers to form a biopolymer (for
example, a single amino acid or nucleotide with two linking groups
one or both of which may have removable protecting groups). A
biomonomer fluid or biopolymer fluid refers to a liquid containing
either a biomonomer or biopolymer, respectively (typically in
solution).
[0074] A biopolymer is a polymer of one or more types of repeating
units. Biopolymers are typically found in biological systems and
particularly include polysaccharides (such as carbohydrates), and
peptides (which term is used to include polypeptides, and proteins
whether or not attached to a polysaccharide) and polynucleotides as
well as their analogs such as those compounds composed of or
containing amino acid analogs or non-amino acid groups, or
nucleotide analogs or non-nucleotide groups. This includes
polynucleotides in which the conventional backbone has been
replaced with a non-naturally occurring or synthetic backbone, and
nucleic acids (or synthetic or naturally occurring analogs) in
which one or more of the conventional bases has been replaced with
a group (natural or synthetic) capable of participating in
Watson-Crick type hydrogen bonding interactions.
[0075] Polynucleotides are compounds or compositions that are
polymeric nucleotides or nucleic acid polymers. The polynucleotide
may be a natural compound or a synthetic compound. Polynucleotides
include oligonucleotides and are comprised of natural nucleotides
such as ribonucleotides and deoxyribonucleotides and their
derivatives although unnatural nucleotide mimetics such as
2'-modified nucleosides, peptide nucleic acids and oligomeric
nucleoside phosphonates are also used. The polynucleotide can have
from about 2 to 5,000,000 or more nucleotides. Usually, the
oligonucleotides are at least about 2 nucleotides, usually, about 5
to about 100 nucleotides, more usually, about 10 to about 50
nucleotides, and may be about 15 to about 30 nucleotides, in
length. Polynucleotides include single or multiple stranded
configurations, where one or more of the strands may or may not be
completely aligned with another.
[0076] A nucleotide refers to a sub-unit of a nucleic acid and has
a phosphate group, a 5 carbon sugar and a nitrogen containing base,
as well as functional analogs (whether synthetic or naturally
occurring) of such sub-units which in the polymer form (as a
polynucleotide) can hybridize with naturally occurring
polynucleotides in a sequence specific manner analogous to that of
two naturally occurring polynucleotides. For example, a
"biopolymer" includes DNA (including cDNA), RNA, oligonucleotides,
and PNA and other polynucleotides as described in U.S. Pat. No.
5,948,902 and references cited therein (all of which are
incorporated herein by reference), regardless of the source. An
"oligonucleotide" generally refers to a nucleotide multimer of
about 10 to 100 nucleotides in length, while a "polynucleotide"
includes a nucleotide multimer having any number of
nucleotides.
[0077] The nature of the support or substrate to which a plurality
of chemical compounds is attached is discussed above. The support
can be hydrophilic or capable of being rendered hydrophilic or it
may be hydrophobic. The support is usually glass such as flat glass
whose surface has been chemically activated for binding thereto or
synthesis thereon, glass available as Bioglass and the like. The
surface of a support is normally treated to create a primed or
functionalized surface, that is, a surface that is able to support
the attachment of a fully formed chemical compound or the synthetic
steps involved in the production of the chemical compound on the
surface of the support. Functionalization relates to modification
of the surface of a support to provide a plurality of functional
groups on the support surface. By the term "functionalized surface"
is meant a support surface that has been modified so that a
plurality of functional groups are present thereon usually at
discrete sites on the surface. The manner of treatment is dependent
on the nature of the chemical compound to be synthesized and on the
nature of the support surface. In one approach a reactive
hydrophilic site or reactive hydrophilic group is introduced onto
the surface of the support. Such hydrophilic moieties can be used
as the starting point in a synthetic organic process.
[0078] In one embodiment, the surface of the support, such as a
glass support, is siliceous, i.e., the surface comprises silicon
oxide groups, either present in the natural state, e.g., glass,
silica, silicon with an oxide layer, etc., or introduced by
techniques well known in the art. One technique for introducing
siloxyl groups onto the surface involves reactive hydrophilic
moieties on the surface. These moieties are typically epoxide
groups, carboxyl groups, thiol groups, and/or substituted or
unsubstituted amino groups as well as a functionality that may be
used to introduce such a group such as, for example, an olefin that
may be converted to a hydroxyl group by means well known in the
art. One approach is disclosed in U.S. Pat. No. 5,474,796
(Brennan), the relevant portions of which are incorporated herein
by reference. A siliceous surface may be used to form silyl
linkages, i.e., linkages that involve silicon atoms. Usually, the
silyl linkage involves a silicon-oxygen bond, a silicon-halogen
bond, a silicon-nitrogen bond, or a silicon-carbon bond.
[0079] Another method for attachment is described in U.S. Pat. No.
6,219,674 (Fulcrand, et al.). A surface is employed that comprises
a linking group consisting of a first portion comprising a
hydrocarbon chain, optionally substituted, and a second portion
comprising an alkylene oxide or an alkylene imine wherein the
alkylene is optionally substituted. One end of the first portion is
attached to the surface and one end of the second portion is
attached to the other end of the first portion chain by means of an
amine or an oxy functionality. The second portion terminates in an
amine or a hydroxy functionality. The surface is reacted with the
substance to be immobilized under conditions for attachment of the
substance to the surface by means of the linking group.
[0080] Another method for attachment is described in U.S. Pat. No.
6,258,454 (Lefkowitz, et al.). A solid support having hydrophilic
moieties on its surface is treated with a derivatizing composition
containing a mixture of silanes. A first silane provides the
desired reduction in surface energy, while the second silane
enables functionalization with molecular moieties of interest, such
as small molecules, initial monomers to be used in the solid phase
synthesis of oligomers, or intact oligomers. Molecular moieties of
interest may be attached through cleavable sites.
[0081] A procedure for the derivatization of a metal oxide surface
uses an aminoalkyl silane derivative, e.g., trialkoxy
3-aminopropylsilane such as aminopropyltriethoxy silane (APS),
4-aminobutyltrimethoxysilane, 4-aminobutyltriethoxysilane,
2-aminoethyltriethoxysilane, and the like. APS reacts readily with
the oxide and/or siloxyl groups on metal and silicon surfaces. APS
provides primary amine groups that may be used to carry out the
present methods. Such a derivatization procedure is described in EP
0 173 356 B1, the relevant portions of which are incorporated
herein by reference. Other methods for treating the surface of a
support will be suggested to those skilled in the art in view of
the teaching herein.
[0082] The devices and methods of the present invention are
particularly useful for the preparation of individual supports with
an array of biopolymers. An array includes any one-, two- or
three-dimensional arrangement of addressable regions bearing a
particular biopolymer such as polynucleotides, associated with that
region. An array is addressable in that it has multiple regions of
different moieties, for example, different polynucleotide
sequences, such that a region or feature or spot of the array at a
particular predetermined location or address on the array can
detect a particular target molecule or class of target molecules
although a feature may incidentally detect non-target molecules of
that feature.
[0083] Normally, the surface of the support opposite the surface
with the array (opposing surface) does not carry any arrays. The
arrays can be designed for testing against any type of sample,
whether a trial sample, a reference sample, a combination of the
foregoing, or a known mixture of components such as
polynucleotides, proteins, polysaccharides and the like (in which
case the arrays may be composed of features carrying unknown
sequences to be evaluated).
[0084] The devices and methods of the present invention are
particularly useful in the preparation of individual supports with
oligonucleotide arrays for determinations of polynucleotides. As
explained briefly above, in the field of bioscience, arrays of
oligonucleotide probes, fabricated or deposited on a surface of a
support, are used to identify DNA sequences in cell matter. The
arrays generally involve a surface containing a mosaic of different
oligonucleotides or sample nucleic acid sequences or
polynucleotides that are individually localized to discrete, known
areas of the surface.
[0085] As mentioned above, biopolymer arrays can be fabricated by
depositing previously obtained biopolymers (such as from synthesis
or natural sources) onto a substrate, or by in situ synthesis
methods. The in situ method for fabricating a polynucleotide array
typically follows, at each of the multiple different addresses at
which features are to be formed, the same conventional iterative
sequence used in forming polynucleotides from nucleoside reagents
on a support by means of known chemistry. This iterative sequence
is as follows: (a) coupling a selected nucleoside through a
phosphite linkage to a functionalized support in the first
iteration, or a nucleoside bound to the substrate (i.e. the
nucleoside-modified substrate) in subsequent iterations; (b)
optionally, but preferably, blocking unreacted hydroxyl groups on
the substrate bound nucleoside; (c) oxidizing the phosphite linkage
of step (a) to form a phosphate linkage; and (d) removing the
protecting group ("deprotection") from the now substrate bound
nucleoside coupled in step (a), to generate a reactive site for the
next cycle of these steps. The functionalized support (in the first
cycle) or deprotected coupled nucleoside (in subsequent cycles)
provides a substrate bound moiety with a linking group for forming
the phosphite linkage with a next nucleoside to be coupled in step
(a). A number of reagents involved in the above synthetic steps
such as, for example, phosphoramidite reagents, are sensitive to
moisture and anhydrous conditions and solvents are employed. Final
deprotection of nucleoside bases can be accomplished using alkaline
conditions such as ammonium hydroxide, in a known manner.
[0086] The foregoing chemistry of the synthesis of polynucleotides
is described in detail, for example, in Caruthers, Science 230:
281-285, 1985; Itakura, et al., Ann. Rev. Biochem. 53: 323-356;
Hunkapillar, et al., Nature 310: 105-110, 1984; and in "Synthesis
of Oligonucleotide Derivatives in Design and Targeted Reaction of
Oligonucleotide Derivatives", CRC Press, Boca Raton, Fla., pages
100 et seq., U.S. Pat. Nos. 4,458,066, 4,500,707, 5,153,319, and
5,869,643, EP 0294196, and elsewhere.
[0087] As mentioned above, various ways may be employed to
introduce the reagents for producing an array of polynucleotides on
the surface of a support such as a glass support. Such methods are
known in the art. One such method involves solid phase chemistry,
photolabile protecting groups and photolithography. Binary masking
techniques are employed in one embodiment of the above. Arrays are
fabricated in situ, adding one base pair at a time to a primer
site. Photolithography is used to uncover sites, which are then
exposed and reacted with one of the four base pair
phosphoramidites. In photolithography the surface is first coated
with a light-sensitive resist, exposed through a mask and the
predetermined area is revealed by dissolving away the exposed or
the unexposed resist and, subsequently, a surface layer. A separate
mask is usually made for each predetermined area, which may involve
one for each base pair in the length of the probe.
[0088] Another in situ method employs inkjet printing technology to
dispense the appropriate phosphoramidite reagents and other
reagents onto individual sites on a surface of a support.
Oligonucleotides are synthesized on a surface of a substrate in
situ using phosphoramidite chemistry. Solutions containing
nucleotide monomers and other reagents as necessary such as an
activator, e.g., tetrazole, are applied to the surface of a support
by means of thermal ink-jet technology. Individual droplets of
reagents are applied to reactive areas on the surface using, for
example, a thermal ink-jet type nozzle. The surface of the support
may have an alkyl bromide trichlorosilane coating to which is
attached polyethylene glycol to provide terminal hydroxyl groups.
These hydroxyl groups provide for linking to a terminal primary
amine group on a monomeric reagent. Excess of non-reacted chemical
on the surface is washed away in a subsequent step. For example,
see U.S. Pat. No. 5,700,637.
[0089] Another approach for fabricating an array of biopolymers on
a substrate using a biopolymer or biomonomer fluid and using a
fluid dispensing head is described in U.S. Pat. No. 6,242,266
(Schleifer, et al.). The head has at least one jet that can
dispense droplets onto a surface of a support. The jet includes a
chamber with an orifice and an ejector, which, when activated,
causes a droplet to be ejected from the orifice. Multiple droplets
of the biopolymer or biomonomer fluid are dispensed from the head
orifice so as to form an array of droplets on the surface of the
substrate.
[0090] In another embodiment (U.S. Pat. No. 6,232,072) (Fisher) a
method of, and apparatus for, fabricating a biopolymer array is
disclosed. Droplets of fluid carrying the biopolymer or biomonomer
are deposited onto a front side of a transparent substrate. Light
is directed through the substrate from the front side, back through
a substrate backside and a first set of deposited droplets on the
first side to an image sensor.
[0091] An example of another method for chemical array fabrication
is described in U.S. Pat. No. 6,180,351 (Cattell). The method
includes receiving from a remote station information on a layout of
the array and an associated first identifier. A local identifier is
generated corresponding to the first identifier and associated
array. The local identifier is shorter in length than the
corresponding first identifier. The addressable array is fabricated
on the substrate in accordance with the received layout
information.
[0092] Other methods for synthesizing arrays of oligonucleotides on
a surface include those disclosed by Gamble, et al., WO97/44134;
Gamble, et al., WO98/10858; Baldeschwieler, et al., WO95/25116;
Brown, et al., U.S. Pat. No. 5,807,522; and the like.
[0093] In accordance with the present invention and as described
above, a sheet of glass, by way of illustration and not limitation,
is first treated to form score lines on one of its surfaces. Next,
arrays are formed on the surface of the sheet of glass in
predetermined areas within the boundaries of the scored sections of
the sheet of glass where each of the sections has a single array.
Then, the sheet of glass is divided along to form individual
supports or slides in accordance with the present invention where
the slides have non-standard size dimensions. The sheet of glass
may be divided by any suitable technique for physically separating
portions of the sheet from other portions. For example, score lines
may be made on the surface of the glass to correspond with the size
of the individual supports or slides. The sheet may be divided by
breaking, dicing or otherwise severing the sheet along the score
lines. As is evident, such an approach may be applied to any
breakable substrate employed for attaching the array features.
[0094] All publications and patent applications cited in this
specification are herein incorporated by reference as if each
individual publication or patent application were specifically and
individually indicated to be incorporated by reference.
[0095] Although the foregoing invention has been described in some
detail by way of illustration and example for purposes of clarity
of understanding, it will be readily apparent to those of ordinary
skill in the art in light of the teachings of this invention that
certain changes and modifications may be made thereto without
departing from the spirit or scope of the appended claims.
Furthermore, the foregoing description, for purposes of
explanation, used specific nomenclature to provide a thorough
understanding of the invention. However, it will be apparent to one
skilled in the art that the specific details are not required in
order to practice the invention. Thus, the foregoing descriptions
of specific embodiments of the present invention are presented for
purposes of illustration and description; they are not intended to
be exhaustive or to limit the invention to the precise forms
disclosed. Many modifications and variations are possible in view
of the above teachings. The embodiments were chosen and described
in order to explain the principles of the invention and its
practical applications and to thereby enable others skilled in the
art to utilize the invention.
* * * * *