(12) United States Patent
`Nova et al.
`
`US006372428B1
`US 6,372,428 B1
`*Apr. 16, 2002
`
`(10) Patent N0.:
`(45) Date of Patent:
`
`(54) REMOTELY PROGRAMMABLE MATRICES
`WITH MEMORIES
`
`(75) Inventors: Michael P. Nova, Rancho Santa Fe;
`Andrew E. Senyei; Gary S. David,
`both of La Jolla, all of CA (US)
`
`(73) Assignee: Discovery Partners International,
`Inc., San Diego, CA (US)
`
`( * ) NOIiCeI
`
`Subject to any disclaimer, the term of this
`patent is extended or adjusted under 35
`U.S.C. 154(b) by 317 days.
`
`This patent is subject to a terminal dis
`claimer.
`
`(21) Appl. No.: 09/098,122
`(22) Filed:
`Jun. 16, 1998
`
`Related US. Application Data
`
`(63) Continuation of application No. 08/538,387, ?led on Oct. 3,
`1995, now Pat. No. 5,874,214, which is a continuation-in
`part of application No. 08/480,147, ?led on Jun. 7, 1995, and
`a continuation-in-part of application No. 08/484,486, ?led
`on Jun. 7, 1995, and a continuation-in-part of application
`No. 08/484,504, ?led on Jun. 7, 1995, now Pat. No. 5,751,
`629, and a continuation-in-part of application No. 08/480,
`196, ?led on Jun. 7, 1995, now Pat. No. 5,925,562, and a
`continuation-in-part of application No. 08/473,660, ?led on
`Jun. 7, 1995, which is a continuation-in-part of application
`No. 08/428,662, ?led on Apr. 25, 1995, now Pat. No.
`5,741,462.
`(51) Im. c1? ............................................... .. C12M 1/34
`(52) US. Cl. .............................. .. 435/6; 435/6; 435/71;
`435/288.1; 435/288.3; 435/288.4; 435/288.7;
`436/501; 536/243
`(58) Field of Search ........................ .. 435/6, 7.1, 287.1,
`435/2872, 288.1, 288.3, 288.4, 288.7; 436/501;
`536/243
`
`(56)
`
`References Cited
`
`U.S. PATENT DOCUMENTS
`
`1/1964 Biernat
`3,119,099 A
`9/1972 Frank et al.
`3,692,498 A
`3,781,120 A 12/1973 Engelhardt
`3,882,318 A
`5/1975 Mioduski
`4,133,642 A
`1/1979 Nosaka et al.
`4,154,795 A
`5/1979 Thorne
`4,855,909 A
`8/1989 Vincent et al.
`4,935,875 A
`6/1990 Shah et al.
`5,075,077 A 12/ 1991 Durley, III et al.
`5,120,662 A
`6/1992 Chan et al.
`5,214,409 A
`5/1993 Beigel
`5,348,705 A
`9/1994 Koreyasu et al.
`5,474,796 A 12/1995 Brennan et al.
`5,565,324 A 10/1996 Still et al.
`5,585,275 A 12/1996 Hudson et al.
`5,605,662 A
`2/1997 Heller et al.
`5,639,603 A
`6/1997 Dower et al.
`5,641,634 A
`6/1997 Mandecki
`5,708,153 A
`1/1998 Dower et al.
`5,741,462
`A
`4/1998 Nova et al.
`5,751,629
`A
`5/1998 Nova et al.
`
`6/1998 Cargill et al.
`5,770,455 A
`2/1999 Nova et al.
`5,874,214 A
`7/1999 Nova et al.
`5,925,562 A
`5,961,923 A 10/1999 Nova et al.
`
`FOREIGN PATENT DOCUMENTS
`
`DE
`DE
`DE
`DE
`EP
`EP
`EP
`EP
`EP
`EP
`FR
`GB
`JP
`WO
`WO
`WO
`WO
`WO
`WO
`
`9/1993
`43 10 169 A1
`43 13 807 A1 11/1993
`43 06 563 A1
`9/1994
`94 16 270 U1 12/1994
`0 196 174 A2 10/1986
`0 410 688 A2 10/1991
`0 541 340 A2
`5/1993
`0 554 955 A1
`53/1993
`0 378 059 B1
`9/1993
`0 640 826 A1
`3/1995
`2555744
`5/1985
`2129551
`5/1984
`57 016359 A
`5/1982
`WO94/13402
`6/1994
`WO97/15390
`5/1997
`WO98/11036
`3/1998
`WO98/46548
`10/1998
`WO98/46549
`10/1998
`WO98/46550
`10/1998
`
`OTHER PUBLICATIONS
`Beck—Sickinger et al., “Semiautomated T—Bag Peptide Syn
`thesis Using 9—Fluorenyl—Methoxycarbonyl Strategy and
`BenZotriaZol—1—yl—Tetramethyl—Uronium Tetra?uoroborate
`Activation”, Peptide Research, vol. 4, No. 2, Mar.—Apr.
`1991, pp. 88—94.
`(List continued on next page.)
`Primary Examiner—Stephanie Zitomer
`(74) Attorney, Agent, or Firm—Kilpatrick Stockton LLP;
`Eleanor M. Musick
`(57)
`
`ABSTRACT
`
`Combinations, called matrices with memories, of matrix
`materials with remotely addressable or remotely program
`mable recording devices that contain at least one data
`storage unit are provided. The matrix materials are those that
`are used in as supports in solid phase chemical and bio
`chemical syntheses, immunoassays and hybridization reac
`tions. The data storage units are non-volatile antifuse memo
`ries or volatile memories, such as EEPROMS, DRAMS or
`?ash memory. By virtue of this combination, molecules and
`biological particles, such as phage and viral particles and
`cells, that are in proximity or in physical contact with the
`matrix combination can be labeled by programming the
`memory with identifying information and can be identi?ed
`by retrieving the stored information. Combinations of matrix
`materials, memories, and linked molecules and biological
`materials are also provided. The combinations have a mul
`tiplicity of applications, including combinatorial chemistry,
`isolation and puri?cation of target macromolecules, capture
`and detection of macromolecules for analytical purposes,
`selective removal of contaminants, enzymatic catalysis, cell
`sorting, drug delivery, chemical modi?cation and other uses.
`Methods for electronically tagging molecules, biological
`particles and matrix support materials, immunoassays,
`receptor binding assays, and other methods are also pro
`vided.
`
`59 Claims, 9 Drawing Sheets
`
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`

`US 6,372,428 B1
`Page 2
`
`OTHER PUBLICATIONS
`
`Jung et a1., “Multiple Peptide Synthesis Methods and Their
`App1ications”,Angew. Chem. Int. Ed. EngL, vol. 31, No. 4,
`Apr. 1992, pp. 367—383.
`J anda, K.D., “Tagged versus untagged libraries: Methods for
`the generation and screening of combinatorial chemical
`libraries”, Proc. Natl. Acad. Sci. USA, vol. 91, Nov. 1994,
`pp. 10779—10785.
`
`Nicolaou et a1., “RadiofrequenZ—versch1iisselte kornbina
`torische Chernie”, Angewandte Chemie, vol. 107, No. 20,
`Oct. 16, 1995, pp. 2476—2479, Weinheirn, DE.
`Moran et a1., “Radio Frequency Tag Encoded Cornbinatorial
`Library Method for the Discovery of Tripeptide—Substituted
`Cinnarnic Acid Inhibitors of the Protein Tyrosine Phos
`phatase PTPlB,” J. Am. Chem. Soc, 117:10787—10788,
`1995.
`
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`

`

`U.S. Patent
`
`Apr. 16, 2002
`
`Sheet 1 0f 9
`
`US 6,372,428 B1
`
`COMBINE
`
`$2
`
`COMBINE
`
`COMBINE
`
`COMBINE
`
`SPLIT
`
`COMBINE \A/V
`¢
`(28
`
`E
`
`Y
`
`COMBINE
`
`SPLIT
`
`[H]
`
`FIG. I
`
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`

`

`U.S. Patent
`
`Apr. 16, 2002
`
`Sheet 2 0f 9
`
`US 6,372,428 B1
`
`COMBINE
`
`LPIPERIDINE
`
`COMBINE
`
`[Lys—Fm0c], DIC
`
`1
`@ @
`
`9
`
`O
`
`SPLIT
`
`[Phe—Fmoc], BIG
`8d
`l
`@ -
`
`'
`
`liys—Fmoc
`
`Lys—Fm0c
`I
`COMBINE
`
`LPIPERIDINE
`
`—Fmoc
`
`Phe-Fmoc
`J
`COMBINE
`
`000-1-——
`
`QOQ4———
`
`FIG. 2
`
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`Luminex/Irori - Page 4
`
`

`

`Luminex Ex. 1002
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`
`

`

`U.S. Patent
`
`Apr. 16, 2002
`
`Sheet 4 0f 9
`
`US 6,372,428 B1
`
`'-
`
`'
`
`1. SPLIT
`2. ADD R1
`3. COMBINE
`S1
`
`R1
`
`'52
`
`1. SPLIT
`2. ADD R2
`:5. COMBINE
`
`R
`1
`
`L
`
`R2
`
`L
`
`I
`R2
`
`1. SPLIT
`2. ADD R3
`3. COMBINE
`Rslf S3
`R1
`
`FIG. 4
`
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`

`

`U.S. Patent
`
`Apr. 16, 2002
`
`Sheet 5 0f 9
`
`US 6,372,428 B1
`
`780
`
`~~12O
`
`Y
`O
`1
`2
`
`g
`6
`g
`
`.
`
`120
`f
`#XY
`
`A 18
`c 24
`E 32
`
`FIG. 6
`
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`
`

`

`U.S. Patent
`
`Apr. 16, 2002
`
`Sheet 6 6f 9
`
`US 6,372,428 B1
`
`W122
`
`"-120
`
`FIG. 7
`
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`

`

`U.S. Patent
`
`Apr. 16, 2002
`
`Sheet 7 0f 9
`
`US 6,372,428 B1
`
`$2“- 212
`
`fzos 290
`204
`\ n
`H
`/
`210
`u?
`
`208
`
`202
`/
`(
`
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`
`

`

`U.S. Patent
`
`Apr. 16, 2002
`
`Sheet 8 0f 9
`
`US 6,372,428 B1
`
`80
`
`Y
`
`PHmqDEl-Em/ AMPLIFIER
`
`100
`
`H
`II
`
`II ::
`ii
`
`II
`
`1
`TRANSISTOR
`
`FIG. 9
`
`120
`
`'/
`
`I? X Y
`A 18
`_, c 24
`E 32
`
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`

`US 6,372,428 B1
`
`1
`REMOTELY PROGRAMMABLE MATRICES
`WITH MEMORIES
`
`RELATED APPLICATIONS
`
`This application is a continuation on US. application Ser.
`No. 08/538,387, ?led Oct. 3, 1995, now issued as US. Pat.
`No. 5,874,214, which is a continuation-in-part of each of
`application Ser. Nos. 08/480,147, 08/484,486, 08/484,504,
`now issued as US. Pat. No. 5,751,629, Ser. No. 08/480,196,
`now issued as US. Pat. No. 5,925,562, and Ser. No. 08/473,
`660, each ?led Jun. 7, 1995, which are continuations-in-part
`of US. application Ser. No. 08/428,662, ?led Apr. 25, 1995,
`now issued as US. Pat. No. 5,741,462. The subject matter of
`each cited application is incorporated herein by reference in
`its entirety.
`
`10
`
`15
`
`FIELD OF THE INVENTION
`
`The present invention relates to the application of data
`storage technology to molecular tracking and identi?cation
`and to biological and biochemical assays.
`
`20
`
`BACKGROUND OF THE INVENTION
`
`There has been a convergence of progress in chemistry
`and biology. Among the important advances resulting from
`this convergence is the development of methods for gener
`ating molecular diversity and for detecting and quantifying
`biological or chemical material. This advance has been
`facilitated by fundamental developments in chemistry,
`including the development of highly sensitive analytical
`methods, solid state chemical synthesis, and sensitive and
`speci?c biological assay systems.
`Analyses of biological interactions and chemical
`reactions, however, require the use of labels or tags to track
`and identify the results of such analyses. Typically biologi
`cal reactions, such as binding, catalytic, hybridiZation and
`signaling reactions, are monitored by labels, such as
`radioactive, ?uorescent, photoabsorptive, luminescent and
`other such labels, or by direct or indirect enZyme labels.
`Chemical reactions are also monitored by direct or indirect
`means, such as by linking the reactions to a second reaction
`in which a colored, ?uorescent, chemiluminescent or other
`such product results. These analytical methods, however, are
`often time consuming, tedious and, when practiced in vivo,
`invasive. In addition, each reaction is typically measured
`individually, in a separate assay. There is, thus, a need to
`develop alternative and convenient methods for tracking and
`identifying analytes in biological interactions and the reac
`tants and products of chemical reactions.
`HybridiZation Reactions
`For example, it is often desirable to detect or quantify
`very small concentrations of nucleic acids in biological
`samples. Typically, to perform such measurements, the
`nucleic acid in the sample [i.e., the target nucleic acid] is
`hybridiZed to a detection oligonucleotide. In order to obtain
`a detectable signal proportional to the concentration of the
`target nucleic acid, either the target nucleic acid in the
`sample or the detection oligonucleotide is associated with a
`signal generating reporter element, such as a radioactive
`atom, a chromogenic or ?uorogenic molecule, or an enZyme
`[such as alkaline phosphatase] that catalyZes a reaction that
`produces a detectable product. Numerous methods are avail
`able for detecting and quantifying the signal.
`Following hybridiZation of a detection oligonucleotide
`with a target, the resulting signal-generating hybrid mol
`ecules must be separated from unreacted target and detection
`
`25
`
`30
`
`35
`
`40
`
`45
`
`50
`
`55
`
`60
`
`65
`
`2
`oligonucleotides. In order to do so, many of the commonly
`used assays immobiliZe the target nucleic acids or detection
`oligonucleotides on solid supports. Presently available solid
`supports to which oligonucleotides are linked include nitro
`cellulose or nylon membranes, activated agarose supports,
`diaZotiZed cellulose supports and non-porous polystyrene
`latex solid microspheres. Linkage to a solid support permits
`fractionation and subsequent identi?cation of the hybridiZed
`nucleic acids, since the target nucleic acid may be directly
`captured by oligonucleotides immobiliZed on solid supports.
`More frequently, so-called “sandwich” hybridiZation sys
`tems are used. These systems employ a capture oligonucle
`otide covalently or otherwise attached to a solid support for
`capturing detection oligonucleotide-target nucleic acid
`adducts formed in solution [see, eg., EP 276,302 and Gin
`geras et al. (1989) Proc. Natl. Acad. Sci. USA 86:1173].
`Solid supports with linked oligonucleotides are also used in
`methods of affinity puri?cation. Following hybridiZation or
`af?nity puri?cation, however, if identi?cation of the linked
`molecule or biological material is required, the resulting
`complexes or hybrids or compounds must be subjected to
`analyses, such as sequencing.
`Immunoassays
`Immunoassays also detect or quantify very small concen
`trations of analytes in biological samples. Many immunoas
`says utiliZe solid supports in which antigen or antibody is
`covalently, non-covalently, or otherwise, such as via a linker,
`attached to a solid support matrix. The support-bound anti
`gen or antibody is then used as an analyte in the assay. As
`with nucleic acid analysis, the resulting antibody-antigen
`complexes or other complexes, depending upon the format
`used, rely on radiolabels or enZyme labels to detect such
`complexes.
`The use of antibodies to detect and/or quantitate reagents
`[“antigens”] in blood or other body ?uids has been widely
`practiced for many years. Two methods have been most
`broadly adopted. The ?rst such procedure is the competitive
`binding assay, in which conditions of limiting antibody are
`established such that only a fraction [usually 30— 50%] of a
`labeled [e.g., radioisotope, ?uorophore or enZyme] antigen
`can bind to the amount of antibody in the assay medium.
`Under those conditions, the addition of unlabeled antigen
`[eg., in a serum sample to be tested] then competes with the
`labeled antigen for the limiting antibody binding sites and
`reduces the amount of labeled antigen that can bind. The
`degree to which the labeled antigen is able to bind is
`inversely proportional to the amount of unlabeled antigen
`present. By separating the antibody-bound from the
`unbound labeled antigen and then determining the amount of
`labeled reagent present, the amount of unlabeled antigen in
`the sample [e.g., serum] can be determined.
`As an alternative to the competitive binding assay, in the
`labeled antibody, or “immunometric” assay [also known as
`“sandwich” assay], an antigen present in the assay ?uid is
`speci?cally bound to a solid substrate and the amount of
`antigen bound is then detected by a labeled antibody [see,
`eg., Miles et al. (1968) Nature 29:186—189; US. Pat. Nos.
`3,867,517; 4,376,110]. Using monoclonal antibodies two
`site immunometric assays are available [see, eg., US. Pat.
`No. 4,376,110]. The “sandwich” assay has been broadly
`adopted in clinical medicine. With increasing interest in
`“panels” of diagnostic tests, in which a number of different
`antigens in a ?uid are measured, the need to carry out each
`immunoassay separately becomes a serious limitation of
`current quantitative assay technology.
`Some semi-quantitative detection systems have been
`developed [see, eg., Buechler et al. (1992) Clin. Chem.
`
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`

`US 6,372,428 B1
`
`3
`38:1678—1684; and US. Pat. No. 5,089,391] for use With
`immunoassays, but no good technologies yet exist to care
`fully quantitate a large number of analytes simultaneously
`[see, eg., Ekins et al. (1990) J. Clin. Immunoassay
`13:169—181] or to rapidly and conveniently track, identify
`and quantitate detected analytes.
`Combinatorial Libraries
`Drug discovery relies on the ability to identify compounds
`that interact With a selected target, such as cells, an antibody,
`receptor, enZyme, transcription factor or the like. Traditional
`drug discovery involves screening natural products form
`various sources, or random screening of archived synthetic
`material. The current trend, hoWever, is to identify such
`molecules by rational design and/or by screening combina
`torial libraries of molecules.
`Combinatorial chemistry is a poWerful tool in drug dis
`covery and materials science. Methods and strategies for
`generating diverse libraries, primarily peptide- and
`nucleotide-based oligomer libraries, have been developed
`using molecular biology methods and/or simultaneous
`chemical synthesis methodologies [see, e.g., DoWer et al.
`(1991) Annu. Rep. Med. Chem. 26:271—280; Fodor et al.
`(1991) Science 251:767—773; Jung et al. (1992) Angew.
`Chem. Ind. Ed. Engl. 31:367—383; Zuckerman et al. (1992)
`Proc. Natl.Acad. Sci. USA 89:4505—4509; Scott et al. (1990)
`Science 249:386—390; Devlin et al. (1990) Science
`249:404—406; CWirla et al. (1990) Proc. Natl. Acad. Sci.
`USA 87:6378—6382; and Gallop et al. (1994) J. Medicinal
`Chemistry 37:1233—1251]. The resulting combinatorial
`libraries potentially contain millions of pharmaceutically
`relevant compounds and can be rapidly screened to identify
`compounds that exhibit a selected activity.
`The libraries fall into roughly three categories: fusion
`protein-displayed peptide libraries in Which random pep
`tides or proteins are presented on the surface of phage
`particles or proteins expressed from plasmids; support
`bound synthetic chemical libraries in Which individual com
`pounds or mixtures of compounds are presented on insoluble
`matrices, such as resin beads [see, eg., Lam et al. (1991)
`Nature 354:82—84] and cotton supports [see, e.g., Eichler et
`al. (1993) Biochemistry 32:11035—11041]; and methods in
`Which the compounds are used in solution [see, eg.,
`Houghten et al. (1991) Nature 354:84—86, Houghten et al.
`(1992) BioTechniques 313:412—421; and Scott et al. (1994)
`Curr Opin. Bi0techn0l. 5:40—48]. There are numerous
`examples of synthetic peptide and oligonucleotide combi
`natorial libraries. The present direction in this area is to
`produce combinatorial libraries that contain non-peptidic
`small organic molecules. Such libraries are based on either
`a basis set of monomers that can be combined to form
`mixtures of diverse organic molecules or that can be com
`bined to form a library based upon a selected pharmacoph
`ore monomer.
`There are three critical aspects in any combinatorial
`library:
`the chemical units of Which the library is com
`posed; (ii) generation and categoriZation of the library, and
`(iii) identi?cation of library members that interact With the
`target of interest, and tracking intermediary synthesis prod
`ucts and the multitude of molecules in a single vessel.
`The generation of such libraries often relies on the use of
`solid phase synthesis methods, as Well as solution phase
`methods, to produce combinatorial libraries containing tens
`of millions of compounds that can be screened in diagnos
`tically or pharmacologically relevant in vitro assay systems.
`In generating large numbers of diverse molecules by step
`Wise synthesis, the resulting library is a complex mixture in
`
`10
`
`15
`
`25
`
`35
`
`45
`
`55
`
`65
`
`4
`Which a particular compound is present at very loW
`concentrations, so that it is difficult or impossible to deter
`mine its chemical structure. Various methods exist for
`ordered synthesis by sequential addition of particular
`moieties, or by identifying molecules based on spacial
`positioning on a chip. These methods are cumbersome and
`ultimately impossible to apply to highly diverse and large
`libraries.
`Thus, an essential element of the combinatorial discovery
`process, as Well as other areas in Which molecules are
`identi?ed and tracked, is the ability to extract the informa
`tion made available during synthesis of the library or iden
`ti?cation of the active components of intermediary struc
`tures. While there are several techniques for identi?cation of
`intermediary products and ?nal products, nanosequencing
`protocols that provide exact structures are only applicable on
`mass to naturally occurring linear oligomers such as pep
`tides and amino acids. Mass spectrographic [MS] analysis is
`suf?ciently sensitive to determine the exact mass and frag
`mentation patterns of individual synthesis steps, but com
`plex analytical mass spectrographic strategies are not readily
`automated nor conveniently performed. Also, mass spectro
`graphic analysis provides at best simple connectivity
`information, but no stereoisomeric information, and gener
`ally cannot discriminate among isomeric monomers.
`Another problem With mass spectrographic analysis is that it
`requires pure compounds; structural determinations on com
`plex mixtures is either dif?cult or impossible. Finally, mass
`spectrographic analysis is tedious and time consuming.
`Thus, although there are a multitude of solutions to the
`generation of libraries, there are no ideal solutions to the
`problems of identi?cation, tracking and categoriZation.
`Similar problems arise in any screening or analytical
`process in Which large numbers of molecules or biological
`entities are screened. In any system, once a desired molecule
`(s) has been isolated, it must be identi?ed. Simple means for
`identi?cation do not exist. Because of the problems inherent
`in any labeling procedure, it Would be desirable to have
`alternative means for tracking and quantitating chemical and
`biological reactions during synthesis and/or screening pro
`cesses.
`Therefore, it is an object herein to provide methods for
`identi?cation, tracking and categoriZation of the components
`of complex mixtures of diverse molecules.
`
`SUMMARY OF THE INVENTION
`Combinations of matrix materials With programmable
`data storage or recording devices, herein referred to as
`memories and assays using these combinations are provided.
`By virtue of this combination, molecules, such as antigens,
`antibodies, ligands, proteins and nucleic acids, and biologi
`cal particles, such as phage and viral particles and cells, that
`are in proximity to or in physical contact With the matrix
`combination can be electromagnetically tagged by program
`ming the memory With data corresponding to identifying
`information. In addition, assays can be monitored, because
`occurrence of a reaction can be recorded in the memory, if
`desired, and/or continually monitored in real time. Also, an
`identi?cation code for the particular memory/matrix com
`bination can be permanently recorded, thereby providing a
`means to identify each such encoded combination. The
`molecules and biological particles that are in proximity to or
`in physical contact With the matrix combination can be
`identi?ed and the results of the assays determined by retriev
`ing the stored data points from the memories.
`Combinations of matrix materials, memories, and linked
`or proximate molecules and biological materials and assays
`
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`US 6,372,428 B1
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`using such combinations are also provided. The combina
`tions provided herein have a multiplicity of applications,
`including combinatorial chemistry, isolation and puri?cation
`of target macromolecules, capture and detection of macro
`molecules for analytical purposes, high throughput
`screening, selective removal of contaminants, enZymatic
`catalysis, drug delivery, chemical modi?cation, drug
`delivery, information collection and management and other
`uses. These combinations are particularly advantageous for
`use in multianalyte analyses and assays in Which a electro
`magnetic signal is generated by the reactants or products in
`the assay.
`The combinations are referred to herein as matrices With
`memories. These combinations contain
`a miniature
`recording device that contains one or more programmable
`data storage devices [memories] that can be remotely read
`and in preferred embodiments also remotely programmed;
`and (ii) a matrix, such as a particulate support used in
`chemical syntheses. The remote programming and reading is
`preferably effected using electromagnetic radiation.
`The matrix materials [matrices] are any materials that are
`routinely used in chemical and biochemical synthesis. The
`matrix materials are typically polymeric materials that are
`compatible With chemical and biological syntheses and
`assays, and include, glasses, silicates, celluloses,
`polystyrenes, polysaccharides, polypropylenes, sand, and
`synthetic resins and polymers, including acrylamides, par
`ticularly cross-linked polymers, cotton, and other such mate
`rials. The matrices may be in the form of particles or may be
`continuous in design, such as a test tube or microtiter plate
`or the like.
`The recording device is a miniature device, typically less
`than 10—20 mm3 [or 10—20 mm in its largest dimension] in
`siZe , preferably smaller, that includes at least one data
`storage unit that includes a remotely programmable and
`remotely readable, preferably non-volatile, memory. This
`device With remotely programmable memory is in proximity
`With or in contact With the matrix. In particular, the record
`ing device includes a memory device, preferably having
`non-volatile memory means, for storing a plurality of data
`points and means for receiving a transmitted signal that is
`received by the device and for causing a data point corre
`sponding to the data signal to be permanently stored Within
`the memory means; and, if needed, a shell that is non
`reactive With and impervious to any processing steps or
`solutions in Which the combination of matrix With recording
`device is placed, and that is transmissive of read or Write
`signals transmitted to the memory. The device may also
`include at least one support matrix disposed on an outer
`surface of the shell for retaining molecules or biological
`particles.
`The recording device [containing the memory] is typically
`coated With at least one layer of material, such as a protec
`tive polymer or a glass, including polystyrene, heavy metal
`free glass, plastic, ceramic, and may be coated With more
`than one layers of this and other materials. For example, it
`may be coated With a ceramic or glass, Which is then coated
`With or linked to the matrix material. Alternatively, the glass
`or ceramic or other coating may serve as the matrix.
`In other embodiments the recording device and the matrix
`material are in proximity, such as in a container of a siZe
`approximately that of the device and matrix material. In
`preferred embodiments in Which the device is a semicon
`ductor that is approximately 10 mm or less and the matrix
`material is a bead or the like, such as a polystyrene bead, the
`device and beads, about 1 mg to about 50 mg, are sealed in
`
`10
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`15
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`25
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`35
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`45
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`55
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`a chemically inert porous supports, such as polypropylene
`formed so that it has pores of a selected siZe.
`The data storage device or memory is programmed With
`or encoded With information that identi?es molecules or
`biological particles, either by their process of preparation,
`their identity, their batch number, category, physical or
`chemical properties, combinations of any of such
`information, or other such identifying information. The
`molecules or biological particles are in physical contact,
`direct or indirect, or in proximity With the matrix, Which in
`turn is in physical contact or in the proximity of the
`recording device that contains the data storage memory.
`Typically, the matrix is on the surface of the recording
`device and the molecules and biological particles are in
`physical contact With the matrix material.
`The data storage device or memory can also be pro
`grammed by virtue of a reaction in proximity to the matrix
`With memory. In particular, memories that also include a
`photodectector can detect the occurrence of ?uorescence or
`other optical emission. Coupling this emission With an
`ampli?er and providing a voltage to permit data storage in
`the matrix With memory during the reaction by Way of, for
`example an RF signal transmitted to and received by an
`antenna/recti?er combination Within the data storage device
`or providing voltage suf?cient to Write to memory from a
`battery [see, e.g., US. Pat. Nos. 5,350,645 and 5,089,877],
`permits occurrence of the emission to be recorded in the
`memory.
`The matrix combinations, thus, contain a matrix material,
`typically in particulate form, in physical contact With a tiny
`device containing one or more remotely programmable data
`storage units [memories]. Contact can be effected by placing
`the recording device With memory on or in the matrix
`material or in a solution that is in contact With the matrix
`material or by linking the device, either by direct or indirect
`covalent or non-covalent interactions, chemical linkages or
`by other interactions, to the matrix.
`For example, such contact is effected chemically, by
`chemically coupling the device With data storage unit to the
`matrix, or physically by coating the recording device With
`the matrix material or another material, by physically insert
`ing or encasing the device in the matrix material, by placing
`the device onto the matrix or by any other means by Which
`the device can be placed in contact With or in proximity to
`the matrix material.
`Thus, combinations of a miniature recording device that
`contains or is a data storage unit linked to or in proximity
`With matrices or supports used in chemical and biotechnical
`applications, such as combinatorial chemistry, peptide
`synthesis, nucleic acid synthesis, nucleic acid ampli?cation
`methods, organic template chemistry, nucleic acid
`sequencing, screening for drugs, particularly high through
`put screening, phage display screening, cell sorting, drug
`delivery, tracking of biological particles and other such
`methods, are provided. These combinations of matrix mate
`rial With data storage unit [or recording device including the
`unit] are herein referred to as matrices With memories.
`The matrices are either particulate of a siZe that is roughly
`about 10 to 20 mm3 [or 10—20 mm in its largest dimension],
`preferably about 10 mm3 or smaller, preferably 1 mm3 or
`smaller, or a continuous medium, such as a microtiter plate,
`or other multi-Well plate, or plastic or other solid polymeric
`vial or glass vial or catheter-tube [for drug delivery] or such
`container or device conventionally used in chemistry and
`biological syntheses and reactions. In instances in Which the
`matrix is continuous, the data storage device [memory] may
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`US 6,372,428 B1
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`7
`be placed in, on, or under the matrix medium or may be
`embedded in the material of the matrix.
`More than one data storage device may be in proximity to
`or contact With a matrix particle, or more than one matrix
`particle may be in contact With on device. For example,
`microtiter plates With the recording device containing the
`data storage unit (remotely programmable memory) embed
`ded in each Well or vials (typically With a 1 ml or smaller
`capacity) With an embedded recording device, may be
`manufactured. In other embodiments, the memory device
`may be linked to or in proximity to more than one matrix
`particle. For example, a single device and a plurality of
`particles may be sealed in a porous or semi-permeable inert
`material such as Te?on® or polypropylene or membrane that
`is permeable to the components of the medium, or they may
`be contained in a small closable container that has at least
`one dimension that is a porous or semi-permeable tube.
`Typically such a microvessel, called a “micro-can”, Which
`preferably has an end that can be opened and sealed or
`closed tightly, has a volume of about 200—500 mm3, With
`preferred dimensions of about 1—10 mm in diameter and 5
`to 20 mm in height, more preferably about 5 mm by 15 mm.
`The porous Wall should be non-collapsible With a pore siZe
`in the range of 70 pM to about 100 pM, but can be selected
`to be semi-permeable for selected components of the
`medium in Which the microvessel “micro-can” is placed.
`The preferred geometry of these combinations is cylindrical.
`These porous micro-cans may be sealed by heat or may be
`designed to snap or otherWise close. In some embodiments
`they are designed to be reused. In other embodiments, the
`microvessel “micro-can” With closures may be made out of
`non-porous material, such as a tube of the shape of an
`EppendorfTM tube, and used as container to hold the matrix
`particles and device.
`The combination of matrix With memory is used by
`contacting it With, linking it to, or placing it in proximity
`With a molecule or biological particle, such as a virus or
`phage particle, a bacterium or a cell, to produce a second
`combination of a matrix With memory and a molecule or
`biological particle. In certain instances, such combinations
`of matrix With memory or combination of matrix With
`memory and molecule or biological particle may be pre
`pared When used or may be prepared before use and pack
`aged or stored as such for futures use.
`Since matrix materials have many knoWn uses in con
`junction With molecules and biological particles, there are a
`multitude of methods knoWn to artisans of skill in this art for
`linking, joining or physically contacting the molecule or
`biological particle With the matrix material. In