Brush polymer macromolecular structures and methods for detecting biomolecules

Abstract

Provided herein are macromolecular structures comprising a surface, tethered moiety, and a macromolecular chain, compositions thereof, and methods of making the same. Also provided herein are systems and kits comprising the macromolecular structures.

Description

Cross-references

[0001] This application claims the benefit of U.S. Provisional Application No. 63 / 517,515, filed August 3, 2023, which is incorporated herein by reference in its entirety. Background Technology

[0002] The analysis and separation of biomolecules often require laborious methods and rely on fine-tuning of analytical parameters. Typically, these methods require induced polymerization, which may be infeasible for all proteins or ligands and is often irreversible. There is a need for improved systems for the separation and subsequent analysis of biomolecules. This disclosure provides macromolecular structures, compositions thereof, methods, and systems to address this need. Summary of the Invention

[0003] This article provides a macromolecular structure, which comprises: (I) Surface; (II) The tethered portion coupled to the surface; and (III) A macromolecular chain, wherein the first end of the macromolecular chain is covalently attached to the tethered portion, and wherein the macromolecular chain comprises two or more distinct repeating units derived from monomers selected from the following structures: , , , , , , , and

[0004] Each of X and Y is independently -C-, -O-, or -N-; Each Z is independently -O- or -NH; Q is -CH2- or ethylene glycol; A is a polymer side chain comprising repeating units derived from monomers represented by the following structures: ; m is an integer selected from 1 to 20; Is it a single bond or a double bond? R 1 R 2 R 1’ R 2’ and R 3’ Each of them is independently selected from hydrogen or -C1-C6 alkyl; R 3It is a hydrogen, a C1-C6 alkyl group, or a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl groups; R 4 It is absent, hydrogen, sulfonate, carboxylic acid ester, C1-C4 alkylene, amine, quaternary ammonium cation or C1-C6 alkyl optionally substituted with halogen; R 5 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, an amine, an azide, a sulfonate, a carbamate, an asymmetric disulfide, or a C1-C8 alkyl group or a C1-C6 heterocyclic group substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines optionally further substituted with amine, hydroxyl, aryl or sulfonate, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogens, C1-C8 alkyl groups optionally substituted with one or more pyrene, two or more fused 5-6 membered rings optionally further substituted, -C1-C3 alkyl groups optionally substituted with benzyl, trimethoxysilane or phosphocholine, C1-C 12 Alkylamines or C1-C4 alkylamines; R 6 It is hydrogen or C1-C6 alkyl. R 7 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, amines, azides, sulfonates, carbamates, or asymmetric disulfides, or a 3-, 5-, or 6-membered heterocyclic or C1-C alkyl group optionally substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines optionally further substituted with amines or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogens, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted, optionally substituted benzyl groups, -C1-C3 alkyl or C1-C4 alkylyl groups substituted with trimethoxysilane or phosphocholine; R 8 It is a C1-C6 alkyl group, a divalent metal, or a symmetrical or asymmetrical disulfide; R 9 It is hydrogen or oxo; and n 1 These are integers selected from 1 to 100. This article also provides the value of R. 3 It is a macromolecular structure of methyl groups. This article also provides the R group within it. 5 It is C1-C n1 The macromolecular structure of ethylene glycol.

[0005] This article provides a macromolecular structure, which comprises: (I) Surface; (II) The tethered portion coupled to the surface; and (III) A macromolecular chain, wherein the first end of the macromolecular chain is covalently attached to the tethered portion, and wherein the macromolecular chain comprises repeating units derived from monomers selected from the following structures: , , , , , , , and

[0006] Each of X and Y is independently -C-, -O-, or -N-; Each Z is independently -O- or -NH; Q is -CH2- or ethylene glycol; A is a polymer side chain comprising repeating units derived from monomers represented by the following structures: ; m is 1-6; R 1 R 2 R 1’ R 2’ and R 3’ Each of them is independently selected from hydrogen or C1-C6 alkyl. R 3 It is a hydrogen, a C1-C6 alkyl group, or a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl groups; R 4 It is absent, hydrogen, or optionally halogenated C1-C6 alkyl, C1-C4 alkylene, C1-C6 alkyl, sulfonate, amine, quaternary ammonium cation or carboxylic acid ester; R 5 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, an amine, an azide, a sulfonate, a carbamate, an asymmetric disulfide, or a C1-C8 alkyl group or a C1-C6 heterocyclic group substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines further substituted with amines, hydroxyl groups, aryl groups, or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogen groups, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted (e.g., two or more fused 6 membered rings optionally further substituted), optionally substituted benzyl groups, trimethoxysilanes, or phosphoric acid choline-substituted -C1-C3 alkyl groups, C1-C 12Alkylamines or C1-C4 alkylamines; R 6 It is hydrogen or a linear C1-C6 alkyl group. R 7 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, amines, azides, sulfonates, carbamates, or asymmetric disulfides, or a 3-, 5-, or 6-membered heterocyclic or C1-C alkyl group optionally substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines further substituted with amines or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogen groups, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted (e.g., two or more fused 6 membered rings optionally further substituted), optionally substituted benzyl groups, -C1-C3 alkyl or C1-C4 alkylyl groups substituted with trimethoxysilane or phosphocholine; and R 8 It is a C1-C6 alkyl or symmetrical disulfide or asymmetrical disulfide; R 9 It is hydrogen or oxygen; n 1 It is an integer selected from 1 to 100; and The condition is when R 3 It is CH3, R 4 When it is CH3 or when R 5 When the C1-C8 alkyl group is substituted with a hydroxyl group, the C1-C8 groups are further substituted.

[0007] This document provides a macromolecular structure comprising: (I) a surface and (II) macromolecular chains coupled to the surface, wherein the macromolecular chains comprise repeating units of formula (I): (I) Where R 1” R 2” and R 3” Each of them is independently hydrogen or a C1-C6 alkyl group; L represents the connector section; A is a polymer side chain comprising repeating units derived from monomers selected from the following structures: , , , , , , and

[0008] Each of X and Y is independently -C-, -O-, or -N-; Z is -O- or -NH; R 1 R 2 R 1’ R 2’ and R 3’ Each of them is independently selected from hydrogen or C1-C6 alkyl; R 3 It is a hydrogen, a C1-C6 alkyl group, or a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl groups; R 4 It is absent, hydrogen, sulfonate, carboxylic acid ester, C1-C4 alkylene, amine, quaternary ammonium cation or C1-C6 alkyl optionally substituted with halogen; R 5 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, an amine, an azide, a sulfonate, a carbamate, an asymmetric disulfide, or a C1-C8 alkyl group or a C1-C6 heterocyclic group substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines further substituted with amines, hydroxyl groups, aryl groups, or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogen groups, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted (e.g., two or more fused 6 membered rings optionally further substituted), optionally substituted benzyl groups, trimethoxysilanes, or phosphoric acid choline-substituted -C1-C3 alkyl groups, C1-C 12 Alkylamines or C1-C4 alkylamines; R 6 It is hydrogen or a linear C1-C6 alkyl group. R 7 It is a hydrogen, C1-C6 alkyl, a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl or oxo compounds, a substituted benzene, a C1-C6 alkyl substituted with a hydroxyl group, or a C1-C8 alkyl sulfonate optionally substituted. R 8 It is a C1-C6 alkyl group, a divalent metal, or a symmetrical or asymmetrical disulfide; R 9 It is hydrogen or oxygen; n 1 It is an integer selected from 1 to 100; and n is an integer selected from 1 to 10,000.

[0009] This article also provides macromolecular structures, wherein the macromolecular chains comprise repeating units derived from monomers, represented by the following structures: This document also provides macromolecular structures wherein the macromolecular chains comprise monomer-derived repeating units represented by the structures in Table 1. This document also provides macromolecular structures wherein the macromolecular chains comprise monomer-derived repeating units represented by the structures in Table 2. This document also provides macromolecular structures having structures represented by the following: , , , , , or .

[0010] This document provides a method for preparing the macromolecular structure described herein, the method comprising: providing a surface; coupling a polymeric initiator to the surface to form an initiator surface; and contacting the initiator surface with a monomer described herein to form the macromolecular structure. This document also provides a method wherein the surface comprises particles. This document further provides a method wherein the particles have a diameter of about 100 nm to about 500 nm.

[0011] This document provides a method for preparing the macromolecular structure described herein, the method comprising: providing a surface; coupling a vinyl group to the surface to form a vinyl-functionalized surface; contacting the vinyl-functionalized surface with a crosslinking monomer and a monomer selected from hydroxyalkyl methacrylate, aminoalkyl methacrylate, alkynyl methacrylate, glycidyl methacrylate, hydroxyalkyl acrylate, aminoalkyl acrylate, alkynyl acrylate, or glycidyl acrylate to form a crosslinked polymer coupled to the surface; coupling a polymer initiator to the crosslinked polymer to form an initiator surface; and contacting the initiator surface with a monomer described herein to form the macromolecular structure. This document also provides a method wherein the polymer initiator is represented by the following structure: Where: X is a halogen; R 10 It is an initiator group.

[0012] This document provides compositions comprising the macromolecular structure described herein and biomolecules adsorbed onto said macromolecular structure. It also provides compositions wherein at least 100 different biomolecules are adsorbed onto said macromolecular structure.

[0013] This document provides a method for identifying proteins in a sample, the method comprising: incubating one or more macromolecular structures described herein with a biological sample containing biomolecules to form a biomolecular crown; isolating at least a portion of the biomolecules in the biomolecular crown; and measuring the biomolecular crown. This document also provides a method wherein the measurement is capable of identifying 1 to 20,000 protein groups.

[0014] This article provides a kit for identifying molecules in biological samples, the kit containing one or more macromolecular structures provided herein.

[0015] This document provides a system for identifying biomolecules in biological samples, the system comprising: a macromolecular structure provided herein; a suspension; a biological sample containing a certain concentration of protein; and an automated system comprising a network of differentiated functional units for isolating biomolecules adsorbed to the macromolecular structure, wherein the automated system is programmed to perform a series of steps. Attached Figure Description

[0016] The novel features of this disclosure are set forth in the appended claims. The features and advantages of this disclosure can be better understood by referring to the following detailed description and accompanying drawings (also referred to herein as “Figures” and “FIG.”), in which illustrative embodiments utilizing the principles of this disclosure are set forth, and in which: Figure 1 An exemplary synthetic scheme for preparing the macromolecular structure described herein is shown by contacting the monomer with the surface of the initiator.

[0017] Figure 2 An exemplary synthetic scheme is shown to prepare the macromolecular structure described herein by contacting a vinyl-functionalized surface with a crosslinking monomer and a second monomer.

[0018] Figure 3A This paper presents scanning electron micrographs of the macromolecular structures described in this paper. Figure 3B Transmission electron micrographs of the macromolecular structures described in this paper are shown.

[0019] Figure 4 The synthetic scheme for preparing compound 1 is shown.

[0020] Figure 5 The synthetic scheme for preparing compound 2 is shown.

[0021] Figure 6 The synthetic scheme for preparing compound 3 is shown.

[0022] Figure 7 The synthetic scheme for preparing block copolymers is shown.

[0023] Figure 8 Synthetic schemes for preparing macromolecular chains containing different monomers are shown. Detailed Implementation

[0024] Some definitions As used herein and in the appended claims, unless the context clearly specifies otherwise, the singular forms “a,” “and,” and “the” include plural references. Thus, for example, reference to “an agent” includes multiple such agents, and reference to “the cell” includes reference to one or more cells and their equivalents known to those skilled in the art. When the scope is used herein to refer to physical properties (such as molecular weight) or chemical properties (such as chemical formula), it is intended to include all combinations and sub-combinations of the scope and the scope in the particular embodiment. When referring to a numerical value or a numerical range, the term “about” means an approximation of the mentioned numerical value or range within experimental variability (or statistical experimental error), and therefore the numerical value or range may vary between 1% and 15% of the stated numerical value or range. The term “comprising” (and related terms such as “comprise”, “comprises”, “having”, or “including”) is not intended to exclude certain other embodiments, for example, embodiments of any material composition, composition, method, or process described herein may be “composed” or “substantially composed” of the described features.

[0025] "Amino" refers to the -NH2 group.

[0026] "Cyano" refers to the -CN group.

[0027] "Nitro" refers to the -NO2 group.

[0028] "Oxo" refers to the =O group.

[0029] "Hydroxy group" refers to the -OH group.

[0030] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure pertains. All patents and publications cited herein are incorporated herein by reference.

[0031] "alkyl" refers to a monovalent group of a straight-chain or branched saturated hydrocarbon with optional substitution, and preferably has one to fifteen carbon atoms (i.e., C1-C1). 15 Alkyl group). In some embodiments, the alkyl group comprises one to thirteen carbon atoms (i.e., C1-C1). 13Alkyl group. In some embodiments, the alkyl group comprises one to eight carbon atoms (i.e., C1-C8 alkyl). In other embodiments, the alkyl group comprises one to five carbon atoms (i.e., C1-C5 alkyl). In other embodiments, the alkyl group comprises one to four carbon atoms (i.e., C1-C4 alkyl). In other embodiments, the alkyl group comprises one to three carbon atoms (i.e., C1-C3 alkyl). In other embodiments, the alkyl group comprises one to two carbon atoms (i.e., C1-C2 alkyl). Whenever appearing herein, numerical ranges such as "C1-C3 alkyl" mean that the alkyl group consists of 1, 2, or 3 carbon atoms. In other embodiments, the alkyl group comprises one carbon atom (i.e., C1 alkyl). In other embodiments, the alkyl group comprises five to fifteen carbon atoms (i.e., C5-C6 alkyl). 15 Alkyl group. In other embodiments, the alkyl group comprises five to eight carbon atoms (i.e., C5-C8 alkyl). In other embodiments, the alkyl group comprises two to five carbon atoms (i.e., C2-C5 alkyl). In other embodiments, the alkyl group comprises three to five carbon atoms (i.e., C3-C5 alkyl). In some embodiments, the alkyl group is selected from methyl, ethyl, 1-propyl (n-propyl), 1-methylethyl (isopropyl), 1-butyl (n-butyl), 1-methylpropyl (sec-butyl), 2-methylpropyl (isobutyl), 1,1-dimethylethyl (tert-butyl), and 1-pentyl (n-pentyl). In other embodiments, examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, 2-methyl-1-propyl, 2-methyl-2-propyl, 2-methyl-1-butyl, 3-methyl-1-butyl, 2-methyl-3-butyl, 2,2-dimethyl-1-propyl, 2-methyl-1-pentyl, 3-methyl-1-pentyl, 4-methyl-1-pentyl, 2-methyl-2-pentyl, 3-methyl-2-pentyl, 4-methyl-2-pentyl, 2,2-dimethyl-1-butyl, 3,3-dimethyl-1-butyl, 2-ethyl-1-butyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, tert-pentyl, and hexyl, as well as longer alkyl groups such as heptyl, octyl, etc. The alkyl group is connected to the rest of the molecule by a single bond. Unless otherwise expressly stated in this specification, the alkyl group is optionally substituted with, for example, oxo, halogen, amino, nitrile, nitro, hydroxyl, sulfone, mercapto, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, the alkyl group is optionally substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, -NO2, or -C≡CH. In some embodiments, the alkyl group is optionally substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, the alkyl group is optionally substituted with a halogen (such as F). In some embodiments, the alkyl group is not substituted.

[0032] As used in this article, C1-Cx (or C) 1-x (Including C1-C2, C1-C3...C1-C) x For example, a group designated "C1-C4" indicates that the moiety contains one to four carbon atoms, i.e., a group containing 1, 2, 3, or 4 carbon atoms. Therefore, for example, "C1-C4 alkyl" indicates that the alkyl group contains one to four carbon atoms, i.e., the alkyl group is selected from methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, sec-butyl, and tert-butyl. Additionally, for example, C0-C2 alkylene groups include direct bonds, -CH2-, and -CH2CH2- linkages.

[0033] "Alkoxy" refers to a group of the formula -O-alkyl bonded by an oxygen atom, wherein the alkyl group is an alkyl chain as defined above. Unless otherwise expressly stated in this specification, the alkoxy group may optionally be substituted with, for example, oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, the alkoxy group is optionally substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, the alkoxy group is optionally substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, the alkoxy group is optionally substituted with a halogen. In some embodiments, the alkoxy group is unsubstituted.

[0034] "Alkenyl" refers to an optionally substituted straight-chain or branched hydrocarbon chain group containing at least one carbon-carbon double bond, and preferably having two to twelve carbon atoms (i.e., C2-C). 12Alkenyl group. In some embodiments, the alkenyl group comprises two to eight carbon atoms (i.e., C2-C8 alkenyl). In some embodiments, the alkenyl group comprises four to eight carbon atoms (i.e., C4-C6 alkenyl). In other embodiments, the alkenyl group comprises six to eight carbon atoms (i.e., C6-C8 alkenyl). In some embodiments, the alkenyl group comprises at least one double bond at the end of the carbon chain. In other embodiments, the alkenyl group comprises at least one double bond in the middle of the carbon chain. The group can be in a cis or trans configuration around the double bond and should be understood to include both isomers. Examples include, but are not limited to, vinyl (-CH=CH2), 1-propenyl (-CH2CH=CH2), isopropenyl [-C(CH3)=CH2], butenyl, 1,3-butadienyl, etc. Whenever it appears herein, numerical ranges such as "C2-C6 alkenyl" mean that the alkenyl group can consist of 2, 3, 4, 5, or 6 carbon atoms. Unless otherwise expressly stated in this specification, the alkenyl group is optionally substituted with, for example, oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, the alkenyl group is optionally substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, the alkenyl group is optionally substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, the alkenyl group is optionally substituted with halogen. The alkenyl group is connected to the rest of the molecule by a single bond, such as ethenyl, propenyl (i.e., allyl), butenyl, pentenyl, pent-1,4-dienyl, etc. Unless otherwise expressly stated in this specification, the alkenyl group is optionally substituted with, for example, oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, the alkenyl group is optionally substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, the alkenyl group is optionally substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, the alkenyl group is optionally substituted with halogen. In some embodiments, the alkenyl group is unsubstituted.

[0035] "Alkyne" refers to an optionally substituted straight-chain or branched hydrocarbon chain group containing at least one carbon-carbon triple bond, and preferably having two to twelve carbon atoms (i.e., C2-C). 12The alkynyl group comprises two to eight carbon atoms (i.e., C2-C8 alkynyl). In other embodiments, the alkynyl group comprises two to six carbon atoms (i.e., C2-C6 alkynyl). In still other embodiments, the alkynyl group comprises two to four carbon atoms (i.e., C2-C4 alkynyl). Whenever appearing herein, numerical ranges such as "C2-C6 alkynyl" mean that the alkynyl group can consist of 2, 3, 4, 5, or 6 carbon atoms. The alkynyl group is connected to the rest of the molecule by a single bond, such as ethynyl, propynyl, butynyl, pentyynyl, hexynyl, 2-propynyl, 2-butynyl, 1,3-butydynyl, etc. Unless otherwise expressly stated in this specification, the alkynyl group may optionally be substituted with, for example, oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, the alkynyl group is optionally substituted with an oxo group, a halogen group, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, the alkynyl group is optionally substituted with an oxo group, a halogen group, -CN, -CF3, -OH, or -OMe. In some embodiments, the alkynyl group is optionally substituted with a halogen. In some embodiments, the alkynyl group is unsubstituted.

[0036] "alkylene" or "alkylene chain" refers to an optionally substituted straight-chain or branched divalent hydrocarbon chain that links the remainder of the molecule to a group containing an unsaturated group, and preferably has one to twelve carbon atoms, such as methylene, ethylene, propylene, n-butylene, etc. The alkylene chain is linked to the remainder of the molecule by single bonds and to the group by single bonds. The connection points between the alkylene chain and the remainder of the molecule, as well as with the group, can be any two carbon atoms within the chain. In some embodiments, the alkylene comprises one to ten carbon atoms (i.e., C1-C8 alkylene). In some embodiments, the alkylene comprises one to eight carbon atoms (i.e., C1-C8 alkylene). In other embodiments, the alkylene comprises one to five carbon atoms (i.e., C1-C5 alkylene). In other embodiments, the alkylene comprises one to four carbon atoms (i.e., C1-C4 alkylene). In other embodiments, the alkylene comprises one to three carbon atoms (i.e., C1-C3 alkylene). In other embodiments, the alkylene group comprises one or two carbon atoms (i.e., C1-C2 alkylene). In other embodiments, the alkylene group comprises one carbon atom (i.e., C1 alkylene). In other embodiments, the alkylene group comprises five to eight carbon atoms (i.e., C5-C8 alkylene). In other embodiments, the alkylene group comprises two to five carbon atoms (i.e., C2-C5 alkylene). In other embodiments, the alkylene group comprises three to five carbon atoms (i.e., C3-C5 alkylene). Unless otherwise expressly stated in this specification, the alkylene group may optionally be substituted with, for example, oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, the alkylene group may optionally be substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, the alkylene group may optionally be substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, the alkylene group is optionally substituted with a halogen. In some embodiments, the alkylene group is -CH2-, -CH2CH2-, or -CH2CH2CH2-. In some embodiments, the alkylene group is -CH2-. In some embodiments, the alkylene group is -CH2CH2-. In some embodiments, the alkylene group is -CH2CH2CH2-. In some embodiments, the alkylene group is unsubstituted.

[0037] "Aryl" refers to a group derived from a hydrocarbon ring system containing at least one aromatic ring. In some embodiments, the aryl group contains hydrogen and 5 to 30 carbon atoms. The aryl group can be a monocyclic, bicyclic, tricyclic, or tetracyclic system, which can include fused ring systems (where the aryl group is bonded through aromatic ring atoms when fused with a cycloalkyl or heterocyclic alkyl ring) or bridged ring systems. In some embodiments, the aryl group is a 6- to 10-membered aryl group. In some embodiments, the aryl group is a 6-membered aryl group. Aryl groups include, but are not limited to, aryl groups derived from hydrocarbon ring systems of anthracene, naphthyl, phenanthrene, anthracene, azulene, benzene, benzo[a], fluorene, indene, indene, naphthalene, phenanthrene, pleiadene, pyrene, and benzo[a]phenanthrene. In some embodiments, the aryl group is phenyl. Unless otherwise expressly stated in this specification, the aryl group may optionally be substituted with, for example, halogen, amino, alkylamino, aminoalkyl, nitrile, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, heteroalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, -S(O)2NH-C1-C6 alkyl, etc. In some embodiments, the aryl group may optionally be substituted with halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, -NO2, S(O)2NH2, -S(O)2NHCH3, -S(O)2NHCH2CH3, -S(O)2NHCH ( -CH3)2, -S(O)2N(CH3)2, or -S(O)2NHC(CH3)3 are substituted. In some embodiments, the aryl group is optionally substituted with a halogen, methyl, ethyl, -CN, -CF3, -OH, or -OMe. In some embodiments, the aryl group is optionally substituted with a halogen. In some embodiments, the aryl group is substituted with an alkyl, alkenyl, alkynyl, haloalkyl, or heteroalkyl group, wherein each alkyl, alkenyl, alkynyl, haloalkyl, or heteroalkyl group is independently unsubstituted, or substituted with a halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, the aryl group is unsubstituted.

[0038] "Aryl group" refers to the formula -R c -aryl groups, where R c It is an alkylene chain as defined above, such as methylene, ethylene, etc.

[0039] "Aryl" refers to the formula -R d -aryl groups, where R d It is an alkenyl chain as defined above. "Arynyl group" refers to the formula -R e -aryl groups, where R e It is an alkyne chain as defined above.

[0040] "Carbocyclic ring" refers to a saturated, unsaturated, or aromatic ring in which each atom of the ring is carbon. Carbocyclic rings can include 3- to 10-membered monocyclic rings and 6- to 12-membered bicyclic rings (such as spirocyclic, fused, or bridged rings). Each ring of a bicyclic carbocyclic ring can be selected from saturated, unsaturated, and aromatic rings. An aromatic ring (e.g., phenyl) may be fused with a saturated or unsaturated ring (e.g., cyclohexane, cyclopentane, or cyclohexene). Any combination of saturated, unsaturated, and aromatic bicyclic rings is included in the definition of a carbocyclic ring, where valence allows. In an exemplary embodiment, an aromatic ring (e.g., phenyl) may be fused with a saturated or unsaturated ring (e.g., cyclohexane, cyclopentane, or cyclohexene). Bicyclic carbocyclic rings include any combination of saturated, unsaturated, and aromatic bicyclic rings, where valence allows. Bicyclic carbocyclic rings include any combination of ring sizes, such as 4-5 fused ring systems, 5-5 fused ring systems, 5-6 fused ring systems, 6-6 fused ring systems, 5-7 fused ring systems, 6-5 fused ring systems, 6-7 fused ring systems, 5-8 fused ring systems, and 6-8 fused ring systems. Exemplary carbocyclic rings include cyclopentyl, cyclohexyl, cyclohexenyl, adamantyl, phenyl, indanyl, and naphthyl. The term "unsaturated carbocyclic ring" refers to a carbocyclic ring having at least one degree of unsaturation and does not include aromatic carbocyclic rings. Examples of unsaturated carbocyclic rings include cyclohexadiene, cyclohexene, and cyclopentene. The term "saturated cycloalkyl" as used herein refers to a saturated carbocyclic ring. Exemplary carbocyclic rings include cyclopropyl, cyclopentyl, cyclohexyl, cyclohexenyl, adamantyl, phenyl, indanyl, norbornyl, and naphthyl. Carbocyclic rings may optionally be substituted with one or more substituents (such as those described herein).

[0041] "Cycloalkyl" refers to a stable, partially or fully saturated, monocyclic or polycyclic carbocyclic ring, which can include fused ring systems (where the cycloalkyl group is bonded through non-aromatic ring atoms when fused with an aryl or heteroaryl ring), bridged ring systems, or spirocyclic systems. Representative cycloalkyl groups include, but are not limited to, cycloalkyl groups having three to fifteen carbon atoms (C3-C4). 15 cycloalkyl groups, having three to ten carbon atoms (C3-C4). 10Cycloalkyl groups are cyclic alkyl groups having three to eight carbon atoms (C3-C8 cycloalkyl), three to six carbon atoms (C3-C6 cycloalkyl), three to five carbon atoms (C3-C5 cycloalkyl), or three to four carbon atoms (C3-C4 cycloalkyl). In some embodiments, the cycloalkyl group is a 3- to 6-membered cycloalkyl group. In some embodiments, the cycloalkyl group is a 5- to 6-membered cycloalkyl group. Monocyclic cycloalkyl groups include, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. Polycyclic cycloalkyl or carbocyclic compounds include, for example, adamantyl, norbornel, decahydronaphthyl, bicyclo[3.3.0]octane, bicyclo[4.3.0]nonane, cis-decahydronaphthyl, trans-decahydronaphthyl, bicyclo[2.1.1]hexane, bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, bicyclo[3.2.2]nonane, bicyclo[3.3.2]decane, and 7,7-dimethyl-bicyclo[2.2.1]heptyl. Partially saturated cycloalkyl compounds include, for example, cyclopentenyl, cyclohexenyl, cycloheptenyl, and cyclooctenyl. Unless otherwise expressly stated in this specification, cycloalkyl compounds may optionally be substituted with, for example, oxo, halogen, amino, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocyclic alkyl, heteroaryl, etc. In some embodiments, the cycloalkyl group is optionally substituted with an oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, the cycloalkyl group is optionally substituted with an oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, or -OMe. In some embodiments, the cycloalkyl group is optionally substituted with a halogen. In some embodiments, the cycloalkyl group is unsubstituted.

[0042] "Cycloalkylalkyl" refers to the formula -R c -cycloalkyl groups, wherein R c It is an alkylene chain as described above.

[0043] "Cycloalkylalkoxy" refers to the formula -OR c - A cycloalkyl group bonded by an oxygen atom, wherein R c It is an alkylene chain as described above.

[0044] "Halogen" or "halogenated" refers to halogenated substituents, such as bromine, chlorine, fluorine, and iodine substituents.

[0045] As used herein, the term "haloalkyl" or "haloalkane" refers to an alkyl group as defined above that has been substituted with one or more halogen groups, such as trifluoromethyl, dichloromethyl, bromomethyl, 2,2,2-trifluoroethyl, 1-fluoromethyl-2-fluoroethyl, etc. In some embodiments, the alkyl portion of the fluoroalkyl group may optionally be further substituted. Examples of halogen-substituted alkanes (“haloalkanes”) include halomethanes (e.g., chloromethane, bromomethane, fluoromethane, iodomethane), dihalomethanes and trihalomethanes (e.g., chloroform, tribromomethane, trifluoromethane, triiodomethane), 1-haloethane, 2-haloethane, 1,2-dihaloethane, 1-halopropane, 2-halopropane, 3-halopropane, 1,2-dihalopropane, 1,3-dihalopropane, 2,3-dihalopropane, 1,2,3-trihalopropane, and any other suitable combination of alkanes (or substituted alkanes) and halogens (e.g., Cl, Br, F, I, etc.). When the alkyl group is substituted by more than one halogen group, each halogen can be chosen independently, for example, 1-chloro,2-fluoroethane.

[0046] "Fluoroalkyl" refers to an alkyl group as defined above that is substituted with one or more fluorine groups, such as trifluoromethyl, difluoromethyl, fluoromethyl, 2,2,2-trifluoroethyl, 1-fluoromethyl-2-fluoroethyl, etc.

[0047] "Hydroxyalkyl" refers to an alkyl group as defined above that is substituted with one or more hydroxyl groups. In some embodiments, the alkyl group is substituted with one hydroxyl group. In some embodiments, the alkyl group is substituted with one, two, or three hydroxyl groups. Hydroxyalkyl groups include, for example, hydroxymethyl, hydroxyethyl, hydroxypropyl, hydroxybutyl, or hydroxypentyl. In some embodiments, the hydroxyalkyl group is hydroxymethyl.

[0048] "Aminoalkyl" refers to an alkyl group as defined above that is substituted with one or more amines. In some embodiments, the alkyl group is substituted with one amine. In some embodiments, the alkyl group is substituted with one, two, or three amines. Aminoalkyl groups include, for example, aminomethyl, aminoethyl, aminopropyl, aminobutyl, or aminopentyl. In some embodiments, the aminoalkyl group is aminomethyl.

[0049] A "disulfide" refers to a compound in which two sulfur atoms are bonded together, each sulfur atom comprising an optionally substituted alkyl chain. In some embodiments, the disulfide may be RSS-R'. In some embodiments, R and R' may be the same. In some embodiments, R and R' are different. Each R and R' may be independently selected from C1-C2. 12 Alkyl group. In some embodiments, R or R' may be substituted with an amine, sulfone, or carboxylic acid.

[0050] The term "heteroalkyl" refers to an alkyl group in which one or more skeletal atoms are selected from atoms other than carbon (e.g., oxygen, nitrogen (e.g., -NH-, -N(alkyl)-), sulfur, or combinations thereof). The heteroalkyl group is attached to the remainder of the molecule at a carbon atom. In one aspect, the heteroalkyl group is a C1-C6 heteroalkyl group, wherein it consists of 1 to 6 carbon atoms and one or more atoms other than carbon (e.g., oxygen, nitrogen (e.g., -NH-, -N(alkyl)-), sulfur, or combinations thereof), wherein the heteroalkyl group is attached to the remainder of the molecule at a carbon atom. Examples of such heteroalkyl groups are, for example, -CH2OCH3, -CH2CH2OCH3, -CH2CH2OCH2CH2OCH3, or -CH(CH3)OCH3. Unless otherwise expressly stated in this specification, the heteroalkyl group may optionally be substituted with, for example, oxo, halogen, amino, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, the heteroalkyl group is optionally substituted with an oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, the heteroalkyl group is optionally substituted with an oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, or -OMe. In some embodiments, the heteroalkyl group is optionally substituted with a halogen. In some embodiments, the heteroalkyl group is unsubstituted.

[0051] "Heterocyclic alkyl" refers to a stable 3- to 24-membered or fully saturated cyclic group comprising 2 to 23 carbon atoms and at least one cyclic heteroatom. In some embodiments, the heterocyclic alkyl group contains 1 to 8 heteroatoms selected from nitrogen, oxygen, phosphorus, and sulfur. Unless otherwise expressly stated in this specification, the heterocyclic alkyl group can be a monocyclic, bicyclic, tricyclic, or tetracyclic system, which may include fused ring systems (where the heterocyclic alkyl group is bonded through non-aromatic ring atoms when fused with an aryl or heteroaryl ring) or bridged ring systems; and the nitrogen, carbon, or sulfur atom in the heterocyclic alkyl group may optionally be oxidized; the nitrogen atom may optionally be quaternized.

[0052] Representative heterocyclic alkyl groups include, but are not limited to, heterocyclic alkyl groups having two to fifteen carbon atoms (C2-C4). 15 Heterocyclic alkyl groups (C2-C4) having two to ten carbon atoms. 10Heterocyclic alkyl groups are alkyl groups having two to eight carbon atoms (C2-C8 heterocyclic alkyl groups), two to six carbon atoms (C2-C6 heterocyclic alkyl groups), two to five carbon atoms (C2-C5 heterocyclic alkyl groups), or two to four carbon atoms (C2-C4 heterocyclic alkyl groups). In some embodiments, the heterocyclic alkyl group is a 3- to 6-membered heterocyclic alkyl group. In some embodiments, the heterocyclic alkyl group is a 5- to 6-membered heterocyclic alkyl group. Examples of such heterocyclic alkyl groups include, but are not limited to, azircyclic propane, azircyclic butane, dioxacyclopentane, thienyl[1,3]dithiaalkyl, decahydroisoquinolinyl, imidazolinyl, imidazoalkyl, isothiazolyl, isoxazolyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, 2-oxopiperidinyl, 2-oxopiperidinyl, 2-oxopiperidinyl, 2-oxopiperylalkyl, oxazolyl, piperidinyl, piperazine, 4-piperidinoneyl, pyrrolylalkyl, pyrazolylalkyl, quininecycloyl, thiazoalkyl. Tetrahydrofuranyl, trithiaalkyl, tetrahydropyranyl, thiomorpholinyl, thiamorpholinyl, 1-oxothiomorpholinyl, 1,1-dioxothiomorpholinyl, 1,3-dihydroisobenzofuran-1-yl, 3-oxo-1,3-dihydroisobenzofuran-1-yl, methyl-2-oxo-1,3-dioxacyclopenten-4-yl, and 2-oxo-1,3-dioxacyclopenten-4-yl. The term heterocyclic alkyl also includes all cyclic forms of carbohydrates, including but not limited to monosaccharides, disaccharides, and oligosaccharides. It should be understood that when referring to the number of carbon atoms in a heterocyclic alkyl group, the number of carbon atoms in the heterocyclic alkyl group is different from the total number of atoms (including heteroatoms) constituting the heterocyclic alkyl group (i.e., the skeletal atoms of the heterocyclic alkyl ring). Unless otherwise expressly stated in this specification, heterocyclic alkyl groups are optionally substituted with, for example, oxo, halogen, amino, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocyclic alkyl, heteroaryl, etc. In some embodiments, heterocyclic alkyl groups are optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, heterocyclic alkyl groups are optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, or -OMe. In some embodiments, heterocyclic alkyl groups are optionally substituted with halogens. In some embodiments, the heterocyclic alkyl groups are unsubstituted.

[0053] "Heterocycle" or "heterocyclic group" refers to a saturated, unsaturated, or aromatic ring containing one or more ring heteroatoms. Exemplary heteroatoms include N, O, Si, P, B, and S atoms. Heterocycles include, for example, 3- to 10-membered monocyclic rings and 6- to 12-membered bicyclic rings (such as spirocyclic, fused, or bridged rings). Unless otherwise expressly stated in this specification, the heterocyclic group is a monocyclic, bicyclic, tricyclic, or tetracyclic system, optionally including fused, bridged, or spirocyclic systems. The heteroatoms in the heterocyclic group are optionally oxidized. If present, one or more nitrogen atoms are optionally quaternized. The heterocyclic group can be partially or fully saturated. The heterocyclic group is connected to the rest of the molecule by any atoms on the ring. Examples of such heterocyclic groups include, but are not limited to, dioxacyclopentyl, thienyl[1,3]dithiaalkyl, decahydroisoquinolinyl, imidazolinyl, imidazoalkyl, isothiazolyl, isoxazolyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopiperidinyl, oxazolyl, piperidinyl, piperazinyl, 4-piperidinoneyl, pyrrolyl, pyrazolyl, quininecycloyl, thiazoalkyl, tetrahydrofuranyl, trithiaalkyl, tetrahydropyranyl, thiomorpholinyl, thiomorpholinyl, 1-oxothiomorpholinyl, and 1,1-dioxothiomorpholinyl. Unless otherwise expressly stated in this specification, the term "heterocyclic" is intended to include heterocyclic groups as defined above, optionally substituted with one or more substituents. For example, the heterocyclic group may optionally be substituted by one or more substituents selected from the following: alkyl, alkenyl, ynyl, halo, fluoroalkyl, oxo, thio, cyano, nitro, optionally substituted aryl, optionally substituted aralkyl, optionally substituted arenel, optionally substituted arynyl, optionally substituted carbocyclic, optionally substituted carbocyclic alkyl, optionally substituted heterocyclic, optionally substituted heterocyclic alkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, -R b -OR a -R b -OC(O)-R a -R b -OC(O)-OR a -R b -OC(O)-N(R a )2、-R b -N(R a )2、-R b -C(O)R a -R b -C(O)OR a -R b -C(O)N(R a )2、-R b -CN、-R b -OR e -C(O)N(R a )2、-Rb -N(R a )C(O)OR a -R b -N(R a )C(O)R a -R b -N(R a S(O) t R a (where t is 1 or 2), -R b -S(O) t R a (where t is 1 or 2), -R b -S(O) t OR a (where t is 1 or 2) and -R b -S(O) t N(R a )2 (where t is 1 or 2), where each R a Independently, it is hydrogen, alkyl (optionally substituted with halogen, hydroxyl, methoxy, or trifluoromethyl), fluoroalkyl, cycloalkyl (optionally substituted with halogen, hydroxyl, methoxy, or trifluoromethyl), cycloalkylalkyl (optionally substituted with halogen, hydroxyl, methoxy, or trifluoromethyl), aryl (optionally substituted with halogen, hydroxyl, methoxy, or trifluoromethyl), aralkyl (optionally substituted with halogen, hydroxyl, methoxy, or trifluoromethyl), heterocyclic (optionally substituted with halogen, hydroxyl, methoxy, or trifluoromethyl), heterocyclic alkyl (optionally substituted with halogen, hydroxyl, methoxy, or trifluoromethyl), heteroaryl (optionally substituted with halogen, hydroxyl, methoxy, or trifluoromethyl), or heteroarylalkyl (optionally substituted with halogen, hydroxyl, methoxy, or trifluoromethyl), each R b Independently, it is a direct bond or a straight-chain or branched alkylene or alkenylene chain, and R e It is a straight-chain or branched alkylene or alkenylene chain, and each of the above substituents is unsubstituted unless otherwise stated.

[0054] "Heteroaryl" or "aromatic heterocycle" refers to a cyclic group comprising a carbon atom and one or more cyclic heteroatoms (e.g., selected from nitrogen, oxygen, phosphorus, silicon, and sulfur) and at least one aromatic ring. In some embodiments, the heteroaryl group is a 5- to 14-membered cyclic group comprising one to thirteen carbon atoms and one to six heteroatoms selected from nitrogen, oxygen, phosphorus, and sulfur. The heteroaryl group can be a monocyclic, bicyclic, tricyclic, or tetracyclic system, which may include a fused ring system (where the heteroaryl group is bonded through aromatic ring atoms when fused with a cycloalkyl or heterocyclic alkyl ring) or a bridged ring system; and the nitrogen, carbon, or sulfur atom in the heteroaryl group may optionally be oxidized; the nitrogen atom may optionally be quaternized. In some embodiments, the heteroaryl group is a 5- to 10-membered heteroaryl group. In some embodiments, the heteroaryl group is a 5- to 6-membered heteroaryl group. Examples include, but are not limited to, azepinyl, acridine, benzimidazolyl, benzothiazolyl, benzoindolyl, benzodioxolyl, benzofuranyl, benzooxazolyl, benzothiazolyl, benzothiadiazolyl, benzo[b][1,4]dioxepinyl, 1,4-benzodioxylalkyl, benzonaphthofuranyl, benzooxazolyl, benzodioxolyl, benzodioxolyl, benzodioxolyl, benzodioxinyl, benzopyranyl, benzopyranoneyl, benzofuranyl, benzofuranoneyl, benzothienyl / benzothiophenyl, benzotriazolyl, benzo[4,6]imidazo[1,2-a]pyridine Carbazolyl, cyclophosphinyl, dibenzofuranyl, dibenzothiophenyl, furanyl, furanoneyl, isothiazolyl, imidazolyl, indazole, indoleyl, indazoleyl, isoindoleyl, indolelinyl, isoindolelinyl, isoquinolinyl, indoleazinyl, isoxazolyl, naphthidyl, oxadiazolyl, 2-oxoazapyridine, oxazolyl, ethylene oxide, 1-pyridinyl oxide, 1-pyrimidinyl oxide, 1-pyrazinyl oxide, 1-oxopyridinyl oxide The heteroaryl group can be substituted with, for example, pyridazinyl, 1-phenyl-1H-pyrrolyl, phenazinyl, phenothiazinyl, phenoxazinyl, phthalazinyl, pteridinyl, purine, pyrrolyl, pyrazolyl, pyridinyl, pyridinyl, pyrazinyl, quinazolinyl, quinoxolinyl, quinolinyl, quininecycloyl, isoquinolinyl, tetrahydroquinolinyl, thiazolyl, thiadiazolyl, triazolyl, tetrazolyl, triazinyl, and thiophenyl / thienyl. Unless otherwise expressly stated in this specification, the heteroaryl group may optionally be substituted with, for example, halogen, amino, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, the heteroaryl group may optionally be substituted with halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2.In some embodiments, the heteroaryl group is optionally substituted with a halogen, methyl, ethyl, -CN, -CF3, -OH, or -OMe. In some embodiments, the heteroaryl group is optionally substituted with a halogen. In some embodiments, the heteroaryl group is unsubstituted.

[0055] The term “substitution” refers to a portion in a structure where hydrogen atoms on one or more carbon atoms or substituted heteroatoms (e.g., NH) are replaced by a substituent. It should be understood that “substitution” or “substituted” includes the implicit condition that such substitution occurs according to the permissible valence of the substituted atom and the substituent, and that the substitution produces a stable compound, i.e., a compound that does not spontaneously undergo transformations such as rearrangement, cyclization, elimination, etc. In some embodiments, substitution refers to a portion having a substituent that replaces two hydrogen atoms on the same carbon atom, such as replacing two hydrogen atoms on a single carbon atom with an oxo, imino, or thio group. As used herein, the term “substitution” is intended to include all permissible substituents in an organic compound. Broadly speaking, permissible substituents include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, aromatic and non-aromatic substituents in organic compounds. For a suitable organic compound, permissible substituents may be one or more, and may be the same or different. For the purposes of this disclosure, heteroatoms (such as nitrogen) may have hydrogen substituents and / or any permissible substituents in the organic compounds described herein that satisfy the valence of the heteroatom.

[0056] In some embodiments, the substituents may include any substituents described herein, such as: halogen, hydroxyl, oxo (=O), thio (=S), cyano (-CN), nitro (-NO2), imino (=NH), oxime (=N-OH), hydrazine (=N-NH2), -R b -OR a -R b -OC(O)-R a -R b -OC(O)-OR a -R b -OC(O)-N(R a )2、-R b -N(R a )2、-R b -C(O)R a -R b -C(O)OR a -R b -C(O)N(R a )2、-R b -OR c -C(O)N(R a )2、-R b -N(R a)C(O)OR a -R b -N(R a )C(O)R a -R b -N(R a S(O) t R a (where t is 1 or 2), -R b -S(O) t R a (where t is 1 or 2), -R b -S(O) t OR a (where t is 1 or 2) and -R b -S(O) t N(R a )2 (where t is 1 or 2); and alkyl, alkenyl, alkynyl, aryl, aralkyl, areneyl, arynyl, cycloalkyl, cycloalkylalkyl and heterocyclic, any of which may optionally be replaced by alkyl, alkenyl, alkynyl, halogen, haloalkyl, haloalkenyl, haloalkynyl, oxo (=O), thio (=S), cyano (-CN), nitro (-NO2), imino (=NH), oxime (=N-OH), hydrazine (=N-NH2), SF 5 -R b -OR a -R b -OC(O)-R a -R b -OC(O)-OR a -R b -OC(O)-N(R a )2、-R b -N(R a )2、-R b -C(O)R a -R b -C(O)OR a -R b -C(O)N(R a )2、-R b -OR c -C(O)N(R a )2、-R b -N(R a )C(O)OR a -R b -N(R a )C(O)R a -R b -N(R a S(O) t R a(where t is 1 or 2), -R b -S(O) t R a (where t is 1 or 2), -R b -S(O) t OR a (where t is 1 or 2) and -R b -S(O) t N(R a )2 (where t is 1 or 2) replace; where each R a Independently selected from hydrogen, alkyl, cycloalkyl, cycloalkylalkyl, aryl, aralkyl, and heterocyclic, wherein each R a Where the oxidation state allows, it may be optionally replaced by alkyl, alkenyl, alkynyl, halogen, haloalkyl, haloalkenyl, haloalkynyl, oxo (=O), thio (=S), cyano (-CN), nitro (-NO2), imino (=NH), oxime (=N-OH), hydrazine (=N-NH2), or -R. b -OR a -R b -OC(O)-R a -R b -OC(O)-OR a -R b -OC(O)-N(R a )2、-R b -N(R a )2、-R b -C(O)R a -R b -C(O)OR a -R b -C(O)N(R a )2、-R b -OR c -C(O)N(R a )2、-R b -N(R a )C(O)OR a -R b -N(R a )C(O)R a -R b -N(R a S(O) t R a (where t is 1 or 2), -R b -S(O) t R a (where t is 1 or 2), -R b -S(O) t OR a(where t is 1 or 2) and -R b -S(O) t N(R a )2 (where t is 1 or 2) replace; where each R b Independently selected from direct-chain or straight-chain or branched alkylene, alkenylene, or ynylene chains, and each R c It is a straight-chain or branched alkylene, alkenylene, or ynylene chain.

[0057] The terms “optional” or “optionally” mean that the event or condition described below may or may not occur, and the description includes both the occurrence and non-occurrence of the event or condition. For example, “optionally substituted alkyl” means “alkyl” or “substituted alkyl” as defined above. Furthermore, the optionally substituted group can be unsubstituted (e.g., -CH2CH3), fully substituted (e.g., -CF2CF3), monosubstituted (e.g., -CH2CH2F), or substituted at any level between full and monosubstituted (e.g., -CH2CHF2, -CH2CF3, -CF2CH3, -CFHCHF2, etc.).

[0058] The term "biomolecule" refers to biological components that can participate in crown formation, including but not limited to, proteins, polypeptides, polysaccharides, sugars, lipids, lipoproteins, metabolites, oligonucleotides, metabolomics, or combinations thereof. It is conceivable that a biomolecular crown of unique particles may contain some of the same biomolecules, may contain unique biomolecules related to other sensor elements, and / or may differ in level or amount, or in the type or structure of the biomolecules bound to each sensor element. In one embodiment, the biomolecules are selected from proteins, nucleic acids, lipids, and metabolomics.

[0059] The ranges provided in this document should be understood as abbreviations of all values ​​within that range. For example, the range 1 to 50 should be understood to include any number, combination of numbers, or subrange of numbers that come from the following groups: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50, as well as all intermediate decimal values ​​between the above integers, such as, for example, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, and 1.9. For subranges, consider in particular “nested subranges” that extend from either end of the range. For example, nested subranges of the example range 1 to 50 could include 1 to 10, 1 to 20, 1 to 30, and 1 to 40 in one direction, or 50 to 40, 50 to 30, 50 to 20, and 50 to 10 in another direction.

[0060] The compounds and structures provided herein may be stereoisomers. In some cases, the compounds or structures disclosed herein may form stereoisomers. In some cases, the stereoisomers may be diastereomers (e.g., cis / trans isomers, E / Z isomers, conformational isomers, or rotational isomers). In some cases, the stereoisomers may be enantiomers (R, S enantiomers, or + / - enantiomers). In some cases, the compounds or structures disclosed herein may be enantiomerically pure (e.g., 100% pure). In some cases, the compounds or structures may form racemic mixtures of enantiomers (e.g., 50% pure). In some cases, the compounds or structures disclosed herein can be stabilized into stereoisomers, wherein the compounds or structures disclosed herein comprise at least about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 96%, about 97%, about 98%, about 98.5%, about 99%, about 99.1%, about 99.2%, about 99.3%, about 99.4%, about 99.5%, about 99.6%, about 99.7%, about 99.8%, about 99.9% or more of the compounds or structures and the corresponding stereoisomers.

[0061] macromolecular structure This document provides macromolecular structures comprising a surface, a tethered portion coupled to the surface, and a macromolecular chain. In some embodiments, the macromolecular structure provided herein comprises a surface and a macromolecular chain. In some embodiments, the macromolecular structure provided herein comprises a surface. In some embodiments, the macromolecular structure provided herein comprises a tethered portion (e.g., a tethered portion coupled to a surface). In some embodiments, the macromolecular structure provided herein comprises a macromolecular chain. In some embodiments, a first end of the macromolecular chain is covalently attached to the tethered portion. In some embodiments, a second end of the macromolecular chain is not coupled to the surface. In some embodiments, the macromolecular chain comprises one or more (e.g., different) repeating units derived from a monomer.

[0062] In some embodiments, this document provides a macromolecular structure comprising a surface, a tethered portion coupled to the surface, and a macromolecular chain. In some embodiments, the macromolecular structure comprises a surface. In some embodiments, the macromolecular structure comprises a tethered portion (e.g., a tethered portion coupled to a surface). In some embodiments, the macromolecular structure comprises a macromolecular chain. In some embodiments, the macromolecular chain comprises two or more distinct repeating units derived from different monomers. In some embodiments, the macromolecular chain comprises at least two (e.g., at least three, at least four, at least five) distinct repeating units derived from different monomers. In some embodiments, the macromolecular chain comprises up to ten (e.g., up to nine, up to eight, up to six, up to four, up to three) distinct repeating units derived from different monomers. In some embodiments, the macromolecular chain comprises two distinct repeating units derived from different monomers. In some embodiments, the macromolecular chain comprises three distinct repeating units derived from different monomers. In some embodiments, the macromolecular chain comprises four distinct repeating units derived from different monomers. In some embodiments, the macromolecular chain comprises five distinct repeating units derived from different monomers.

[0063] In some embodiments, this document provides a macromolecular structure comprising a surface, a tethered portion coupled to the surface, and a macromolecular chain. In some embodiments, the macromolecular structure comprises a surface. In some embodiments, the macromolecular structure comprises a tethered portion (e.g., a tethered portion coupled to a surface). In some embodiments, the macromolecular structure comprises a macromolecular chain. In some embodiments, the macromolecular chain comprises monomer-derived repeating units. In some embodiments, the macromolecular chain consists of a single monomer repeating unit.

[0064] In some cases, the tethering and repeating units used for functionalized surfaces affect the physicochemical properties of the surface, such as size, surface charge, hydrophobicity, hydrophilicity, surface functionality, surface morphology, surface curvature, porosity, shape, and any combination thereof. Changes in physicochemical properties can affect the binding properties of macromolecular structures with other compounds, such as biomolecules (e.g., proteins), leading to increases or decreases in binding efficiency.

[0065] In some embodiments, the macromolecular structures provided herein comprise crosslinked polymers (e.g., crosslinked macromolecular chains). For example, the relative amount of the crosslinking monomer to the total monomers in the polymer, by weight or quantity, may be at least 0.1%, at least 0.5%, at least 1%, or at least 2%. In some embodiments, the macromolecular structures provided herein comprise substantially non-crosslinked polymers (e.g., non-crosslinked macromolecular chains). For example, the relative amount of the crosslinking monomer to the total monomers in the polymer, by weight or quantity, may be less than 0.1%, less than 0.05%, less than 0.01%, or about 0%. In some embodiments, the macromolecular structures provided herein comprise polymer brushes (e.g., PEG brushes). For example, the polymer brush may comprise side chains having at least 5, at least 10, at least 15, or at least 20 repeating units derived from one or more monomers.

[0066] In some implementations, it is a macromolecular structure comprising formula (A): ATC In some embodiments, A is a surface, such as the surface provided elsewhere herein. In some embodiments, T is a tethered portion, such as the tethered portion provided elsewhere herein. In some embodiments, C is a macromolecular chain, such as the macromolecular chain provided elsewhere herein. In some embodiments, C comprises a plurality of monomer-derived repeating units. In some embodiments, the plurality of repeating units comprises a single monomer. In other embodiments, the plurality of repeating units comprises two or more different monomers. In some embodiments, the monomers have a controlled distribution throughout the macromolecular chain. In some embodiments, the monomers have a random distribution throughout the macromolecular chain.

[0067] In some embodiments, the macromolecular chain provided herein comprises repeating units derived from monomers represented by the following structures:

[0068] In some implementation schemes, R 1 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1 It is hydrogen. In some implementations, R 1 It is a C1-C6 alkyl group.

[0069] In some implementation schemes, R2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2 It is hydrogen. In some implementations, R 2 It is a C1-C6 alkyl group.

[0070] In some implementation schemes, R 3 It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3 It is hydrogen. In some implementations, R 3 It is a C1-C6 alkyl group. In some embodiments, R 3 It is a methyl group.

[0071] In some implementation schemes, R 4 It is hydrogen, a sulfonate, a carboxylic acid ester, a C1-C4 alkylene group, an amine (e.g., a quaternary ammonium cation), or a C1-C6 alkyl group optionally substituted with a halogen. In some embodiments, R 4 It is hydrogen. In some implementations, R 4 It does not exist. In some implementations, R 4 It is a sulfonate. In some embodiments, R 4 It is an amine. In some implementations, R 4 It is a quaternary ammonium cation. In some embodiments, R 4 It is a carboxylic acid ester. In some embodiments, R 4 It is a C1-C4 alkylene group. In some embodiments, R 4 It is a C1-C6 alkyl group that is optionally substituted with a halogen (e.g., a haloalkyl group).

[0072] In some implementations, X is -C- or -N-. In some implementations, X is -C-. In some implementations, X is -N-.

[0073] In some implementations, Y is -C- or -N-. In some implementations, Y is -C-. In some implementations, Y is -N-.

[0074] In some embodiments, the macromolecular chain provided herein comprises repeating units derived from monomers represented by the following structures:

[0075] In some implementation schemes, R 1 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1 It is hydrogen. In some implementations, R 1 It is a C1-C6 alkyl group.

[0076] In some implementation schemes, R2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2 It is hydrogen. In some implementations, R 2 It is a C1-C6 alkyl group.

[0077] In some implementation schemes, R 3 It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3 It is hydrogen. In some implementations, R 3 It is a C1-C6 alkyl group. In some embodiments, R 3 It is a methyl group.

[0078] In some implementation schemes, R 9 It is either hydrogen or oxygen. In some implementations, R 9 It is hydrogen. In some implementations, R 9 It is oxygenation.

[0079] In some implementations, X is -C- or -N-. In some implementations, X is -C-. In some implementations, X is -N-.

[0080] In some implementations, Y is -C-, -N-, or -O-. In some implementations, Y is -C-. In some implementations, Y is -O-. In some implementations, Y is -N-.

[0081] In some implementation schemes, It can be a single bond or a double bond. In some implementations, It is a single bond. In some implementations, It is a double bond. In some implementations, One of them is a single bond and One of them is a double bond. In some implementations, the two... It is a single bond. In some implementations, two... It is a double bond.

[0082] In some embodiments, the macromolecular chain provided herein comprises repeating units derived from monomers represented by the following structures:

[0083] In some embodiments, the repeating unit is derived from an acrylate monomer. In some embodiments, the repeating unit is derived from a methacrylate monomer.

[0084] In some implementation schemes, R 1 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1It is hydrogen. In some implementations, R 1 It is a C1-C6 alkyl group.

[0085] In some implementation schemes, R 2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2 It is hydrogen. In some implementations, R 2 It is a C1-C6 alkyl group.

[0086] In some implementation schemes, R 3 It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3 It is hydrogen. In some implementations, R 3 It is a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3 It is a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more methyl groups. In some embodiments, R 3 It is a C1-C6 alkyl group. In some embodiments, R 3 It is a methyl group.

[0087] In some implementation schemes, R 5 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, an amine, an azide, a sulfonate, a carbamate, an asymmetric disulfide, or a C1-C8 alkyl group or a C1-C6 heterocyclic group substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines further substituted with amines, hydroxyl groups, aryl groups, or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogen groups, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted (e.g., two or more fused 6 membered rings optionally further substituted), optionally substituted benzyl groups, trimethoxysilanes, or phosphoric acid choline-substituted -C1-C3 or C1-C4 alkylylenes. In some embodiments, R 5 It is C1-C 12 Alkylamines.

[0088] In some implementation schemes, R 5 It is hydrogen. In some implementations, R 5 It is a C1-C6 alkyl group. In some embodiments, R 5 It is a C1-C4 alkylyne. In some embodiments, R 5It is a C1-C8 alkyl or C1-C6 alkyl group substituted with one or more hydroxyl groups, amines, azides, sulfonates, carbamates, asymmetric disulfides, or optionally a C1-C6 alkyl or oxo-substituted 3-, 5-, or 6-membered heterocycles. n1 Ethylene glycol. In some implementations, R 5 It is a C1-C8 alkyl or C1-C6 alkyl group substituted with one or more hydroxyl groups, amines, azides, carbamates, asymmetric disulfides, or optionally a C1-C6 alkyl group or an oxo-substituted 3-, 5-, or 6-membered heterocycle. n1 Ethylene glycol. In some implementations, R 5 It is a C1-C8 alkylamine. In some embodiments, R 5 It is a C1-C8 alkylamine substituted with an amine. In some embodiments, R 5 It is a C1-C8 alkylamine substituted with hydroxyl and amine groups. In some embodiments, R 5 It is a C1-C8 alkylamine substituted with a sulfonate. In some embodiments, R 5 It is an aryl-substituted C1-C8 alkylamine. In some embodiments, R 5 It is a C1-C8 alkoxy group. In some embodiments, R 5 It is a C1-C8 alkoxy group substituted with one or more oxo or halogen groups. In some embodiments, R 5 It is a C1-C8 alkoxy group substituted with one or more oxo groups. In some embodiments, R 5 It is a C1-C8 alkoxy group substituted with one or more halogens. In some embodiments, alkylamines include quaternary ammonium cations.

[0089] In some implementation schemes, R 5 It is a C1-C3 alkyl group optionally substituted with one or more pyrene, two or more optionally further substituted fused 5-6 membered rings (e.g., two or more optionally further substituted fused 6 membered rings), optionally substituted benzyl, trimethoxysilane, or phosphocholine. In some embodiments, R 5 It is a C1-C3 alkyl group substituted with pyrene. In some embodiments, R 5 It is a C1-C3 alkyl group substituted with two or more optionally further substituted 5-6-membered rings (such as two or more optionally further substituted fused 6-membered rings). In some embodiments, R 5 It is a C1-C3 alkyl group that has been optionally substituted with a benzyl group. In some embodiments, R 5 It is a C1-C3 alkyl group substituted with trimethoxysilane. In some embodiments, R 5 It is a C1-C3 alkyl group substituted with phosphocholine.

[0090] In some implementation schemes, R 5 It is a hydrogen, C1-C6 alkyl, or a C1-C3 alkyl group substituted with pyrene or two or more optionally further substituted fused 5-6 membered rings. In some embodiments, R 5 It is a hydrogen, C1-C6 alkyl, or a C1-C3 alkyl group substituted with pyrene or two or more fused 5-6 membered rings optionally further substituted, and the macromolecular chain contains two or more distinct repeating units.

[0091] In some embodiments, the macromolecular chain provided herein comprises repeating units derived from monomers represented by the following structures:

[0092] In some implementations, the repeating unit is derived from an acrylamide monomer.

[0093] In some implementation schemes, R 1 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1 It is hydrogen. In some implementations, R 1 It is a C1-C6 alkyl group.

[0094] In some implementation schemes, R 2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2 It is hydrogen. In some implementations, R 2 It is a C1-C6 alkyl group.

[0095] In some implementation schemes, R 3 It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3 It is hydrogen. In some implementations, R 3 It is a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3 It is a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more methyl groups. In some embodiments, R 3 It is 3,4-dimethyl-1 H -pyrrole-2,5-dione. In some embodiments, R 3 It is a C1-C6 alkyl group. In some embodiments, R 3 It is a methyl group.

[0096] In some implementation schemes, R 6 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 6 It is hydrogen. In some implementations, R 6It is a C1-C6 alkyl group.

[0097] In some implementation schemes, R 7 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, amines, azides, sulfonates, carbamates, or asymmetric disulfides, or a 3-, 5-, or 6-membered heterocyclic or C1-C alkyl group optionally substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines further substituted with amines or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogen groups, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6-membered rings optionally further substituted (e.g., two or more fused 6-membered rings optionally further substituted), optionally substituted benzyl groups, trimethoxysilanes, or -C1-C3 or C1-C4 alkylyl groups substituted with phosphocholine.

[0098] In some implementation schemes, R 7 It is hydrogen. In some implementations, R 7 It is a C1-C6 alkyl group. In some embodiments, R 7 It is a C1-C4 alkylyne. In some embodiments, R 7 It is a C1-C8 alkyl group substituted with one or more hydroxyl groups, amines, azides, sulfonates, carbamates, or asymmetric disulfides, or a 3-, 5-, or 6-membered heterocyclic or C1-C alkyl group substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol. In some implementations, R 7 It is a C1-C8 alkyl group substituted with one or more hydroxyl groups, amines, azides, urethanes, or asymmetric disulfides, or a 3-, 5-, or 6-membered heterocyclic or C1-C substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol. In some implementations, R 7 It is a C1-C8 alkylamine. In some embodiments, R 7 It is a C1-C8 alkylamine substituted with an amine. In some embodiments, R 7 It is a C1-C8 alkylamine substituted with a sulfonate. In some embodiments, R 7 It is a C1-C8 alkoxy group. In some embodiments, R 7 It is a C1-C8 alkoxy group substituted with one or more oxo or halogen groups. In some embodiments, R 7 It is a C1-C8 alkoxy group substituted with one or more oxo groups. In some embodiments, R 7 It is a C1-C8 alkoxy group substituted with one or more halogens. In some embodiments, R 7It is a C1-C6 alkyl group optionally substituted with a hydroxyl group, a substituted benzene, or a hydrogen.

[0099] In some implementation schemes, R 7 It is a C1-C3 alkyl group optionally substituted with one or more pyrene, two or more optionally further substituted fused 5-6 membered rings (e.g., two or more optionally further substituted fused 6 membered rings), optionally substituted benzyl, trimethoxysilane, or phosphocholine. In some embodiments, R 7 It is a C1-C3 alkyl group substituted with pyrene. In some embodiments, R 7 It is a C1-C3 alkyl group substituted with two or more optionally further substituted 5-6-membered rings (such as two or more optionally further substituted fused 6-membered rings). In some embodiments, R 7 It is a C1-C3 alkyl group that has been optionally substituted with a benzyl group. In some embodiments, R 7 It is a C1-C3 alkyl group substituted with trimethoxysilane. In some embodiments, R 7 It is a C1-C3 alkyl group substituted with phosphocholine.

[0100] In some embodiments, the macromolecular chain provided herein comprises repeating units derived from monomers represented by the following structures:

[0101] In some embodiments, the monomer-derived repeating unit can be a diene, such as a cis-diene. In some embodiments, the diene can be used as a crosslinking monomer.

[0102] In some implementation schemes, R 1 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1 It is hydrogen. In some implementations, R 1 It is a C1-C6 alkyl group.

[0103] In some implementation schemes, R 2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2 It is hydrogen. In some implementations, R 2 It is a C1-C6 alkyl group.

[0104] In some implementation schemes, R 3 It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3 It is hydrogen. In some implementations, R 3 It is a C1-C6 alkyl group. In some embodiments, R 3 It is a methyl group.

[0105] In some implementation schemes, R 1’ It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1’ It is hydrogen. In some implementations, R 1’ It is a C1-C6 alkyl group.

[0106] In some implementation schemes, R 2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2’ It is hydrogen. In some implementations, R 2’ It is a C1-C6 alkyl group.

[0107] In some implementation schemes, R 3’ It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3’ It is hydrogen. In some implementations, R 3’ It is a C1-C6 alkyl group. In some embodiments, R 3’ It is a methyl group.

[0108] In some embodiments, the macromolecular chain provided herein comprises repeating units derived from monomers represented by the following structures:

[0109] In some implementations, the repeating unit is derived from acrylonitrile monomer.

[0110] In some implementation schemes, R 1 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1 It is hydrogen. In some implementations, R 1 It is a C1-C6 alkyl group.

[0111] In some implementation schemes, R 2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2 It is hydrogen. In some implementations, R 2 It is a C1-C6 alkyl group.

[0112] In some implementation schemes, R 3 It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3 It is hydrogen. In some implementations, R 3 It is a C1-C6 alkyl group. In some embodiments, R 3 It is a methyl group.

[0113] In some implementation schemes, R1 It is hydrogen, R 2 It is hydrogen, and R 3 It is hydrogen.

[0114] In some embodiments, the macromolecular chain provided herein comprises repeating units derived from monomers represented by the following structures:

[0115] In some implementation schemes, R 1 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1 It is hydrogen. In some implementations, R 1 It is a C1-C6 alkyl group.

[0116] In some implementation schemes, R 2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2 It is hydrogen. In some implementations, R 2 It is a C1-C6 alkyl group.

[0117] In some embodiments, the macromolecular chain provided herein comprises repeating units derived from monomers represented by the following structures:

[0118] In some implementation schemes, R 1 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1 It is hydrogen. In some implementations, R 1 It is a C1-C6 alkyl group.

[0119] In some implementation schemes, R 2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2 It is hydrogen. In some implementations, R 2 It is a C1-C6 alkyl group.

[0120] In some implementation schemes, R 3 It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3 It is hydrogen. In some implementations, R 3 It is a C1-C6 alkyl group. In some embodiments, R 3 It is a methyl group.

[0121] In some implementation schemes, R 6 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 6 It is hydrogen. In some implementations, R 6It is a C1-C6 alkyl group. In some embodiments, R 6 It is a methyl group.

[0122] In some embodiments, Q is -CH2- or ethylene glycol. In some embodiments, Q is -CH2-. In some embodiments, Q is ethylene glycol.

[0123] In some implementations, m is 1-20. In some implementations, m is 1-10. In some implementations, m is 1-5. In some implementations, m is 5-10. In some implementations, m is 10-20. In some implementations, m is 2. In some implementations, m is 1. In some implementations, m is 3, 4, 5, 6, 7, 8, 9, or 10.

[0124] In some embodiments, A is a polymer side chain comprising repeating units derived from monomers represented by the following structures:

[0125] In some implementation schemes, R 1 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1 It is hydrogen. In some implementations, R 1 It is a C1-C6 alkyl group.

[0126] In some implementation schemes, R 2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2 It is hydrogen. In some implementations, R 2 It is a C1-C6 alkyl group.

[0127] In some implementation schemes, R 3 It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3 It is hydrogen. In some implementations, R 3 It is a C1-C6 alkyl group. In some embodiments, R 3 It is a methyl group.

[0128] In some implementation schemes, R 1 It is hydrogen, R 2 It is hydrogen, and R 3 It is hydrogen.

[0129] In some implementation schemes, R 5It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, an amine, an azide, a sulfonate, a carbamate, an asymmetric disulfide, or a C1-C8 alkyl group or a C1-C6 heterocyclic group substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines optionally further substituted with amines or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogen groups, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted, optionally substituted benzyl groups, trimethoxysilanes, or phosphoric acid choline-substituted -C1-C3 or C1-C4 alkylyl groups. In some embodiments, R 5 It is polyethylene glycol. In some implementations, R 5 It is polyethylene glycol with a chain length of 9.

[0130] In some implementation schemes, R 1 It is hydrogen, R 2 It is hydrogen, R 3 It is a methyl group, Q is -CH2-, and R is... 6 It is methyl, and R 5 It is polyethylene glycol.

[0131] In some embodiments, the macromolecular chain provided herein comprises repeating units derived from monomers represented by the following structures:

[0132] In some embodiments, the repeating unit is derived from a dimethacrylate monomer. In some embodiments, the dimethacrylate monomer is a crosslinking monomer.

[0133] In some implementation schemes, R 1 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1 It is hydrogen. In some implementations, R 1 It is a C1-C6 alkyl group. In some embodiments, R 2 It is hydrogen or C1-C6 alkyl.

[0134] In some implementation schemes, R 2 It is hydrogen. In some implementations, R 2 It is a C1-C6 alkyl group.

[0135] In some implementation schemes, R 3 It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3 It is hydrogen. In some implementations, R 3 It is a C1-C6 alkyl group. In some embodiments, R3 It is a methyl group.

[0136] In some implementation schemes, R 1’ It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1’ It is hydrogen. In some implementations, R 1’ It is a C1-C6 alkyl group.

[0137] In some implementation schemes, R 2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2’ It is hydrogen. In some implementations, R 2’ It is a C1-C6 alkyl group.

[0138] In some implementation schemes, R 3’ It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3’ It is hydrogen. In some implementations, R 3’ It is a C1-C6 alkyl group. In some embodiments, R 3’ It is a methyl group.

[0139] In some implementations, Z is -O- or -NH. In some implementations, Z is -O-. In some implementations, Z is -NH.

[0140] In some implementation schemes, R 8 It is a C1-C6 alkyl group, a divalent metal, or a symmetrical or asymmetric disulfide. In some embodiments, R 8 It is a C1-C6 alkyl group. In some embodiments, R 8 It is a divalent metal. In some implementations, R 8 It is cadmium (II). In some implementations, R 8 It is a symmetrical disulfide or an asymmetrical disulfide. In some embodiments, R 8 It is a symmetrical disulfide. In some embodiments, R 8 It is a symmetrical disulfide (e.g., CH2CH2S-SCH2CH2) or a divalent metal.

[0141] In any macromolecular structure presented in this article, n 1 It is an integer selected from 1 to 100. In some implementations, n 1 It is an integer selected from 1 to 20. In some implementations, n 1 It is an integer selected from 1 to 10. In some implementations, n 1 It is an integer selected from 1 to 5. In some implementations, n 1It is an integer selected from 5 to 10. In some implementations, n 1 It is an integer selected from 10 to 20. In some implementations, n 1 It is 9. In some implementations, n 1 It is an integer selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0142] In some embodiments, any macromolecular structure provided herein may be terminated by a terminating group. The terminating group may be a halogen, an epoxide, or an alkene (e.g., an alkylene group). In some embodiments, the terminating group is a halogen. In some embodiments, the terminating group is bromine. In some embodiments, the terminating group is iodine. In some embodiments, the terminating group is chlorine. In some embodiments, the terminating group is an epoxide. In some embodiments, the terminating group is an alkene.

[0143] In some embodiments, the macromolecular chains provided herein comprise monomer-derived repeating units represented by the structures in Table 1. In some embodiments, the macromolecular chains comprise two or more distinct monomer-derived repeating units represented by the structures in Table 1.

[0144] Table 1

[0145] In some embodiments, the macromolecular chains provided herein comprise monomer-derived repeating units (e.g., one or more) represented by the structures in Table 2.

[0146] Table 2

[0147] In some embodiments, the repeating units provided herein are randomly distributed throughout the macromolecular chain. In some embodiments, the repeating units provided herein have a controlled distribution throughout the macromolecular chain. In some embodiments, the macromolecular chain is a homopolymer. In some embodiments, the macromolecular chain is a block copolymer. In some embodiments, the macromolecular chain is a random copolymer. Those skilled in the art will understand, through the guidance of the disclosure herein, how to provide polymers with a random or controlled distribution of repeating units.

[0148] When multiple unique repeating units are present in a macromolecular chain, these repeating units can exist in the same or different ratios. The ratio of the unique repeating units can be controlled in a manner known to those skilled in the art, including by modifying the stoichiometry of the added monomers.

[0149] In some embodiments, this document provides a macromolecular structure comprising (I) a surface and (II) macromolecular chains coupled to the surface. In some embodiments, the macromolecular chains comprise repeating units of formula (I): Formula (I).

[0150] In some implementation schemes, R 1’’ It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1’’ It is hydrogen. In some implementations, R 1’’ It is a C1-C6 alkyl group. In some embodiments, R 1’’ It is a methyl group.

[0151] In some implementation schemes, R 2’’ It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2’’ It is a C1-C6 alkyl group. In some embodiments, R 2’’ It is a methyl group.

[0152] In some implementation schemes, R 3’’ It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 3’’ It is hydrogen. In some implementations, R 3’’ It is a C1-C6 alkyl group. In some embodiments, R 3’’ It is a methyl group.

[0153] In some implementation schemes, R 1’’ R 2’’ and R 3’’ It is a C1-C6 alkyl group (e.g., methyl).

[0154] In some implementations, n is an integer from 1 to 10,000. In some implementations, n is an integer from 1 to 200. In some implementations, n is an integer not exceeding 25,000 (e.g., not exceeding 20,000, 15,000, 10,000, 5,000, 2,500, 1,000, 500, 100, or 50). In some implementations, n is an integer from 1 to 5,000. In some implementations, n is an integer from 1 to 2,500. In some implementations, n is an integer from 1 to 1,000. In some implementations, n is an integer not exceeding 200. In some implementations, n is an integer from 1 to 100. In some implementations, n is an integer from 1 to 50.

[0155] In some embodiments, A is a polymer side chain comprising any repeating unit provided elsewhere herein. In some embodiments, A is poly(oxyethylene) methacrylate or poly(oxyethylene) acrylate. In some embodiments, A is poly(ethylene glycol) methacrylate or poly(ethylene glycol) acrylate. In some embodiments, A is poly(oxyethylene) methacrylamide or poly(oxyethylene) acrylamide. In some embodiments, A is poly(ethylene glycol) methacrylamide or poly(ethylene glycol) acrylamide.

[0156] In some embodiments, L represents the joint portion. In some embodiments, the joint portion is represented by the following structure:

[0157] In some implementations, each Z is independently -O- or -N-. In some implementations, Z is -O-. In some implementations, Z is -N-.

[0158] In some embodiments, X' is a C1-C6 alkyl group. In some embodiments, X' is a methyl group. In some embodiments, X' is an ethyl group. In some embodiments, X' is a propyl group.

[0159] In some embodiments, the macromolecular structure further includes a cross-linked portion. In some embodiments, the cross-linked portion comprises a structure represented by the following:

[0160] In some embodiments, the crosslinking portion is derived from a dimethacrylate monomer, such as a dimethacrylate monomer as described elsewhere herein. In some embodiments, R 1 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1 It is hydrogen. In some implementations, R 1It is a C1-C6 alkyl group.

[0161] In some implementation schemes, R 2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2 It is hydrogen. In some implementations, R 2 It is a C1-C6 alkyl group.

[0162] In some implementation schemes, R 3 It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3 It is hydrogen. In some implementations, R 3 It is a C1-C6 alkyl group. In some embodiments, R 3 It is a methyl group.

[0163] In some implementation schemes, R 1’ It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1’ It is hydrogen. In some implementations, R 1’ It is a C1-C6 alkyl group.

[0164] In some implementation schemes, R 2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2’ It is hydrogen. In some implementations, R 2’ It is a C1-C6 alkyl group.

[0165] In some implementation schemes, R 3’ It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3’ It is hydrogen. In some implementations, R 3’ It is a C1-C6 alkyl group. In some embodiments, R 3’ It is a methyl group.

[0166] In some implementations, Z is -O- or -NH. In some implementations, Z is -O-. In some implementations, Z is -NH.

[0167] In some implementation schemes, R 8 It is a C1-C6 alkyl group, a divalent metal, or a symmetrical or asymmetric disulfide. In some embodiments, R 8 It is a C1-C6 alkyl group. In some embodiments, R 8 It is a divalent metal. In some implementations, R 8 It is cadmium (II). In some implementations, R 8It is a symmetrical disulfide or an asymmetrical disulfide. In some embodiments, R 8 It is a symmetrical disulfide.

[0168] In some embodiments, the crosslinking portion is ethylene glycol dimethacrylate (EGDMA). In some embodiments, the crosslinking portion is ethylene glycol dimethacrylamide. In some embodiments, the crosslinking portion is ethylene glycol diacrylate. In some embodiments, the crosslinking portion is ethylene glycol diacrylamide.

[0169] In some embodiments, the crosslinked portion comprises a structure represented by the following:

[0170] In some implementation schemes, R 1 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1 It is hydrogen. In some implementations, R 1 It is a C1-C6 alkyl group.

[0171] In some implementation schemes, R 2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2 It is hydrogen. In some implementations, R 2 It is a C1-C6 alkyl group.

[0172] In some implementation schemes, R 3 It is a hydrogen, C1-C6 alkyl, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl groups. In some embodiments, R 3 It is hydrogen. In some implementations, R 3 It is a C1-C6 alkyl group. In some embodiments, R 3 It is a methyl group.

[0173] In some implementation schemes, R 4 It is a C1-C4 alkylene group. In some embodiments, the crosslinking portion is divinylbenzene.

[0174] In some embodiments, the macromolecular structures provided herein include a tethering moiety that surfaces-couples the macromolecular chain. In some embodiments, the tethering moiety is a C1-C6 chain optionally substituted with one or more C1-C6 alkyl, oxo, halogenated, or hydroxyl groups. 20 Heteroalkyl groups.

[0175] In some implementations, the tethering portion includes structures:

[0176] In some implementations, the tethering portion includes structures:

[0177] In some implementations, the tethering portion is optionally secured by one or more C1-C 20 Heteroalkyl-substituted C1-C 12 Alkoxy, C1-C 12 Alkoxy and C1-C 20 Each of the heteroalkyl groups is optionally substituted with one or more C1-C6 alkyl, oxo, halogenated, or hydroxyl groups. In some embodiments, the tethering moiety is represented by the following structure:

[0178] In some implementations, Y is C1-C 20 Heteroalkyl. In some embodiments, Y is a C1-C6 alkyl group optionally substituted with one or more C1-C6 alkyl, oxo, halogenated, or hydroxyl groups. 20 Heteroalkyl groups.

[0179] In some implementations, p is an integer from 1 to 12. In some implementations, p is an integer from 1 to 6. In some implementations, p is an integer from 1 to 3. In some implementations, p is 3. In some implementations, the tethering portion is represented by the following structure:

[0180] In some implementations, the tethering portion is represented by the following structure:

[0181] In any of the macromolecular structures presented in this article, R 5 Or R 7 The product may include polyethylene glycol. In some embodiments, polyethylene glycol may have a chain length of about 1 to about 50. In some embodiments, polyethylene glycol may have a chain length of no more than 100. In some embodiments, polyethylene glycol may have a chain length of at least 5. In some embodiments, polyethylene glycol may have a chain length of about 5 to about 100 or about 5 to about 50. In some embodiments, polyethylene glycol may have a chain length of about 9.

[0182] In some embodiments, the polyethylene glycol provided herein is oligomeric polyethylene glycol. In some embodiments, the polyethylene glycol is diethylene glycol. In some embodiments, the polyethylene glycol is triethylene glycol. In some embodiments, the polyethylene glycol is tetraethylene glycol.

[0183] In some implementations, the macromolecular structures provided herein may have the structures shown in Table 3.

[0184] Table 3

[0185] In some embodiments, “b” in any of the structures in Table 3 indicates a block copolymer structure. In some embodiments, the block copolymer structures depicted in Table 3 may instead be random copolymers.

[0186] In any macromolecular structure provided herein, the macromolecular chain comprises from 1 to 1000 repeating units. In some embodiments, the macromolecular chain comprises at least 1, at least 10, at least 50, at least 100, at least 250, at least 500, at least 750, or at least 1000 repeating units. In some embodiments, the macromolecular chain comprises up to 2500 repeating units. In some embodiments, the macromolecular chain comprises up to 1000 repeating units. In some embodiments, the macromolecular chain comprises up to 750 repeating units. In some embodiments, the macromolecular chain comprises up to 500 repeating units. In some embodiments, the macromolecular chain comprises up to 250 repeating units. In some embodiments, the macromolecular chain comprises up to 100 repeating units. In some embodiments, the macromolecular chain comprises about 1 to about 100 repeating units. In some embodiments, the macromolecular chain comprises about 1 to about 250 repeating units. In some embodiments, the macromolecular chain comprises about 1 to about 500 repeating units. In some embodiments, the macromolecular chain comprises about 1 to about 1,000 repeating units. In some embodiments, the macromolecular chain comprises about 100 to about 1,000 repeating units. In some embodiments, the macromolecular chain comprises about 1 to about 10 repeating units.

[0187] In any of the macromolecular structures provided herein, the macromolecular chain has a molecular weight of about 0.1 kDa to about 500 kDa. In some embodiments, the macromolecular chain has a molecular weight of at least 0.1 kDa, at least 1 kDa, at least 5 kDa, at least 10 kDa, at least 20 kDa, at least 25 kDa, at least 50 kDa, at least 100 kDa, at least 250 kDa, or at least 500 kDa. In some embodiments, the macromolecular chain has a molecular weight of no more than 1000 kDa, no more than 500 kDa, no more than 750 kDa, no more than 500 kDa, no more than 250 kDa, no more than 100 kDa, no more than 75 kDa, no more than 50 kDa, no more than 40 kDa, no more than 30 kDa, no more than 25 kDa, no more than 20 kDa, no more than 15 kDa, or no more than 10 kDa. In some embodiments, the macromolecular chain has a molecular weight of about 0.1 to about 500 kDa, about 0.1 to about 250 kDa, about 0.1 kDa to about 100 kDa, about 0.1 kDa to about 70 kDa, 0.5 kDa to about 10 kDa, 0.5 kDa to about 15 kDa, or about 1 kDa to about 25 kDa. In some embodiments, the macromolecular chain has a molecular weight of about 0.1 kDa to about 100 kDa. In some embodiments, the macromolecular chain has a molecular weight of about 0.1 kDa to about 50 kDa.

[0188] In some embodiments, the macromolecular chain comprises a block copolymer. In some embodiments, the block copolymer comprises a first block derived from a first monomer and a second block derived from a second monomer, wherein the first block is adjacent to the tethered portion, and wherein the first monomer is more hydrophobic than the second monomer. In some embodiments, the block copolymer comprises a first block derived from a first monomer and a second block derived from a second monomer, wherein the first block is adjacent to the tethered portion, and wherein the first monomer is less hydrophobic than the second monomer. In some embodiments, the hydrophobicity of the first and second monomers can be determined by using estimated partition coefficients from XLOGP3. In some embodiments, the absolute difference between the estimated partition coefficients of the first and second monomers is at least 0.3, at least 0.5, at least 0.8, at least 1, at least 1.5, or at least 2. In some embodiments, the absolute difference between the estimated partition coefficients of the first and second monomers is no more than 3, no more than 2.5, no more than 2, no more than 1, or no more than 0.8.

[0189] In some embodiments, the surface is particulate. In some embodiments, the particulate is a nanoparticle or microparticle. In some embodiments, the particulate is a nanoparticle. In some embodiments, the particulate is a microparticle. In some cases, the particulates provided herein have a diameter of at least 10 nm, at least 100 nm, at least 200 nm, at least 300 nm, at least 400 nm, at least 500 nm, at least 600 nm, at least 700 nm, at least 800 nm, or at least 900 nm. In some embodiments, the particulates provided herein have a diameter of no more than 5000 nm, no more than 4000 nm, no more than 3000 nm, no more than 2000 nm, no more than 1000 nm, no more than 750 nm, or no more than 500 nm. In some embodiments, the particles provided herein have a range of wavelengths from 10 nm to 50 nm, from 50 nm to 100 nm, from 100 nm to 150 nm, from 150 nm to 200 nm, from 200 nm to 250 nm, from 250 nm to 300 nm, from 300 nm to 350 nm, from 350 nm to 400 nm, from 400 nm to 450 nm, from 450 nm to 500 nm, from 500 nm to 550 nm, from 550 nm to 600 nm, from 600 nm to 650 nm, from 650 nm to 700 nm, from 700 nm to 750 nm, from 750 nm to 800 nm, from 800 nm to 850 nm, from 850 nm to 900 nm, from 100 nm to 300 nm, from 150 nm to 350 nm, from 200 nm to 400 nm, and from 250 nm to 450 nm. Diameters ranging from 300 nm to 500 nm, 350 nm to 550 nm, 400 nm to 600 nm, 450 nm to 650 nm, 500 nm to 700 nm, 550 nm to 750 nm, 600 nm to 800 nm, 650 nm to 850 nm, 700 nm to 900 nm, or 10 nm to 900 nm. Particle size (e.g., diameter) can be measured as an indirect measure of size by dynamic light scattering (DLS). DLS measurements can be “intensity-weighted” averages, meaning that the size distribution calculated from the average can be weighted by the sixth power of the radius. This may refer to, as in this document, “z-average” or “intensity average”. Particle size can also be measured by electron microscopy (e.g., SEM, TEM).

[0190] In some examples, the particles provided herein may have a diameter of about 100 nm to about 500 nm. In some embodiments, the particles have a diameter of about 100 nm to about 300 nm. In some embodiments, the particles have a diameter of about 100 nm to about 200 nm. In some embodiments, the particles have a diameter of about 150 nm to about 250 nm.

[0191] Furthermore, the particles can have a uniform or non-uniform size distribution. The polydispersity index (PDI) is a measure of size distribution and can be measured using techniques such as dynamic light scattering. A low PDI indicates a more uniform size distribution, while a higher PDI indicates a more non-uniform size distribution. For example, the particles disclosed herein may have a PDI of less than 0.5, less than 0.4, less than 0.3, less than 0.2, less than 0.15, or less than 0.1. In certain embodiments, the particles disclosed herein have a PDI of less than 0.1. In some embodiments, the particles disclosed herein have a PDI of more than 0.5, more than 1, or more than 2.

[0192] The particles described in this article can have different surface charge ranges. The particles can be negatively charged, positively charged, or neutrally charged. In some embodiments, the particles have values ​​from -150 mV to -100 mV, -100 mV to -90 mV, -90 mV to -80 mV, -80 mV to -70 mV, -70 mV to -60 mV, -60 mV to -50 mV, -50 mV to -40 mV, -40 mV to -30 mV, -30 mV to -20 mV, -20 mV to -10 mV, -10 mV to 0 mV, 0 mV to 10 mV, 10 mV to 20 mV, 20 mV to 30 mV, 30 mV to 40 mV, 40 mV to 50 mV, 50 mV to 60 mV, 60 mV to 70 mV, 70 mV to 80 mV, 80 mV to 90 mV, 90 mV to 100 mV, 100 mV to 100 mV. Surface charges range from mV to 110 mV, 110 mV to 120 mV, 120 mV to 130 mV, 130 mV to 140 mV, or 140 mV to 150 mV. In some embodiments, the particles have a surface charge of about 0 to -100 mV. In some embodiments, the particles have a surface charge of about 0 to 100 mV. In some embodiments, the charge can be determined by measuring the zeta potential at neutral pH using a suitable buffer.

[0193] Various particle morphologies are consistent with the particle types disclosed herein. For example, particles can be spherical, colloidal, square, rod-shaped, linear, conical, pyramidal, or rectangular.

[0194] In some embodiments, the surface (e.g., particles) comprises any suitable material according to those skilled in the art. In some embodiments, the particles are magnetic, such as any suitable magnetic material according to those skilled in the art. In some embodiments, the particles comprise a metallic material. In some embodiments, the metallic material comprises any one or any combination of gold, silver, copper, nickel, cobalt, palladium, platinum, iridium, osmium, rhodium, ruthenium, rhenium, vanadium, chromium, manganese, niobium, molybdenum, tungsten, tantalum, iron, and cadmium. In some embodiments, the particles comprise iron oxide. In some embodiments, the particles are superparamagnetic iron oxide particles. In some embodiments, the particles have a core-shell structure. In some embodiments, the particles have an iron oxide core. In some embodiments, the particles comprise a silica shell. In some embodiments, the particles comprise an iron oxide core and a silica shell. In some cases, the silica shell may be functionalized with tethered portions or macromolecular chains provided elsewhere herein. In some embodiments, the particles comprise iron oxide crystals. In some embodiments, the particles comprise polystyrene. In some embodiments, the particles comprise iron oxide crystals embedded in a polystyrene core.

[0195] In some embodiments, the macromolecular structure provided herein comprises at least 5% w / w of the repeating units provided herein. In some embodiments, the macromolecular structure comprises at least 10% w / w, at least 15% w / w, at least 20% w / w, at least 25% w / w, at least 30% w / w, at least 35% w / w, at least 40% w / w, at least 45% w / w, at least 50% w / w, at least 60% w / w, at least 75% w / w, at least 90% w / w, at least 95% w / w, at least 99% w / w, or about 100% w / w of the repeating units provided herein. In some embodiments, the macromolecular structure comprises up to 95% w / w, up to 90% w / w, up to 85% w / w, up to 75% w / w, up to 70% w / w, up to 65% w / w, up to 60% w / w, up to 55% w / w, up to 50% w / w, up to 45% w / w, up to 40% w / w, up to 35% w / w, or up to 30% w / w of the repeating units provided herein. In some embodiments, the macromolecular structure comprises about 5% to about 95% w / w of the repeating units provided herein. In some embodiments, the macromolecular structure comprises about 5% to about 75% w / w of the repeating units provided herein. In some embodiments, the macromolecular structure comprises about 10% to about 50% w / w of the repeating units provided herein. In some embodiments, the macromolecular structure comprises at least 10% w / w of repeating units. In some embodiments, the macromolecular structure comprises up to 50% w / w of repeating units. The weight percentage of repeating units in a macromolecular structure can be determined by thermogravimetric analysis (TGA).

[0196] In some embodiments, the surface provided herein comprises tethered units (e.g., and macromolecular chains) at a suitable density according to those skilled in the art. In some embodiments, the surface comprises at a density of 500 nm. 2 At least one tethering unit (e.g., and macromolecular chains) at a density of [number missing]. In some embodiments, the surface contains [number missing] units per 50 nm. 2 At least one tethering unit (e.g., and a macromolecular chain) at a density of [number missing]. In some embodiments, the surface contains [number missing] units per 5 nm [units missing]. 2 At least one tethering unit (e.g., and a macromolecular chain) at a density of [number missing]. In some embodiments, the surface contains [number missing] units per 1 nm. 2 At least one tethering unit (e.g., and macromolecular chains) at a density of [number missing]. In some embodiments, the surface contains [number missing] units per 50 nm. 2 Approximately 1 to 5nm 2 A density of approximately 1 tethered unit (e.g., and a macromolecular chain).

[0197] Preparation method This document provides methods for preparing any of the macromolecular structures described herein. In some embodiments, this document provides methods for preparing macromolecular structures comprising a surface, a tethered portion, and a macromolecular structure. This document also provides methods for preparing macromolecular structures comprising a surface and a macromolecular chain of formula (I).

[0198] In some implementations, the methods provided herein include surface-initiated polymerization.

[0199] In some embodiments, the macromolecular structure provided herein further includes a terminating group. In some embodiments, the terminating group is a halogen, epoxide, or alkylene (e.g., an olefin). In some embodiments, the terminating group is a halogen. In some embodiments, the terminating group is bromine, chlorine, or iodine. In some embodiments, the terminating group is bromine. In some embodiments, the terminating group participates in surface-initiated polymerization.

[0200] In some embodiments, this document provides methods for preparing macromolecular structures such as any macromolecular structures provided elsewhere herein. In some embodiments, the method includes providing a surface. In some embodiments, the method further includes coupling a polymeric initiator to the surface to form an initiator surface. In some embodiments, the method further includes contacting the initiator surface with a monomer, such as a monomer provided elsewhere herein, to form a macromolecular structure.

[0201] In some cases, such as the exemplary synthesis schemes described by the methods provided herein, Figure 1 Provided by China.

[0202] In some cases, the methods provided herein can also provide block copolymers, such as by contacting a second repeating unit with an initiator surface provided by a first repeating unit. Exemplary synthetic schemes of this kind are detailed in [the document / section]. Figure 7 It is displayed in the middle.

[0203] In some cases, the methods provided herein involve contacting an initiator surface with more than one (different) monomer, such as to form a hybrid polymer. The monomers can be organized in a controlled or random distribution. Exemplary synthetic schemes detailing this are described in [the following text is missing from the original extract]. Figure 8 It is displayed in the middle.

[0204] In some embodiments, the method includes providing a surface. The surface may include any surface (e.g., particles) as described elsewhere herein. In some embodiments, the surface comprises particles. In some embodiments, the particles are nanoparticles. In some embodiments, the particles are microparticles. In some embodiments, the surface comprises silica. In some embodiments, the surface is amine-functionalized, such as by functionalization with (3-aminopropyl)triethoxysilane (APTES). In some embodiments, the surface is epoxide-functionalized—using (3-glycidyloxypropyl)triethoxysilane followed by reaction with diaminohexane (…). Figure 6 Skilled technicians will understand that various silane coupling agents can be used to functionalize surfaces, and, if appropriate, further modify them to achieve a variety of chain lengths and functionalities. Non-limiting examples of silane coupling agents include 3-aminopropyltriethoxysilane (APTES), 3-mercaptopropyltrimethoxysilane, 3-epoxypropoxypropyltrimethoxysilane (GPTMS), (3-triethoxysilylpropyl)diethylenetriamine (DETAS), N-(2-aminoethyl)-3-aminopropyltrimethoxysilane, 3-isocyanopropyltriethoxysilane, 3-chloropropyltrimethoxysilane, 3-bromopropyltrimethoxysilane, 3-iodopropyltrimethoxysilane, 3-[2-(2-aminoethylamino)ethylamino]propyl-dimethoxymethylsilane, 3-[2-(2-aminoethylamino)ethylaminopropyl]trimethoxysilane, propargyltriethoxysilane, 3-glycidyloxypropylheptamethyltrisiloxane, 3-(2-(2-propargyloxy)ethoxy)propyltriethoxysilane, etc.

[0205] In some embodiments, the functionalization of a surface (e.g., a silica surface) is described in Example 1, such as in which the silica surface is functionalized by APTES via amine coupling with a polymer initiator followed by polymerization.

[0206] In some embodiments, the method further includes coupling a polymer initiator to a surface to form an initiator surface. In some embodiments, coupling is accomplished using an organic solvent, such organic solvents are provided elsewhere herein. In some embodiments, the surface (e.g., particles) is contacted with or dispersed in an organic solvent prior to coupling. In some embodiments, the organic solvent comprises dimethylformamide. In some embodiments, the organic solvent comprises tetrahydrofuran. In some embodiments, the organic solvent comprises N,N-dimethylacetamide.

[0207] In some embodiments, coupling further includes the addition of a base. In some embodiments, the base includes a weak base. In some embodiments, the base is an amine. In some embodiments, the base is an alkylamine. In some embodiments, the base is triethylamine.

[0208] In some embodiments, coupling is performed below room temperature. In some embodiments, coupling is performed at temperatures below 30°C. In some embodiments, coupling is performed at temperatures not exceeding 30°C, not exceeding 25°C, not exceeding 20°C, not exceeding 15°C, not exceeding 10°C, not exceeding 5°C, or not exceeding 0°C. In some embodiments, coupling is performed above the freezing point of the solvent. In some embodiments, coupling is performed at temperatures from about 0°C to about 30°C. In some embodiments, coupling is performed at temperatures from about 0°C to about 15°C. In some embodiments, coupling includes temperatures from about -5°C to about 5°C. In some embodiments, coupling is performed at temperatures from about 0°C. In some embodiments, coupling is performed in an ice bath. In some cases, the reduced temperature is maintained when adding a base and a polymer initiator.

[0209] In some implementations, the polymer initiator is an oxygen-substituted C1-C8 haloalkyl group.

[0210] In some embodiments, the polymer initiator is represented by the following structure:

[0211] In some embodiments, X is a halogen. In some embodiments, X is bromine. In some embodiments, X is chlorine.

[0212] In some implementation schemes, R 10 It is an initiator group. In some embodiments, the initiator group is a halogen. In some embodiments, the initiator group is bromine.

[0213] In some implementations, the polymer initiator is 2-bromoisobutyryl bromide.

[0214] In some embodiments, the coupling step occurs for any suitable period of time as known to those skilled in the art, thus obtaining a suitable yield of the product. In some embodiments, the coupling step occurs for at least 0.5 hr, 1 hr, 2 hr, 4 hr, 6 hr, 8 hr, 12 hr, or 16 hr. In some embodiments, the coupling step occurs for up to 36 hr, 24 hr, 20 hr, 18 hr, or 16 hr. In some embodiments, the coupling step occurs for about 0.5 hr to about 36 hr or for about 12 hr to about 20 hr. In some embodiments, the coupling step occurs for about 16 hr, such as at 0°C.

[0215] In some implementations, the coupling step is performed under inert conditions. In some implementations, the coupling step is performed under N2 conditions.

[0216] In some embodiments, the coupling step further includes a washing step, such as after the reaction. In some embodiments, washing includes washing with one or more organic solvents or an aqueous solvent. In some embodiments, washing includes washing with both an organic solvent and an aqueous solvent, respectively.

[0217] In some embodiments, non-limiting examples of the organic solvents provided herein include ethanol, methanol, isopropanol, butanol, dimethyl sulfoxide, dimethylformamide, hexane, pentane, benzene, acetonitrile, acetone, carbon tetrachloride, chloroform, N,N-dimethylacetamide, cyclohexane, diethylene glycol, diethyl ether, ethyl acetate, and tetrahydrofuran. In some embodiments, the organic solvent is tetrahydrofuran. In some embodiments, the organic solvent is dimethylformamide. In some embodiments, the organic solvent is N,N-dimethylacetamide. In some embodiments, the organic solvents provided herein comprise combinations of organic solvents. In some embodiments, the organic solvents provided herein comprise mixtures of organic solvents and water.

[0218] In some implementations, examples of the coupling step are described in Example 2.

[0219] In some embodiments, the method includes contacting the surface of an initiator with a monomer to form a macromolecular structure. In some cases, the monomer includes any monomer provided elsewhere herein.

[0220] In some implementations, contact is performed under inert conditions. In some implementations, contact is performed under N2.

[0221] In some embodiments, the method includes dissolving the monomers separately in a solvent before adding them to the surface dispersion. In some embodiments, the solvent includes an organic solvent, such as dimethylformamide.

[0222] In some embodiments, the contact includes ultrasonic treatment. In some embodiments, the solution is ultrasonicated for at least 1 minute, at least 5 minutes, at least 10 minutes, or at least 15 minutes. In some embodiments, the solution is ultrasonicated for 15 minutes.

[0223] In some embodiments, the contact also includes the addition of a reducing agent. In some embodiments, the reducing agent is L-ascorbic acid. In some embodiments, the reducing agent is added after the monomer is added to the solution. In some embodiments, the reducing agent is added at a specific rate, such as by a syringe pump. In some embodiments, the reducing agent is added at a rate in the range of about 0.05 mL / min. In some embodiments, the reducing agent is added at a rate of about 0.01 mL / min to about 0.5 mL / min.

[0224] In some embodiments, the contact includes any suitable temperature according to those skilled in the art. In some embodiments, the contact includes a temperature of at least 25°C. In some embodiments, the contact includes a temperature of at least 30°C, 40°C, 50°C, 60°C, or 70°C. In some embodiments, the contact includes a temperature at or below the solvent's boiling point but above room temperature. In some embodiments, the contact includes a temperature of about 25°C to about 75°C. In some embodiments, the contact includes a temperature of about 35°C.

[0225] In some embodiments, the contact is carried out in an organic solvent, such as those described elsewhere herein. In some embodiments, the organic solvent is dimethylformamide (DMF), ethanol, methanol, isopropanol, dimethyl sulfoxide (DMSO), or a combination thereof or an aqueous mixture thereof. In some embodiments, the contact is carried out in water. In some embodiments, the contact is carried out in dimethylformamide (DMF).

[0226] In some embodiments, this document provides methods for preparing macromolecular structures. In some embodiments, the method includes providing a surface. In some embodiments, the method includes coupling vinyl groups to the surface to form a vinyl-functionalized surface. In some embodiments, the method includes contacting the vinyl-functionalized surface with a crosslinking monomer and a monomer selected from hydroxyalkyl methacrylate, aminoalkyl methacrylate, alkynyl methacrylate, glycidyl methacrylate, hydroxyalkyl acrylate, aminoalkyl acrylate, alkynyl acrylate, or glycidyl acrylate to form a crosslinked polymer coupled to the surface. In some embodiments, the method further includes coupling a polymer initiator to the crosslinked polymer to form an initiator surface. In some embodiments, the method includes contacting the initiator surface with a monomer, such as a monomer provided elsewhere herein, to form a macromolecular structure.

[0227] In some cases, such as the exemplary synthesis schemes described by the methods provided herein, Figure 2 Provided in [the text]. Vinyl-functionalized surfaces undergo polymerization in the presence of a diallyl amide crosslinking agent and hydroxyalkyl methacrylate monomers. A polymer initiator is coupled to the hydroxyalkyl side chains of the crosslinked polymer. A second polymerization is carried out to incorporate ethylene glycol methacrylate monomers.

[0228] In some embodiments, the method includes providing a surface, such as the surface described elsewhere herein. In some embodiments, the surface is washed with a solvent, such as an organic solvent, prior to subsequent steps. In some embodiments, the surface comprises silica.

[0229] In some embodiments, the method includes coupling vinyl groups to a surface to form a vinyl-functionalized surface. In some embodiments, coupling includes dispersing the surface in a solvent. In some embodiments, the solvent is an organic solvent. In some embodiments, the solvent is dimethylformamide. In some embodiments, dispersion is accomplished by ultrasonic treatment. In some embodiments, vinyl coupling is performed under inert conditions. In some embodiments, vinyl coupling is performed under N2. In some embodiments, vinyl coupling is performed under elevated temperatures. In some embodiments, vinyl coupling involves temperatures of at least 80°C. In some embodiments, vinyl coupling involves temperatures of at least 90°C, 100°C, 110°C, or 120°C. In some embodiments, vinyl coupling includes heating to at most 150°C, at most 140°C, at most 130°C, or at most 120°C. In some embodiments, vinyl coupling includes heating at a temperature of about 120°C. In some embodiments, coupling further includes a washing step, such as washing with an organic solvent (e.g., DMF).

[0230] In some embodiments, the method further includes contacting the vinyl-functionalized surface with a crosslinking monomer and a monomer selected from hydroxyalkyl methacrylate, aminoalkyl methacrylate, alkynyl methacrylate, glycidyl methacrylate, hydroxyalkyl acrylate, aminoalkyl acrylate, alkynyl acrylate, or glycidyl acrylate to form a crosslinked polymer coupled to the surface. In some embodiments, the monomer is hydroxyalkyl methacrylate. In some embodiments, the monomer is aminoalkyl methacrylate. In some embodiments, the monomer is alkynyl methacrylate. In some embodiments, the monomer is glycidyl methacrylate. In some embodiments, the monomer is hydroxyalkyl acrylate. In some embodiments, the monomer is aminoalkyl acrylate. In some embodiments, the monomer is alkynyl acrylate. In some embodiments, the monomer is glycidyl acrylate. In some embodiments, the contact includes contacts as described elsewhere herein. In some embodiments, the vinyl-functionalized surface comprises vinyl acrylate.

[0231] In some embodiments, the crosslinking monomer is a diene. In some embodiments, the crosslinking monomer is ethylene glycol dimethacrylate (EGDMA). In some embodiments, the crosslinking monomer is divinylbenzene.

[0232] In some embodiments, the crosslinking monomer comprises a structure represented by the following:

[0233] In some implementation schemes, R 1 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1 It is hydrogen. In some implementations, R 1 It is a C1-C6 alkyl group.

[0234] In some implementation schemes, R 2 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2 It is hydrogen. In some implementations, R 2 It is a C1-C6 alkyl group.

[0235] In some implementation schemes, R 3 It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 3 It is hydrogen. In some implementations, R 3 It is a C1-C6 alkyl group. In some embodiments, R 3 It is a methyl group.

[0236] In some implementation schemes, R 1’ It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 1’It is hydrogen. In some implementations, R 1’ It is a C1-C6 alkyl group.

[0237] In some implementation schemes, R 2’ It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 2’ It is hydrogen. In some implementations, R 2’ It is a C1-C6 alkyl group.

[0238] In some implementation schemes, R 3’ It is hydrogen or a C1-C6 alkyl group. In some embodiments, R 3’ It is hydrogen. In some implementations, R 3’ It is a C1-C6 alkyl group. In some embodiments, R 3’ It is a methyl group.

[0239] In some embodiments, the method further includes coupling a polymeric initiator to a crosslinked polymer to form an initiator surface. In some embodiments, the coupling is described elsewhere herein. In some embodiments, the polymeric initiator is a polymeric initiator as described elsewhere herein.

[0240] In some embodiments, the methods provided herein include polymerization. In some embodiments, the methods provided herein include surface-initiated polymerization. In some embodiments, the contact initiator surface includes polymerization. In some embodiments, the polymerization includes free radical polymerization. In some embodiments, the polymerization includes "living" / controlled free radical polymerization.

[0241] In some implementations, the methods provided herein can provide any of the macromolecular structures in Table 3.

[0242] Composition In some embodiments, compositions comprising any of the macromolecular structures provided herein are provided. In some embodiments, compositions comprising a macromolecular structure and a biomolecule, the macromolecular structure being such as those provided elsewhere herein (e.g., comprising a surface, tethering portion, and macromolecular chain). In some embodiments, a biomolecule is adsorbed onto the macromolecular structure. In some embodiments, a composition comprising one or more macromolecular structures and a biomolecule adsorbed onto the macromolecular structure, the macromolecular structure being such as those provided elsewhere herein (e.g., comprising a surface, tethering portion, and macromolecular chain).

[0243] In some implementations, biomolecules adsorbed onto macromolecular structures form biomolecular crowns on the macromolecular structures.

[0244] In some embodiments, the compositions provided herein further comprise a biological sample in contact with a macromolecular structure. In some embodiments, the biological sample comprises multiple proteins. In some embodiments, the biological sample comprises plasma, serum, urine, cerebrospinal fluid, synovial fluid, tears, saliva, whole blood, breast milk, nipple aspirate, catheter lavage fluid, vaginal fluid, nasal fluid, ear fluid, gastric juice, pancreatic juice, trabecular fluid, bronchoalveolar lavage fluid, sweat, gingival crevicular fluid, semen, prostatic fluid, sputum, excrement, bronchial lavage fluid, fluid from a swab, bronchial aspirate, fluidized solid, fine needle aspirate sample, tissue homogenate, lymph, cell culture sample, or any combination thereof. In some embodiments, the biological sample comprises plasma, serum, or blood. In some embodiments, the biological sample comprises blood. In some embodiments, the biological sample comprises plasma. In some embodiments, the biological sample comprises serum. In some embodiments, the biological sample is a biofluid. In some embodiments, the biological sample is a cell-free sample.

[0245] In some embodiments, multiple biomolecules may be adsorbed onto the macromolecular structure provided herein. In some embodiments, multiple different biomolecules may be adsorbed onto the macromolecular structure provided herein. In some embodiments, at least 5, at least 10, at least 20, at least 40, at least 60, at least 80, at least 100, at least 200, at least 400, at least 600, at least 800, at least 1000, or at least 2000 (e.g., different) biomolecules are adsorbed onto the macromolecular structure provided herein. In some embodiments, up to 5000, up to 4000, up to 3000, up to 2000, up to 1000, up to 1000, up to 800, up to 600, up to 400, up to 200, up to 100, up to 60, or up to 20 (e.g., different) biomolecules are adsorbed onto the macromolecular structure provided herein. In some embodiments, about 5 to about 5000, about 10 to about 2000, about 100 to about 2000, or about 100 to about 1000 (e.g., different) biomolecules are adsorbed onto the macromolecular structure provided herein. In some embodiments, at least 100 (e.g., different) biomolecules are adsorbed onto the macromolecular structure provided herein.

[0246] In some embodiments, the biomolecule is a protein, polypeptide, polysaccharide, sugar, lipid, lipoprotein, metabolite, oligonucleotide, or metabolome. In some embodiments, the biomolecule is a protein. In some embodiments, the biomolecule is a polypeptide. In some embodiments, the biomolecule is a polysaccharide. In some embodiments, the biomolecule is a sugar. In some embodiments, the biomolecule is a lipid. In some embodiments, the biomolecule is a lipoprotein. In some embodiments, the biomolecule is a metabolite. In some embodiments, the biomolecule is an oligonucleotide. In some embodiments, the biomolecule is a metabolome.

[0247] In some embodiments, compositions comprising a macromolecular structure and a protein are provided herein, such as the macromolecular structures provided herein (e.g., comprising a surface, tethering portions, and macromolecular chains). In some embodiments, compositions comprising one or more macromolecular structures and a protein are provided herein, such as the macromolecular structures provided herein.

[0248] In some embodiments, multiple proteins can be adsorbed onto the macromolecular structure provided herein. In some embodiments, multiple different proteins can be adsorbed onto the macromolecular structure provided herein. In some embodiments, at least 5, at least 10, at least 20, at least 40, at least 60, at least 80, at least 100, at least 200, at least 400, at least 600, at least 800, at least 1000, or at least 2000 (e.g., different) proteins are adsorbed onto the macromolecular structure provided herein. In some embodiments, up to 5000, up to 4000, up to 3000, up to 2000, up to 1000, up to 1000, up to 800, up to 600, up to 400, up to 200, up to 100, up to 60, or up to 20 (e.g., different) proteins are adsorbed onto the macromolecular structure provided herein. In some embodiments, about 5 to about 5000, about 10 to about 2000, about 100 to about 2000, or about 100 to about 1000 (e.g., different) proteins are adsorbed onto the macromolecular structure provided herein. In some embodiments, at least 100 (e.g., different) proteins are adsorbed onto the macromolecular structure provided herein.

[0249] Methods for protein identification This document provides a method for identifying proteins in a sample using macromolecular structures (e.g., those comprising surfaces, tethered portions, and macromolecular chains) as described elsewhere herein. In some embodiments, the method includes incubating one or more macromolecular structures together with a biological sample containing biomolecules. In some embodiments, incubating the resulting one or more macromolecular structures together with a biological sample containing biomolecules results in the formation of a biomolecular crown. In some embodiments, the method includes isolating at least a portion of the biomolecules within the biomolecular crown. In some embodiments, the method includes determining the biomolecular crown. In some embodiments, the biomolecules are as described elsewhere herein. In some embodiments, the biomolecules are proteins.

[0250] In some embodiments, the biomolecular crown assay can identify 1 to 50,000 protein groups or proteins. In some embodiments, 1 to 20,000 protein groups or proteins can be identified. In some embodiments, at least 100 protein groups can be identified. In some embodiments, at least 300 protein groups can be identified. In some embodiments, at least 500 protein groups can be identified. In some embodiments, at least 1,000 protein groups can be identified. In some embodiments, 1,000 to 10,000 protein groups or proteins can be identified. In some embodiments, 1,000 to 5,000 protein groups or proteins can be identified. In some embodiments, 1,800 to 5,000 protein groups or proteins can be identified. In some embodiments, 1,200 to 2,200 protein groups or proteins can be identified. In some embodiments, the protein groups or proteins may comprise peptide sequences of a minimum length of 2 amino acid residues. In some embodiments, the protein groups may comprise peptide sequences of a minimum length of 2 amino acid residues. In some embodiments, the proteome may comprise a peptide sequence of minimum length of 5 amino acid residues. In some embodiments, the proteome may comprise a peptide sequence of minimum length of 7 amino acid residues. In some embodiments, the proteome may comprise a peptide sequence of minimum length of 8 amino acid residues. In some embodiments, the proteome may comprise a peptide sequence of minimum length of 9 amino acid residues. In some embodiments, the proteome may comprise a peptide sequence of minimum length of 10 amino acid residues. In some embodiments, the method may further comprise cleaving the protein with the unique biomolecular crown. In some embodiments, the method may further comprise digesting the protein with the unique biomolecular crown. In some embodiments, the digested protein may be purified.

[0251] In some embodiments, the method further includes repeating the method described herein, wherein when repeated, incubation, isolation, and assay produce a percentage quantile normalized coefficient of variation (QNCV) of 30% or less, as determined by comparing peptide mass spectrometry characteristics of at least three complete assay replicates from each of one or more surfaces. In some embodiments, when repeated, incubation, isolation, and assay produce a percentage quantile normalized coefficient of variation (QNCV) of 25% or less, as determined by comparing peptide mass spectrometry characteristics of at least three complete assay replicates from each of one or more surfaces. In some embodiments, when repeated, incubation, isolation, and assay produce a percentage quantile normalized coefficient of variation (QNCV) of 20% or less, as determined by comparing peptide mass spectrometry characteristics of at least three complete assay replicates from each of one or more surfaces. In some embodiments, the assay is capable of identifying proteins within a dynamic range of at least 7, at least 8, at least 9, or at least 10. In some implementations, the assay can identify proteins within a dynamic range of no more than 12, no more than 11, no more than 10, no more than 9, no more than 8, or no more than 7.

[0252] In some embodiments, the method may further include washing one or more surfaces at least once after isolating them from unbound proteins. In some embodiments, the method may further include washing one or more surfaces at least twice after isolating them from unbound proteins. In some embodiments, the method may further include washing one or more surfaces at least three times after isolating them from unbound proteins. Isolation may be performed, for example, using magnetic isolation or centrifugation.

[0253] In some embodiments, the method may further include desorbing the proteins of the biomolecular crown. In some embodiments, the method may further include denaturing the proteins of the biomolecular crown.

[0254] In some embodiments, the methods herein further include the preparation of an analyte (e.g., protein) from the biomolecular crown for analysis. In some embodiments, the preparation for analysis includes digesting a subset of biomolecules within the biomolecular crown, a protein crown, or biomolecules desorbed from the biomolecular crown to form a digested sample. The preparation of an analyte from the biomolecular crown for analysis may also include chemically modifying the biomolecule from the biomolecular crown, such as methylating or reducing the biomolecule. In some embodiments, the preparation of an analyte from the biomolecular crown for analysis may include denaturing the analyte (e.g., protein).

[0255] In some embodiments, the assay includes identifying proteins in the sample using mass spectrometry. In some embodiments, the assay includes using tandem mass spectrometry. In some embodiments, the assay includes using liquid chromatography-tandem mass spectrometry. In some embodiments, the assay may include ELISA, Edman degradation, immunoaffinity techniques, single-molecule protein sequencing, etc.

[0256] In some implementations, the determination is performed over approximately 2 to approximately 4 hours. In some implementations, the method is performed over approximately 1 to approximately 20 hours. In some implementations, the method is performed over approximately 2 to approximately 10 hours. In some implementations, the method is performed over approximately 4 to approximately 6 hours. In some implementations, the isolation takes no more than approximately 30 minutes, no more than approximately 15 minutes, no more than approximately 10 minutes, no more than approximately 5 minutes, or no more than approximately 2 minutes.

[0257] In some embodiments, multiple spatially isolated samples are processed according to the method. In some embodiments, the multiple samples include at least 10 spatially isolated samples, at least 50 spatially isolated samples, at least 100 spatially isolated samples, at least 150 spatially isolated samples, at least 200 spatially isolated samples, at least 250 spatially isolated samples, or at least 300 spatially isolated samples. In a further embodiment, the multiple samples include at least 96 samples.

[0258] In some embodiments, one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure share at least one physicochemical property and are different in at least one physicochemical property, such that the first unique macromolecular structure and the second unique macromolecular structure are distinct. The two macromolecular structures may have polymeric properties, but may exhibit different macromolecular structures, such as charge measured by zeta potential analysis. In some embodiments, one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure share at least two physicochemical properties and are different in at least two physicochemical properties, such that the first unique macromolecular structure and the second unique macromolecular structure are distinct. In some embodiments, one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure share at least one physicochemical property and are different in at least two physicochemical properties, such that the first unique macromolecular structure and the second unique macromolecular structure are distinct.

[0259] In some embodiments, one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure share at least two physicochemical properties and are different in at least one physicochemical property, such that the first unique macromolecular structure and the second unique macromolecular structure are distinct. In further embodiments, the physicochemical properties include size, charge, core material, shell material, porosity, or hydrophobicity of the macromolecular structure. In further embodiments, size is diameter or radius, as measured by dynamic light scattering, SEM, TEM, or any combination thereof.

[0260] In some embodiments, one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure have a surface charge from 0 mV to -50 mV, and wherein the first unique macromolecular structure, the second unique macromolecular structure, or both have a diameter of less than 400 nm. In some embodiments, one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure have a diameter of 100 to 400 nm, wherein the first unique macromolecular structure has a positive surface charge, and wherein the second unique macromolecular structure has a negative surface charge.

[0261] In some embodiments, one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure are nanoparticles, wherein the first unique macromolecular structure has a surface charge of less than -20 mV and the second unique macromolecular structure has a surface charge of greater than 20 mV. In some embodiments, one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure are microparticles, wherein the first unique macromolecular structure has a negative surface charge and the second unique macromolecular structure has a positive surface charge. In some embodiments, one or more macromolecular structures comprise a subset of negatively charged nanoparticles, wherein each macromolecular structure in the subset is distinct on at least one surface chemical group. In some embodiments, one or more macromolecular structures include a first unique macromolecular structure, a second unique macromolecular structure, and a third unique macromolecular structure, wherein the first unique macromolecular structure, the second unique macromolecular structure, and the third unique macromolecular structure include an iron oxide core and have a diameter of less than about 500 nm, wherein the first unique macromolecular structure has a negative charge of less than -40 mV, the second unique macromolecular structure has a positive charge of greater than 20 mV, and the third unique macromolecular structure has a negative charge of -20 mV to -40 mV.

[0262] In some embodiments, at least one of the one or more unique macromolecular structures comprises a carboxylated polymer, an amination polymer, a zwitterionic polymer, or any combination thereof. In some embodiments, at least one of the one or more unique macromolecular structures comprises a carboxylated polymer. In some embodiments, at least one of the one or more unique macromolecular structures comprises an amination polymer. In some embodiments, at least one of the one or more unique macromolecular structures comprises a zwitterionic polymer.

[0263] In some embodiments, one or more macromolecular structures include at least two unique macromolecular structures. In some embodiments, one or more macromolecular structures include at least three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, twenty, twenty-five, or thirty unique macromolecular structures. In some embodiments, one or more macromolecular structures include up to 50, 45, 35, 25, or twenty unique macromolecular structures. In some embodiments, one or more macromolecular structures include about two to about thirty unique macromolecular structures. In some implementations, one or more macromolecular structures include at least 10 unique macromolecular structures.

[0264] Analysis and Automation Systems In some embodiments, this document provides a system for identifying biomolecules in a biological sample, wherein the system may include (i) one or more macromolecular structures provided elsewhere herein; (ii) a biological sample containing biomolecules; and (iii) an automated system comprising a network of differentiated functional units for isolating biomolecules adsorbed to the macromolecular structure, and wherein the automated system is programmed to perform a series of steps.

[0265] In some embodiments, macromolecules, suspensions, and biological samples containing a certain concentration of protein, immobilized with macromolecular structures provided elsewhere herein, can be incubated at temperatures from about 4 degrees Celsius to about 90 degrees Celsius. In some embodiments, one or more components of the composition can be incubated at temperatures from about 20 degrees Celsius to about 90 degrees Celsius. In some embodiments, one or more components of the composition can be incubated at temperatures from about 20 degrees Celsius to about 50 degrees Celsius. In some embodiments, one or more components of the composition can be incubated at temperatures from about 4 degrees Celsius to about 40 degrees Celsius. In some embodiments, one or more components of the composition can be incubated at temperatures from about 25 degrees Celsius to about 40 degrees Celsius.

[0266] In some implementations, the suspension may contain Tris, EDTA, and a CHAPS buffer. For example, the suspension may be Tris, EDTA in 150 mM KCl, and 0.05% CHAPS buffer. In another example, the suspension may be 10 mM Tris HCl pH 7.4 and 1 mM EDTA.

[0267] In some embodiments, this disclosure provides an automated system comprising a network of units as described in U.S. Patent No. 11,428,688, which is incorporated herein by reference in its entirety. In some embodiments, the network of units may include differentiated functions to distinguish the state of complex biological samples using multiple macromolecular structures with different physicochemical properties, wherein: a first unit includes a multichannel fluid transfer device for transferring fluids between units within the system; a second unit includes a support for storing multiple biological samples; a third unit includes a support for a sensor array plate with partitions comprising macromolecular structures for binding and interacting with a population of analytes from the biological samples; a fourth unit includes a support for storing multiple reagents; a fifth unit includes a support for storing reagents to be disposed of; a sixth unit includes a support for storing consumables used by the multichannel fluid transfer device; and wherein the system is programmed to perform a series of steps including: contacting a complex biological sample with a designated partition of the sensor array; incubating the complex biological sample together with multiple macromolecular structures contained within the partitions of the sensor array plate; removing components from the partitions other than the multiple macromolecular structures and the population of analytes interacting with the macromolecular structures; and optionally preparing a population of analytes for analysis (such as mass spectrometry).

[0268] In some embodiments, the first unit includes a degree of mobility, enabling access to all other units within the system. In some embodiments, the first unit includes the ability to perform pipetting functions.

[0269] In some embodiments, the supports for the second and / or third units include supports for single-plate, 6-well, 12-well, 96-well, or microtube racks. In some embodiments, the second and / or unit includes a heating unit capable of adjusting the temperature of the supports and the sample. In some embodiments, the second and / or third unit includes a rotating unit capable of physically agitating and / or mixing the sample.

[0270] In some embodiments, multiple macromolecular structures with different physicochemical properties to bind analyte populations within a biological sample are immobilized within partitions of a sensor array. In some embodiments, multiple particles comprise multiple magnetic nanoparticles with different physicochemical properties to bind analyte populations within complex biological samples. In some embodiments, the system includes a step in which the sensor array plate is transferred to an additional seventh unit and incubated for an additional amount of time, the seventh unit comprising a magnetized support and a thermal unit capable of adjusting the temperature of the support and the sample.

[0271] In some embodiments, the fourth unit includes a reagent kit for: generating the sensor array plate; washing unbound samples; and / or preparing samples for mass spectrometry. In some embodiments, contacting a biological sample with a designated section of the sensor array includes pipetting a designated volume of biological sample into the designated section of the sensor array.

[0272] In some embodiments, contacting a biological sample with a designated section of the sensor array includes pipetting a volume of biological sample of at least 10 μL, at least 20 μL, at least 50 μL, at least 100 μL, at least 250 μL, at least 500 μL, or at least 1000 μL into the designated section of the sensor array. In some embodiments, contacting a biological sample with a designated section of the sensor array includes transferring a volume of no more than 1000 μL, no more than 500 μL, no more than 250 μL, no more than 150 μL, no more than 100 μL, no more than 75 μL, no more than 50 μL, or no more than 30 μL.

[0273] In some embodiments, the biological sample may be diluted with water or a buffer. In some embodiments, the biological sample may be diluted at least 2, 3, 4, or 5 times. In some embodiments, the biological sample may be diluted no more than 20, 10, 8, or 5 times. In some embodiments, the biological sample is diluted with water or a buffer about 2 to about 5 times.

[0274] In some implementations, incubating the biological sample together with multiple macromolecular structures contained within a partition of the sensor array plate includes incubation periods of at least approximately 10 seconds, at least approximately 15 seconds, at least approximately 20 seconds, at least approximately 25 seconds, at least approximately 30 seconds, at least approximately 40 seconds, at least approximately 50 seconds, at least approximately 60 seconds, at least approximately 90 seconds, at least approximately 2 minutes, at least approximately 3 minutes, at least approximately 4 minutes, at least approximately 5 minutes, at least approximately 6 minutes, at least approximately 7 minutes, at least approximately 8 minutes, at least approximately 9 minutes, at least approximately 10 minutes, at least approximately 15 minutes, at least approximately 20 minutes, and at least... Incubation times of approximately 25 minutes, at least approximately 30 minutes, at least approximately 45 minutes, at least approximately 50 minutes, at least approximately 60 minutes, at least approximately 90 minutes, at least approximately 2 hours, at least approximately 3 hours, at least approximately 4 hours, at least approximately 5 hours, at least approximately 6 hours, at least approximately 7 hours, at least approximately 8 hours, at least approximately 9 hours, at least approximately 10 hours, at least approximately 12 hours, at least approximately 14 hours, at least approximately 15 hours, at least approximately 16 hours, at least approximately 17 hours, at least approximately 18 hours, at least approximately 19 hours, at least approximately 20 hours, or at least approximately 24 hours. In some embodiments, incubating biological samples together with multiple macromolecular structures contained within a partition of the sensor array plate includes incubation times of no more than 24 hours, no more than 12 hours, no more than 6 hours, no more than 3 hours, no more than 2 hours, no more than 90 minutes, no more than 75 minutes, no more than 60 minutes, no more than 45 minutes, or no more than 30 minutes. In some implementations, the biological sample is incubated together with multiple macromolecular structures contained within a partition of the sensor array plate for incubation times of 30 minutes and 3 hours.

[0275] In some embodiments, incubating a biological sample together with multiple macromolecular structures contained within a partition of a substrate includes incubation temperatures between about 4°C and about 40°C. Incubating a biological sample together with multiple macromolecular structures contained within a partition of a substrate may include incubation temperatures between about 4°C and about 37°C. Incubating a biological sample together with multiple macromolecular structures contained within a partition of a substrate may include incubation temperatures between about 20°C and about 50°C. Incubating a biological sample together with multiple macromolecular structures contained within a partition of a substrate may include incubation temperatures between about 4°C and about 100°C.

[0276] In some implementations, the second unit can facilitate the transfer of samples for mass spectrometry to the mass spectrometry unit.

[0277] In some embodiments, this disclosure provides an automated apparatus for identifying proteins in biological samples, the automated apparatus comprising: a sample preparation unit; a substrate including multiple channels; multiple pipettes; and multiple solutions and multiple macromolecular structures as described herein. In some embodiments, the automated apparatus is configured to form and digest protein crowns.

[0278] In some embodiments, the automated device also includes a magnetic source. In some embodiments, the automated device is configured for BCA, gel, or trypsin digestion of protein crowns.

[0279] In some embodiments, the automated device is enclosed. In some embodiments, the automated device is sterilized before use. In some embodiments, the automated device is configured as a mass spectrometer. In some embodiments, the automated device is temperature-controlled.

[0280] A variety of analytical techniques can be used to identify, measure, and quantify proteomics data from samples. For example, SDS-PAGE or any gel-based separation technique can be used to analyze proteomics data. Peptides and proteins can also be identified, measured, and quantified using immunoassays such as ELISA. Alternatively, mass spectrometry, high-performance liquid chromatography, LC-MS / MS, Edman degradation assays, immunoaffinity techniques, and the methods disclosed in EP3548652, WO2019083856, and WO2019133892 (each of which is incorporated herein by reference in its entirety) and other protein separation techniques can be used to identify, measure, and quantify proteomics data. In some embodiments, this disclosure provides an automated apparatus for generating a subset of biomolecules from a biological sample, comprising: a substrate including multiple partitions, a first unit containing the biological sample, and a loading unit movable along the substrate and capable of transferring volumes (e.g., volumes of buffers) between different units of the apparatus. In some cases, the substrate is a porous plate.

[0281] Multiple partitions may include multiple sensor elements. Multiple sensor elements may include surfaces. Multiple sensor elements may be macromolecular structures (e.g., particles) as disclosed herein. For example, sensor elements may include a first unique nanoparticle and a second unique nanoparticle.

[0282] A partition within a plurality of partitions may include 1 to 100 types of sensor elements (e.g., unique macromolecular structures). A partition within a plurality of partitions may include 2 to 50 types of sensor elements. A partition within a plurality of partitions may include 2 to 20 types of sensor elements. A partition within a plurality of partitions may include 2 to 5 types of sensor elements. A partition within a plurality of partitions may include 3 to 8 types of sensor elements. A partition within a plurality of partitions may include 4 to 10 types of sensor elements. A partition within a plurality of partitions may include 5 to 12 types of sensor elements. A partition within a plurality of partitions may include 6 to 15 types of sensor elements. A partition within a plurality of partitions may include 8 to 20 types of sensor elements. A partition within a plurality of partitions may include 2 types of sensor elements. A partition within a plurality of partitions may include 3 types of sensor elements. A partition within a plurality of partitions may include 4 types of sensor elements. A partition within a plurality of partitions may include one type of sensor element.

[0283] Two or more partitions in a plurality of partitions may include different numbers of sensor elements. Two or more partitions in a plurality of partitions may include different types of sensor elements. A partition in a plurality of partitions may include combinations of sensor element types and / or numbers that are different from those in other partitions in a plurality of partitions. A subset of partitions in a plurality of partitions may each contain unique combinations of sensor elements that are different from those in other partitions in a plurality of partitions.

[0284] Sensor elements can be stored in a dry form inside or within a partition. Dry sensor elements can be rehydrated or restored before use. Sensor elements can also be stored in a solution. For example, the substrate partition may include a solution containing a high concentration of macromolecular structures.

[0285] The partitions within the multiple partitions include sensor elements of different concentrations or amounts (e.g., mass / molarity per unit volume of sample). Partitions within the multiple partitions may include sensor elements from 1 pM to 100 nM. Partitions within the multiple partitions may include sensor elements from 1 pM to 500 pM. Partitions within the multiple partitions may include sensor elements from 10 pM to 1 nM. Partitions within the multiple partitions may include sensor elements from 100 pM to 10 nM. Partitions within the multiple partitions may include sensor elements from 500 pM to 100 nM. Partitions within the multiple partitions may include sensor elements from 50 μg / ml to 300 μg / ml. Partitions within the multiple partitions may include sensor elements from 100 μg / ml to 500 μg / ml. Partitions within the multiple partitions may include sensor elements from 250 μg / ml to 750 μg / ml. Partitions within the multiple partitions may include sensor elements from 400 μg / ml to 1 mg / ml. Partitions within the multiple partitions may include sensor elements from 600 μg / ml to 1.5 mg / ml. A partition within a plurality of partitions may include sensor elements with a concentration of 800 μg / ml to 2 mg / ml. A partition within a plurality of partitions may include sensor elements with a concentration of 1 mg / ml to 3 mg / ml. A partition within a plurality of partitions may include sensor elements with a concentration of 2 mg / ml to 5 mg / ml. A partition within a plurality of partitions may include sensor elements with a concentration greater than 5 mg / ml.

[0286] The loading unit can be configured to move and transfer volumes (e.g., volumes of solution or powder) between any units, compartments, or zones within the device. The loading unit can be configured to move precise volumes (e.g., within 0.1%, 0.01%, or 0.001% of a specified volume). The loading unit can be configured to collect volumes from or within a substrate, from a compartment or zone within the substrate, and dispensing the volumes back into or within a compartment or zone of the substrate, or dispensing volumes or portions of volumes into different units, compartments, or zones. The loading unit can be configured to move multiple volumes simultaneously, such as 2 to 400 individual volumes. The loading unit may include multiple pipette tips.

[0287] The loading unit can be configured to move a volume of liquid. The volume can be approximately 0.1 μl, 0.2 μl, 0.3 μl, 0.4 μl, 0.5 μl, 0.6 μl, 0.7 μl, 0.8 μl, 0.9 μl, 1 μl, 2 μl, 3 μl, 4 μl, 5 μl, 6 μl, 7 μl, 8 μl, 9 μl, 10 μl, 12 μl, 15 μl, 20 μl, 25 μl, 30 μl, 40 μl, 50 μl, 60 μl, 70 μl, 80 μl, 90 μl, 100 μl, 120 μl, 150 μl, 180 μl, 200 μl, 250 μl, 300 μl, 400 μl, 500 μl, 600 μl, 800 μl, 1 ml, or greater than 1 ml. The liquid can be a biological sample or a solution.

[0288] In some cases, the solution includes washing solutions, resuspension solutions, denaturing solutions, buffers, reagents (e.g., reducing agents), or any combination thereof. In some cases, the solution contains biological samples.

[0289] Partly by virtue of these capabilities, the loading unit can partition the sample. In some embodiments, this includes partitioning the sample into multiple partitions. The sample can be partitioned into at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 15, 20, 25, 30, 40, 50, 60, 70, 80, 90, 100, 120, 150, 180, 200, 250, 300, 350, 400, 500 or more partitions. The sample can be partitioned into 96, 192 or 384 partitions. The automated device may include multiple substrates containing partitions. The automated device may include 1, 2, 3, 4, 5 or more substrates containing partitions. In some cases, the loading unit loads biological samples of different volumes into different partitions. In some cases, the loading unit loads the same volume into two or more partitions. The volume of biological sample loaded into the partition can be approximately 0.1 μl, 0.2 μl, 0.3 μl, 0.4 μl, 0.5 μl, 0.6 μl, 0.7 μl, 0.8 μl, 0.9 μl, 1 μl, 2 μl, 3 μl, 4 μl, 5 μl, 6 μl, 7 μl, 8 μl, 9 μl, 10 μl, 12 μl, 15 μl, 20 μl, 25 μl, 30 μl, 40 μl, 50 μl, 60 μl, 70 μl, 80 μl, 90 μl, 100 μl, 120 μl, 150 μl, 180 μl, 200 μl, 250 μl, 300 μl, 400 μl, 500 μl, 600 μl, 800 μl, 1 ml or greater than 1 ml. The volume of biological sample loaded into the partition can be from about 10 μl to 400 μl. The volume of biological sample loaded into the partition can be from about 5 μl to 150 μl. The volume of biological sample loaded into the partition can be from about 35 μl to 80 μl. In some cases, the loading unit can partition two or more biological samples. For example, the sample storage unit can contain two biological samples, which the system partitions in a one-well plate. In some embodiments, the loading unit can facilitate the transfer of samples for mass spectrometry to the mass spectrometry unit.

[0290] The system can be configured to dilute samples or sample partitions. Samples or sample partitions can be diluted with buffers, water (e.g., purified water), non-aqueous solvents, or any combination thereof. The diluent can be stored in the automated device before being dispensed into the substrate partitions. The automated device can store multiple diluents differing in pH, salinity, osmolality, viscosity, dielectric constant, or any combination thereof. The diluent can be used to adjust the chemical properties of the sample or sample partition. The automated device can dilute samples or sample partitions by 2, 3, 4, 5, 6, 8, 10, 15, 20, 30, 40, 50, 75, 100, 150, 200, 300, 400, 500, or more times. In some embodiments, the automated device can dilute samples or sample partitions by about 2 to about 5 times. The automated device can perform different dilutions on two samples or sample partitions. The system can perform different dilutions on each of multiple partitions. For example, the system can perform different dilutions for each of the 96 sample partitions in a 96-well plate. In some cases, different dilutions involve different degrees of dilution (e.g., 2-fold vs. 4-fold). In some cases, different dilutions involve dilution with different solutions (e.g., different buffers). In some cases, it can be made possible for two sample partitions to differ in one or more chemical properties (such as pH, salinity, or viscosity).

[0291] In some cases, this system can modify the chemical composition of a sample or sample partition. The system can modify or adjust pH, salinity, osmolality, dielectric constant, viscosity, buffer type, salt type, sugar type, detergent type, or any combination thereof for a sample or sample partition. Such modifications or adjustments may include mixing reagents from Unit 4 with the sample or sample partition. The system can modify the chemical composition of two samples or sample partitions differently.

[0292] The systems or automation devices disclosed herein may also include incubation elements. Incubation elements may contact, support, or hold another component (e.g., a substrate or unit) of the automation device. Incubation elements may contact, support, or hold multiple components of the automation device. Incubation elements may contact a substrate to facilitate heat transfer between the incubation element and the substrate. Incubation elements may be configured to control the temperature of one or more components of the automation device, such as by heating or cooling. Incubation elements may enable a component of the device to cool from 20°C to 1°C. Incubation elements may enable a component of the device to heat from 25°C to 100°C. Incubation elements may enable the temperature of a component of the device to be set from 4°C to 37°C. Incubation elements may be configured to heat or cool different portions of a component of the automation device to different temperatures. For example, an incubation element may maintain a first section in a substrate at 30°C and a second section in a substrate at 35°C. Incubation elements may control the temperature of a sample or section. Incubation elements may include temperature sensors (e.g., thermocouples) for detecting the temperature within a section or container. Incubation elements may calibrate their heating or cooling based on readings from the temperature sensors.

[0293] Incubation elements can be configured to physically agitate components of an automated device. Agitation can take the form of shaking or rotation, vibration, rocking, sonication, or any combination thereof. Incubation elements can be capable of providing a variety of agitation intensities and / or frequencies. For example, an incubation element may include multiple settings for shaking at different frequencies and amplitudes. Incubation elements can also be capable of stirring and / or mixing volumes (e.g., portions of biological samples).

[0294] Automated devices may include a unit containing a resuspension solution. The loading unit may be able to transfer a volume of the resuspension solution to a partition among multiple partitions of a substrate. In some cases, this results in dilution of the sample present within the partition and may further result in the desorption of multiple biomolecules from a biocluster deposited on a sensor element within the partition. The number of biomolecules desorbed from the biocluster can depend on the volume of the resuspension solution added to the partition, the temperature of the partition, the composition of the resuspension solution (e.g., salinity, osmolality, viscosity, dielectric constant, or pH), the volume of the biological sample within the partition, and the type of sensor element and the composition of the biomolecules in the biocluster. Volume transfer of the resuspension solution to the partition may result in less than 5% desorption of biomolecules from the biocluster. Volume transfer of the resuspension solution to the partition may result in 10% to 20% desorption of biomolecules from the biocluster. Volume transfer of the resuspension solution to the partition may result in 20% to 30% desorption of biomolecules from the biocluster. Volume transfer of the resuspension solution to the partition may result in 30% to 40% desorption of biomolecules from the biocluster. The volume transfer of a resuspension into a partition can result in 40% to 50% desorption of biomolecules from the biocluster. The volume transfer of a resuspension into a partition can result in 50% to 60% desorption of biomolecules from the biocluster. The volume transfer of a resuspension into a partition can result in 60% to 70% desorption of biomolecules from the biocluster. The volume transfer of a resuspension into a partition can result in 70% to 80% desorption of biomolecules from the biocluster. The volume transfer of a resuspension into a partition can result in 80% to 90% desorption of biomolecules from the biocluster. The volume transfer of a resuspension into a partition can result in more than 90% desorption of biomolecules from the biocluster.

[0295] In some cases, multiple rounds of desorption are performed. In each round, the supernatant containing the desorbed biomolecules can be collected, analyzed, or discarded. The type and abundance of biomolecules in the supernatant may vary between desorption rounds. Automated devices can perform one or more desorption and discard cycles (i.e., washing), followed by one or more desorption cycles including sample collection and / or analysis.

[0296] Resuspension solutions can be customized to optimize the enrichment of specific biomarkers. Resuspension solutions may contain buffers such as Tris-EDTA (TE), CHAPS, PBS, citrate, HEPES, MES, CHES, or other biological buffers. A resuspension solution may contain Tris-EDTA (TE) 150 mM KCl 0.05% CHAPS buffer. A resuspension solution may contain 10 mM TrisHCl pH 7.4 and 1 mM EDTA. Resuspension solutions may also contain highly purified water (e.g., distilled or deionized water). Biomolecular desorption can be enhanced by heating or agitation using an incubation element. The supernatant can be transferred to a new partition after desorption. Resuspension solutions can be used to dilute samples.

[0297] The automated device may include a unit containing a denaturing solution. The denaturing solution may contain a protease. The denaturing solution may contain chemicals capable of peptide cleavage (e.g., cyanogen bromide, formic acid, or hydroxylamine, 2-nitro-5-thiocyanobenzoic acid). The denaturing solution may contain chemical denaturing agents such as guanidine, urea, sodium deoxycholate, acetonitrile, trichloroacetic acid, acetic acid, sulfosalicylic acid, sodium bicarbonate, ethanol, perchlorate, dodecyl sulfate, or any combination thereof. The denaturing solution may contain a reducing agent such as 2-mercaptoethanol, dithiothreitol, or tris(2-carboxyethyl)phosphine. The protease may be trypsin. The denaturing solution may be added to a partition after desorption. The denaturing solution may be added to a partition containing a biomolecular crown.

[0298] The automated device may include magnets or an array of magnets. The automated device may be capable of moving a substrate to and from a magnet or array of magnets. The magnet array may be configured such that multiple magnets from the magnet array can be placed directly beneath multiple sections of the substrate. The magnets may be capable of securing magnetic sensor elements (e.g., magnetic particles, such as coated or uncoated superparamagnetic iron oxide nanoparticles) within the sections on the substrate. For example, the magnets may prevent the magnetic nanoparticles from being removed from the sections during a washing step. The magnets may also generate clumps from a batch of magnetic particles. The magnets may generate particle clumps in less than 10 minutes. The magnets may generate particle clumps in less than 5 minutes. The particle clumps may include particles with biomolecular crowns.

[0299] The automated apparatus may include a purification unit. The purification unit may include multiple compartments containing an adsorbent or resin. The purification unit may include a solid-phase extraction array or plate. The solid-phase extraction array or plate may contain a polar stationary phase material. The solid-phase extraction array or plate may contain a non-polar stationary phase material. The solid-phase extraction array or plate may contain a C18 stationary phase material (e.g., octadecyl silica gel). The automated apparatus may include a unit containing a conditioning solution for the purification unit (e.g., a conditioning solution for the solid-phase extraction material). The automated apparatus may include a unit having an elution solution for removing biomolecules from the purification unit.

[0300] In some implementations, the supernatant is removed from the sensor array plate. In some cases, automated devices may perform a series of washing steps. Washing steps can remove unbound biomolecules within a partition. Washing steps can desorb a subset of biomolecules bound to the sensor element within a partition. For example, washing steps can result in the desorption and removal of a subset of soft-corona analytes, leaving the majority of hard-corona analytes bound to the sensor element.

[0301] In some embodiments, this disclosure provides an automated apparatus for identifying proteins in biological samples, the automated apparatus comprising: a sample preparation unit; a substrate including multiple channels; multiple pipettes; multiple solutions; multiple macromolecular structures (such as macromolecules provided elsewhere herein); and wherein the automated apparatus is configured to form and digest protein crowns.

[0302] In some embodiments, the automated device also includes a magnetic source. In some embodiments, the automated device is configured for BCA, gel, or trypsin digestion of protein crowns.

[0303] In some embodiments, the automated device is enclosed. In some embodiments, the automated device is sterilized before use. In some embodiments, the automated device is configured as a mass spectrometer. In some embodiments, the automated device is temperature-controlled.

[0304] Reagent test kit In one aspect, this document describes a kit for identifying biomolecules in biological samples, wherein the kit may comprise one or more macromolecular structures as described elsewhere herein. In some embodiments, the kit can be used to perform methods for identifying proteins in samples as provided herein. In some embodiments, the kit may be configured to perform methods using automated devices as provided herein.

[0305] The kit disclosed herein may contain one or more macromolecular structures (e.g., particles) for querying samples. The kit may be pre-packaged as discrete aliquots. In another example, the kit may contain one or more different macromolecular structures (e.g., particles with different surface chemistry) that can be used for querying samples. Multiple different macromolecular structures may be pre-packaged, with each of the multiple macromolecules packaged individually. Alternatively, multiple macromolecular structures may be packaged together to contain a combination of macromolecular structures in a single package. In some embodiments, the kit contains two or more packages containing different macromolecular structures (e.g., particles), wherein at least one package contains two or more different macromolecular structures (e.g., particles). In some embodiments, the macromolecular structures (e.g., particles) may be freeze-dried and stored in a sealed container.

[0306] In some embodiments, the kit also includes a denaturing agent. In some embodiments, the denaturing agent includes at least one of the following: sodium dodecyl sulfate, acetic acid, trichloroacetic acid, sulfosalicylic acid, sodium bicarbonate, ethanol, formaldehyde, glutaraldehyde, urea, guanidine chloride, lithium perchlorate, 2-mercaptoethanol, dithiothreitol, tris(2-carboxyethyl)phosphine (TCEP), or any combination thereof.

[0307] In some embodiments, the kit also includes a reducing agent. In some embodiments, the reducing agent includes TCEP, dithiothreitol, β-mercaptoethanol, glutathione, cysteine, or any combination thereof.

[0308] In some embodiments, the kit also contains an alkylating agent. In some embodiments, the alkylating agent includes iodoacetamide, iodoacetic acid, acrylamide, chloroacetamide, or any combination thereof.

[0309] In some embodiments, the kit also includes a digestive agent. In some embodiments, the digestive agent includes trypsin, lysozyme, serine protease, or any combination thereof.

[0310] In some embodiments, the kit also includes an eluent. In some embodiments, the eluent buffer includes triethylammonium bicarbonate, tris(hydroxymethyl)aminomethane, citrate, Tris, phosphate, ethylenediaminetetraacetic acid, or any combination thereof.

[0311] In some embodiments, the kit also contains a detergent. In some embodiments, the detergent is water or a buffer.

[0312] In some embodiments, the kit further comprises a solid support for solid-phase extraction. In some embodiments, the kit comprises a polar stationary phase material. In some embodiments, the kit comprises a non-polar stationary phase material. In some embodiments, the kit comprises a C18 stationary phase material (e.g., octadecyl silica gel). In some embodiments, the kit comprises a conditioning solution for the solid-phase extraction material.

[0313] In some embodiments, the kit also includes a multi-well plate. In some embodiments, the multi-well plate is a 4-well plate. In some embodiments, the multi-well plate is a 12-well plate. In some embodiments, the multi-well plate is a 24-well plate. In some embodiments, the multi-well plate is a 48-well plate. In some embodiments, the multi-well plate is a 96-well plate. In some embodiments, the multi-well plate is a 384-well plate. In some embodiments, the multi-well plate is a 1536-well plate.

[0314] In some embodiments, the kit also includes a diluent. In some embodiments, the diluent is an organic solvent. In some embodiments, the diluent is water. In some embodiments, the diluent is a buffer. In some embodiments, the diluent is an organic solvent, water, a buffer, or any combination thereof.

[0315] In some embodiments, the kit further comprises an organic solvent. In some embodiments, the kit further comprises a cysteine ​​blocking agent. In some embodiments, the cysteine ​​blocking agent comprises methyl methanethiosulfonate, iodoacetamide, N-ethylmaleimide, methanesulfonylbenzothiazole, or any combination thereof.

[0316] Implementation plan with numbering This document provides implementation scheme 1-271, numbered as follows.

[0317] Implementation Scheme 1. A macromolecular structure, said macromolecular structure comprising: (I) Surface; (II) The tethered portion coupled to the surface; and (III) A macromolecular chain, wherein the first end of the macromolecular chain is covalently attached to the tethered portion, and wherein the macromolecular chain comprises two or more distinct repeating units derived from monomers selected from the following structures: , , , , , , , and

[0318] Each of X and Y is independently -C-, -O-, or -N-; Each Z is independently -O- or -NH; Q is -CH2- or ethylene glycol; A is a polymer side chain comprising repeating units derived from monomers represented by the following structures: ; m is an integer selected from 1 to 20; Is it a single bond or a double bond? R 1 R 2 R 1’ R 2’ and R 3’ Each of them is independently selected from hydrogen or -C1-C6 alkyl; R 3 It is a hydrogen, a C1-C6 alkyl group, or a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl groups; R 4 It is absent, hydrogen, sulfonate, carboxylic acid ester, C1-C4 alkylene, amine, quaternary ammonium cation or C1-C6 alkyl optionally substituted with halogen; R 5 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, an amine, an azide, a sulfonate, a carbamate, an asymmetric disulfide, or a C1-C8 alkyl group or a C1-C6 heterocyclic group substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines optionally further substituted with amine, hydroxyl, aryl or sulfonate, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogens, C1-C8 alkyl groups optionally substituted with one or more pyrene, two or more fused 5-6 membered rings optionally further substituted, -C1-C3 alkyl groups optionally substituted with benzyl, trimethoxysilane or phosphocholine, C1-C 12 Alkylamines or C1-C4 alkylamines; R 6 It is hydrogen or C1-C6 alkyl. R 7 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, amines, azides, sulfonates, carbamates, or asymmetric disulfides, or a 3-, 5-, or 6-membered heterocyclic or C1-C alkyl group optionally substituted with one or more C1-C6 alkyl groups or oxo groups. n1Ethylene glycol, C1-C8 alkylamines optionally further substituted with amines or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogens, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted, optionally substituted benzyl groups, -C1-C3 alkyl or C1-C4 alkylyl groups substituted with trimethoxysilane or phosphocholine; R 8 It is a C1-C6 alkyl group, a divalent metal, or a symmetrical or asymmetrical disulfide; R 9 It is hydrogen or oxygen; and n 1 It is an integer selected from 1 to 100.

[0319] Implementation Scheme 2. The macromolecular structure as described in Implementation Scheme 1, wherein the macromolecular chain comprises repeating units derived from monomers represented by the following structures: .

[0320] Implementation Scheme 3. A macromolecular structure as described in Implementation Scheme 1 or 2, wherein R 1 It is hydrogen.

[0321] Implementation Scheme 4. The macromolecular structure as described in any one of Implementation Schemes 1-3, wherein R 2 It is hydrogen.

[0322] Implementation Scheme 5. The macromolecular structure as described in any one of Implementation Schemes 1-4, wherein R 3 It is hydrogen.

[0323] Implementation Scheme 6. The macromolecular structure as described in any one of Implementation Schemes 1-5, wherein R 5 It is a hydrogen, C1-C6 alkyl, or a C1-C3 alkyl group substituted with pyrene or two or more fused 5-6 membered rings optionally further substituted.

[0324] Implementation Scheme 7. The macromolecular structure as described in any one of Implementation Schemes 1-4, wherein R 3 It is a methyl group.

[0325] Implementation Scheme 8. The macromolecular structure as described in Implementation Scheme 7, wherein R 5 It is C1-C n1 Ethylene glycol.

[0326] Implementation Scheme 9. The macromolecular structure as described in any one of Implementation Schemes 1-8, wherein the macromolecular chain comprises repeating units derived from monomers represented by the following structures: .

[0327] Implementation Scheme 10. The macromolecular structure as described in Implementation Scheme 9, wherein R1 It is hydrogen.

[0328] Implementation Scheme 11. The macromolecular structure as described in Implementation Scheme 9 or 10, wherein R 2 It is hydrogen.

[0329] Implementation Scheme 12. The macromolecular structure as described in any one of Implementation Schemes 9-11, wherein R 3 It is H.

[0330] Implementation Scheme 13. The macromolecular structure as described in any one of Implementation Schemes 9-11, wherein R 3 It is a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl groups.

[0331] Implementation Scheme 14. The macromolecular structure as described in any one of Implementation Schemes 9-13, wherein R 6 It is H.

[0332] Implementation Scheme 15. The macromolecular structure as described in any one of Implementation Schemes 9-14, wherein R 7 It is a C1-C6 alkyl group optionally substituted with a hydroxyl group, a substituted benzene, or a hydrogen.

[0333] Implementation Scheme 16. The macromolecular structure as described in any one of Implementation Schemes 1-15, wherein the macromolecular chain comprises repeating units derived from monomers represented by the following structures: .

[0334] Implementation Scheme 17. The macromolecular structure as described in Implementation Scheme 16, wherein R 1 It is hydrogen.

[0335] Implementation Scheme 18. A macromolecular structure as described in Implementation Scheme 16 or 17, wherein R 2 It is hydrogen.

[0336] Implementation Scheme 19. The macromolecular structure as described in any one of Implementation Schemes 16-18, wherein R 1’ It is hydrogen.

[0337] Implementation Scheme 20. The macromolecular structure as described in any one of Implementation Schemes 16-19, wherein R 2’ It is hydrogen.

[0338] Implementation Scheme 21. The macromolecular structure as described in any one of Implementation Schemes 16-20, wherein R 3 It is a methyl group.

[0339] Implementation Scheme 22. The macromolecular structure as described in any one of Implementation Schemes 16-21, wherein R 3’ It is a methyl group.

[0340] Implementation Scheme 23. The macromolecular structure as described in any one of Implementation Schemes 16-22, wherein each Z is O.

[0341] Implementation Scheme 24. The macromolecular structure as described in any one of Implementation Schemes 16-23, wherein R 8 It is a C1-C6 alkyl group.

[0342] Implementation Scheme 25. The macromolecular structure as described in any one of Implementation Schemes 16-23, wherein R 8 It is a symmetrical disulfide (e.g., CH2CH2S-SCH2CH2) or a divalent metal.

[0343] Implementation Scheme 26. The macromolecular structure as described in any one of Implementation Schemes 16-20, wherein R 3 It is hydrogen.

[0344] Implementation Scheme 27. The macromolecular structure as described in any one of Implementation Schemes 16-20, wherein R 3’ It is hydrogen.

[0345] Implementation Scheme 28. A macromolecular structure as described in any one of Implementation Schemes 26 or 27, wherein each Z is -N-.

[0346] Implementation Scheme 29. The macromolecular structure as described in any one of Implementation Schemes 26-28, wherein R 8 It is a C1-C6 alkyl group.

[0347] Implementation Scheme 30. A macromolecular structure, said macromolecular structure comprising: (I) Surface; (II) The tethered portion coupled to the surface; and (III) A macromolecular chain, wherein the first end of the macromolecular chain is covalently attached to the tethered portion, and wherein the macromolecular chain comprises repeating units derived from monomers selected from the following structures: , , , , , , , and

[0348] Each of X and Y is independently -C-, -O-, or -N-; Each Z is independently -O- or -NH; Q is -CH2- or ethylene glycol; A is a polymer side chain comprising repeating units derived from monomers represented by the following structures: ; m is 1-6; R 1 R 2 R 1’ R 2’ and R 3’ Each of them is independently selected from hydrogen or C1-C6 alkyl. R 3 It is a hydrogen, a C1-C6 alkyl group, or a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl groups; R 4 It is absent, hydrogen, or optionally halogenated C1-C6 alkyl, C1-C4 alkylene, C1-C6 alkyl, sulfonate, amine, quaternary ammonium cation or carboxylic acid ester; R 5 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, an amine, an azide, a sulfonate, a carbamate, an asymmetric disulfide, or a C1-C8 alkyl group or a C1-C6 heterocyclic group substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines further substituted with amines, hydroxyl groups, aryl groups, or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogen groups, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted (e.g., two or more fused 6 membered rings optionally further substituted), optionally substituted benzyl groups, trimethoxysilanes, or phosphoric acid choline-substituted -C1-C3 alkyl groups, C1-C 12 Alkylamines or C1-C4 alkylamines; R 6 It is hydrogen or a linear C1-C6 alkyl group. R 7 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, amines, azides, sulfonates, carbamates, or asymmetric disulfides, or a 3-, 5-, or 6-membered heterocyclic or C1-C alkyl group optionally substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines further substituted with amines or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogen groups, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted (e.g., two or more fused 6 membered rings optionally further substituted), optionally substituted benzyl groups, -C1-C3 alkyl or C1-C4 alkylyl groups substituted with trimethoxysilane or phosphocholine; and R 8 It is a C1-C6 alkyl or symmetrical disulfide or asymmetrical disulfide; R 9 It is hydrogen or oxygen; n 1 It is an integer selected from 1 to 100; and The condition is when R 3 It is CH3, R 4 When it is CH3 or when R 5 When the C1-C8 alkyl group is substituted with a hydroxyl group, the C1-C8 groups are further substituted.

[0349] Implementation Scheme 31. The macromolecular structure as described in any one of Implementation Schemes 1-30, wherein the macromolecular chain comprises repeating units derived from monomers represented by the following structures: .

[0350] Implementation Scheme 32. The macromolecular structure as described in Implementation Scheme 31, wherein Q is -CH2-.

[0351] Implementation Scheme 33. A macromolecular structure as described in Implementation Scheme 31 or 32, wherein R 1 It is hydrogen.

[0352] Implementation Scheme 34. The macromolecular structure as described in any one of Implementation Schemes 31-33, wherein R 2 It is hydrogen.

[0353] Implementation Scheme 35. The macromolecular structure as described in any one of Implementation Schemes 31-34, wherein R 3 It is a methyl group.

[0354] Implementation Scheme 36. The macromolecular structure as described in any one of Implementation Schemes 31-35, wherein m is 2.

[0355] Implementation Scheme 37. The macromolecular structure as described in any one of Implementation Schemes 31-36, wherein R 6 It is a C1-C6 alkyl group.

[0356] Implementation Scheme 38. The macromolecular structure as described in Implementation Scheme 31, wherein Q is ethylene glycol.

[0357] Implementation Scheme 39. The macromolecular structure as described in Implementation Scheme 38, wherein R 1 It is hydrogen.

[0358] Implementation Scheme 40. A macromolecular structure as described in Implementation Scheme 38 or 39, wherein R 2 It is hydrogen.

[0359] Implementation Scheme 41. The macromolecular structure as described in any one of Implementation Schemes 38-40, wherein R 6 It is a C1-C6 alkyl group.

[0360] Implementation Scheme 42. The macromolecular structure as described in any one of Implementation Schemes 38-41, wherein R 5 It is C1-C n1 Ethylene glycol.

[0361] Implementation Scheme 43. A macromolecular structure, said macromolecular structure comprising: (I) a surface and (II) a macromolecular chain coupled to the surface, wherein the macromolecular chain comprises repeating units of formula (I): (I) Where R 1” R 2” and R 3” Each of them is independently hydrogen or a C1-C6 alkyl group; L represents the connector section; A is a polymer side chain comprising repeating units derived from monomers selected from the following structures: , , , , , , and

[0362] Each of X and Y is independently -C-, -O-, or -N-; Z is -O- or -NH; R 1 R 2 R 1’ R 2’ and R 3’ Each of them is independently selected from hydrogen or C1-C6 alkyl; R 3 It is a hydrogen, a C1-C6 alkyl group, or a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl groups; R 4 It is absent, hydrogen, sulfonate, carboxylic acid ester, C1-C4 alkylene, amine, quaternary ammonium cation or C1-C6 alkyl optionally substituted with halogen; R 5It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, an amine, an azide, a sulfonate, a carbamate, an asymmetric disulfide, or a C1-C8 alkyl group or a C1-C6 heterocyclic group substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines further substituted with amines, hydroxyl groups, aryl groups, or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogen groups, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted (e.g., two or more fused 6 membered rings optionally further substituted), optionally substituted benzyl groups, trimethoxysilanes, or phosphoric acid choline-substituted -C1-C3 alkyl groups, C1-C 12 Alkylamines or C1-C4 alkylamines; R 6 It is hydrogen or a linear C1-C6 alkyl group. R 7 It is a hydrogen, C1-C6 alkyl, a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl or oxo compounds, a substituted benzene, a C1-C6 alkyl substituted with a hydroxyl group, or a C1-C8 alkyl sulfonate optionally substituted. R 8 It is a C1-C6 alkyl group, a divalent metal, or a symmetrical or asymmetrical disulfide; R 9 It is hydrogen or oxygen; n 1 It is an integer selected from 1 to 100; and n is an integer selected from 1 to 10,000.

[0363] Implementation Scheme 44. The macromolecular structure as described in Implementation Scheme 43, wherein the linker portion is represented by the following structure:

[0364] in: Each Z is independently O or N; and X' is a C1-C6 alkyl group.

[0365] Implementation Scheme 45. A macromolecular structure as described in Implementation Scheme 43 or 44, wherein the macromolecule further comprises a tethering portion.

[0366] Implementation Scheme 46. The macromolecular structure as described in any one of Implementation Schemes 43-45, wherein the macromolecular structure further comprises a crosslinked portion.

[0367] Implementation Scheme 47. The macromolecular structure as described in Implementation Scheme 46, wherein the crosslinking portion comprises... The structure represented, in: R 1 R 2 R 1’ R 2’ and R 3’ Each of them is independently selected from hydrogen or C1-C6 alkyl; Z is -O- or -NH; and R 8 It is a -C1-C6 alkyl, divalent metal, or symmetrical or asymmetrical disulfide.

[0368] Implementation Scheme 48. The macromolecular structure as described in any one of Implementation Schemes 1-47, wherein the macromolecular chain comprises repeating units derived from monomers represented by the following structures: .

[0369] Implementation Scheme 49. The macromolecular structure as described in Implementation Scheme 48, wherein R 1 It is H.

[0370] Implementation Scheme 50. A macromolecular structure as described in Implementation Scheme 48 or 49, wherein R 2 It is H.

[0371] Implementation Scheme 51. The macromolecular structure as described in any one of Implementation Schemes 48-50, wherein R 3 It is H.

[0372] Implementation Scheme 52. The macromolecular structure as described in any one of Implementation Schemes 48-51, wherein R 4 It is a C1-C6 alkyl group that is optionally substituted with halogen, absent, hydrogen, carboxylic acid ester or sulfonate.

[0373] Implementation Scheme 53. The macromolecular structure as described in any one of Implementation Schemes 48-51, wherein R 4 It is a C1-C4 alkylene group.

[0374] Implementation Scheme 54. The macromolecular structure as described in any one of Implementation Schemes 48-53, wherein X is -C- or -N-.

[0375] Implementation Scheme 55. The macromolecular structure as described in any one of Implementation Schemes 48-54, wherein Y is -C- or -N-.

[0376] Implementation Scheme 56. The macromolecular structure as described in any one of Implementation Schemes 1-55, wherein the macromolecular chain comprises repeating units derived from monomers represented by the following structures: .

[0377] Implementation Scheme 57. The macromolecular structure as described in Implementation Scheme 56, wherein R1 It is H.

[0378] Implementation Scheme 58. A macromolecular structure as described in Implementation Scheme 56 or 57, wherein R 2 It is H.

[0379] Implementation Scheme 59. The macromolecular structure as described in any one of Implementation Schemes 56-58, wherein R 3 It is H.

[0380] Implementation Scheme 60. The macromolecular structure as described in any one of Implementation Schemes 56-59, wherein R 5 It is a C1 alkyl, a pyrene-substituted C1-C3 alkyl, a -C1-C3 alkyl comprising two or more fused 5-6 membered rings optionally further substituted, or H.

[0381] Implementation Scheme 61. The macromolecular structure as described in any one of Implementation Schemes 56-58, wherein R 3 It is a methyl group.

[0382] Implementation Scheme 62. The macromolecular structure as described in Implementation Scheme 61, wherein R 5 It is an H, C2-C6 alkyl, C1-C8 alkyl or C1-C6 alkyl substituted with one or more hydroxyl groups, amines, azides, asymmetric disulfides, or optionally a 3-, 5-, or 6-membered heterocycle substituted with one or more C1-C6 alkyl or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines further substituted with amines or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogens, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted, -C1-C3 alkyl or C1-C4 alkylyne substituted with trimethoxysilane or phosphocholine.

[0383] Implementation Scheme 63. The macromolecular structure as described in Implementation Scheme 61, wherein R 5 It is C1-C n1 Ethylene glycol.

[0384] Implementation Scheme 64. The macromolecular structure as described in Implementation Scheme 61, wherein when R 5 When the C1-C6 alkyl group is substituted with a hydroxyl group, the C1-C6 group is substituted with at least one other hydroxyl group or azide.

[0385] Implementation Scheme 65. The macromolecular structure as described in any one of Implementation Schemes 1-64, wherein the macromolecular chain comprises repeating units derived from monomers represented by the following structures: .

[0386] Implementation Scheme 66. The macromolecular structure as described in Implementation Scheme 65, wherein R1 It is H.

[0387] Implementation Scheme 67. A macromolecular structure as described in Implementation Scheme 65 or 66, wherein R 2 It is H.

[0388] Implementation Scheme 68. The macromolecular structure as described in any one of Implementation Schemes 65-67, wherein R 3 It is a 3-, 5-, or 6-membered heterocycle, which is H or optionally substituted with one or more C1-C6 alkyl groups.

[0389] Implementation Scheme 69. The macromolecular structure as described in any one of Implementation Schemes 65-68, wherein R 6 It is a 3-, 5-, or 6-membered heterocycle, which is H or optionally substituted with one or more C1-C6 alkyl groups.

[0390] Implementation Scheme 70. The macromolecular structure as described in any one of Implementation Schemes 65-69, wherein R 7 It is a C1-C2 alkyl group, a -C1-C6 alkyl group substituted with a hydroxyl group, a substituted benzene, or a hydrogen.

[0391] Implementation Scheme 71. The macromolecular structure as described in any one of Implementation Schemes 1-70, wherein the macromolecular chain comprises repeating units derived from monomers represented by the following structures: .

[0392] Implementation Scheme 72. The macromolecular structure as described in Implementation Scheme 71, wherein R 1 It is hydrogen.

[0393] Implementation Scheme 73. A macromolecular structure as described in Implementation Scheme 71 or 72, wherein R 2 It is hydrogen.

[0394] Implementation Scheme 74. The macromolecular structure as described in any one of Implementation Schemes 71-73, wherein R 3 It is hydrogen.

[0395] Implementation Scheme 75. The macromolecular structure as described in any one of Implementation Schemes 71-74, wherein It is a double bond.

[0396] Implementation Scheme 76. The macromolecular structure as described in any one of Implementation Schemes 71-75, wherein X is -N-.

[0397] Implementation Scheme 77. The macromolecular structure as described in any one of Implementation Schemes 71-76, wherein Y is -N-.

[0398] Implementation Scheme 78. The macromolecular structure as described in any one of Implementation Schemes 71-77, wherein R 9 It is hydrogen.

[0399] Implementation Scheme 79. The macromolecular structure as described in any one of Implementation Schemes 71-74, wherein It is a single key.

[0400] Implementation scheme 80. The macromolecular structure as described in implementation scheme x, wherein 79 is -N-.

[0401] Implementation Scheme 81. The macromolecular structure as described in Implementation Scheme 79 or 80, wherein Y is -C-.

[0402] Implementation Scheme 82. The macromolecular structure as described in any one of Implementation Schemes 79-81, wherein R 9 It is oxygenation.

[0403] Implementation Scheme 83. The macromolecular structure as described in any one of Implementation Schemes 1-82, wherein the macromolecular chain comprises repeating units derived from monomers represented by the following structures: .

[0404] Implementation Scheme 84. The macromolecular structure as described in Implementation Scheme 83, wherein X is -O-.

[0405] Implementation Scheme 85. A macromolecular structure as described in Implementation Scheme 83 or 84, wherein R 1 It is hydrogen.

[0406] Implementation Scheme 86. The macromolecular structure as described in any one of Implementation Schemes 83-85, wherein R 2 It is hydrogen.

[0407] Implementation Scheme 87. The macromolecular structure as described in any one of Implementation Schemes 1-86, wherein the macromolecular chain comprises repeating units derived from monomers represented by the following structures: .

[0408] Implementation Scheme 88. The macromolecular structure as described in Implementation Scheme 87, wherein R 1 It is hydrogen.

[0409] Implementation Scheme 89. A macromolecular structure as described in Implementation Scheme 87 or 88, wherein R 2 It is hydrogen.

[0410] Implementation Scheme 90. The macromolecular structure as described in any one of Implementation Schemes 87-89, wherein R 3 It is hydrogen.

[0411] Implementation Scheme 91. The macromolecular structure as described in any one of Implementation Schemes 87-90, wherein R 3’ It is a methyl group.

[0412] Implementation Scheme 92. The macromolecular structure as described in any one of Implementation Schemes 87-91, wherein R 1’ It is hydrogen.

[0413] Implementation Scheme 93. The macromolecular structure as described in any one of Implementation Schemes 87-92, wherein R 2’ It is hydrogen.

[0414] Implementation Scheme 94. The macromolecular structure as described in any one of Implementation Schemes 1-93, wherein the macromolecular chain comprises repeating units derived from monomers represented by the following structures: .

[0415] Implementation Scheme 95. The macromolecular structure as described in Implementation Scheme 94, wherein R 1 It is hydrogen.

[0416] Implementation Scheme 96. A macromolecular structure as described in Implementation Scheme 94 or 95, wherein R 2 It is hydrogen.

[0417] Implementation Scheme 97. The macromolecular structure as described in any one of Implementation Schemes 94-96, wherein R 3 It is hydrogen.

[0418] Implementation Scheme 98. The macromolecular structure as described in any one of Implementation Schemes 1-97, wherein the macromolecular chain comprises repeating units derived from monomers represented by the following structures: .

[0419] Implementation Scheme 99. The macromolecular structure as described in Implementation Scheme 98, wherein R 1 It is hydrogen.

[0420] Implementation Scheme 100. A macromolecular structure as described in Implementation Scheme 98 or 99, wherein R 2 It is hydrogen.

[0421] Implementation Scheme 101. The macromolecular structure as described in any one of Implementation Schemes 98-100, wherein R 1’ It is hydrogen.

[0422] Implementation Scheme 102. The macromolecular structure as described in any one of Implementation Schemes 98-101, wherein R 2’ It is hydrogen.

[0423] Implementation Scheme 103. The macromolecular structure as described in any one of Implementation Schemes 98-102, wherein R 3 It is a methyl group.

[0424] Implementation Scheme 104. The macromolecular structure as described in any one of Implementation Schemes 98-103, wherein R 3’ It is a methyl group.

[0425] Implementation Scheme 105. The macromolecular structure as described in any one of Implementation Schemes 98-104, wherein Z is O.

[0426] Implementation Scheme 106. The macromolecular structure as described in any one of Implementation Schemes 98-105, wherein R 8 It is a C1-C6 alkyl group.

[0427] Implementation Scheme 107. The macromolecular structure as described in any one of Implementation Schemes 98-105, wherein R 8 It is a symmetrical disulfide (e.g., CH2CH2S-SCH2CH2).

[0428] Implementation Scheme 108. The macromolecular structure as described in any one of Implementation Schemes 98-102, wherein R 3 It is hydrogen.

[0429] Implementation Scheme 109. The macromolecular structure as described in any one of Implementation Schemes 98-102, wherein R 3’ It is hydrogen.

[0430] Implementation Scheme 110. The macromolecular structure as described in any one of Implementation Schemes 108 or 109, wherein Z is -N-.

[0431] Implementation Scheme 111. The macromolecular structure as described in any one of Implementation Schemes 108-110, wherein R 8 It is a C1-C6 alkyl group.

[0432] Scheme 112. The macromolecular structure as described in any one of Schemes 1-111, wherein the macromolecular chain comprises repeating units derived from monomers represented by the structures in Table 1.

[0433] Implementation Scheme 113. The macromolecular structure as described in any one of Implementation Schemes 1-112, wherein the macromolecular chain comprises repeating units derived from monomers represented by the structures in Table 2.

[0434] Implementation Scheme 114. The macromolecular structure as described in any one of Implementation Schemes 1-113, wherein the macromolecular structure has a structure represented by the following: , , , , , or .

[0435] Implementation Scheme 115. The macromolecular structure as described in any one of Implementation Schemes 1-114, wherein the surface comprises particles.

[0436] Implementation Scheme 116. The macromolecular structure as described in Implementation Scheme 115, wherein the particles are nanoparticles.

[0437] Implementation Scheme 117. The macromolecular structure as described in Implementation Scheme 115, wherein the particles are microparticles.

[0438] Implementation Scheme 118. The macromolecular structure as described in any one of Implementation Schemes 115-117, wherein the particles have a diameter of about 100 nm to about 500 nm.

[0439] Implementation Scheme 119. The macromolecular structure as described in any one of Implementation Schemes 115-118, wherein the particles have a diameter of about 100 nm to about 300 nm.

[0440] Implementation Scheme 120. The macromolecular structure as described in any one of Implementation Schemes 115-119, wherein the particles have a polydispersity index (PDI) of about 0.01 to about 1.

[0441] Implementation Scheme 121. The macromolecular structure as described in any one of Implementation Schemes 115-120, wherein the particles have a PDI of less than about 0.1.

[0442] Implementation Scheme 122. The macromolecular structure as described in any one of Implementation Schemes 115-121, wherein the particles comprise iron oxide.

[0443] Implementation Scheme 123. The macromolecular structure as described in any one of Implementation Schemes 115-122, wherein the particles have an iron oxide core.

[0444] Implementation Scheme 124. The macromolecule as described in any one of Implementation Schemes 115-123, wherein the particles are magnetic.

[0445] Implementation Scheme 125. The macromolecular structure as described in any one of Implementation Schemes 115-124, wherein the particles are superparamagnetic iron oxide particles.

[0446] Implementation Scheme 126. The macromolecular structure as described in any one of Implementation Schemes 115-125, wherein the particles have a core-shell structure.

[0447] Implementation Scheme 127. The macromolecular structure as described in any one of Implementation Schemes 115-126, wherein the particles comprise an iron oxide core and a silicon dioxide shell.

[0448] Implementation Scheme 128. The macromolecular structure as described in any one of Implementation Schemes 115-126, wherein the particles comprise iron oxide crystals embedded in a polystyrene core.

[0449] Implementation Scheme 129. The macromolecular structure as described in any one of Implementation Schemes 1-128, wherein the macromolecular structure comprises at least 10% w / w repeating units.

[0450] Implementation Scheme 130. The macromolecular structure as described in any one of Implementation Schemes 1-129, wherein the macromolecular structure comprises up to 50% w / w repeating units.

[0451] Implementation Scheme 131. The macromolecular structure as described in any one of Implementation Schemes 1-130, wherein the tethering portion is covalently coupled to the surface.

[0452] Implementation Scheme 132. The macromolecular structure as described in any one of Implementation Schemes 1-131, wherein the tethered portion is non-covalently coupled to the surface.

[0453] Implementation Scheme 133. The macromolecular structure as described in any one of Implementation Schemes 1-132, wherein the tethered moiety is a C1-C6 alkyl, oxo, halogenated, or hydroxylated moiety. 20 Heteroalkyl groups.

[0454] Implementation Scheme 134. The macromolecular structure as described in Implementation Scheme 133, wherein the tethered portion is represented by the following structure: .

[0455] Implementation Scheme 135. The macromolecular structure as described in any one of Implementation Schemes 1-132, wherein the tethered portion is represented by the following structure: .

[0456] Implementation Scheme 136. The macromolecular structure as described in any one of Implementation Schemes 1-132, wherein the tethering portion is optionally bounded by one or more C1-C1 bonds. 20 Heteroalkyl-substituted C1-C 12 Alkoxy, C1-C 12 Alkoxy and C1-C 20 Each of the heteroalkyl groups is optionally substituted with one or more C1-C6 alkyl, oxo, halogenated, or hydroxyl groups.

[0457] Implementation Scheme 137. The macromolecular structure as described in Implementation Scheme 135, wherein the tethered portion is represented by the following structure:

[0458] in: Y is a C1-C6 alkyl, oxo, halogenated, or hydroxyl-substituted C1-C6 alkyl group.20 Heteroalkyl; and p is an integer from 1 to 12.

[0459] Implementation Scheme 138. The macromolecular structure as described in Implementation Scheme 135 or 137, wherein the tethered portion is represented by the following structure: .

[0460] Implementation Scheme 139. The macromolecular structure as described in any one of Implementation Schemes 1-137, wherein the macromolecular chain is a homopolymer.

[0461] Implementation Scheme 140. The macromolecular structure as described in any one of Implementation Schemes 1-137, wherein the macromolecular chain is a block copolymer.

[0462] Implementation Scheme 141. The macromolecular structure as described in any one of Implementation Schemes 1-137, wherein the macromolecular chain is a random copolymer.

[0463] Implementation Scheme 142. The macromolecular structure as described in any one of Implementation Schemes 1-137, wherein the macromolecular chains are not cross-linked.

[0464] Implementation Scheme 143. The macromolecular structure as described in any one of Implementation Schemes 1-142, wherein the macromolecular chain comprises about 1 to about 100 repeating units.

[0465] Implementation Scheme 144. The macromolecular structure as described in any one of Implementation Schemes 1-143, wherein the macromolecular chain comprises about 1 to about 50 repeating units.

[0466] Implementation Scheme 145. The macromolecular structure as described in any one of Implementation Schemes 1-144, wherein the macromolecular chain has a molecular weight of about 0.5 kDa to about 25 kDa.

[0467] Implementation Scheme 146. The macromolecular structure as described in any one of Implementation Schemes 1-145, wherein the macromolecular chain has a molecular weight of about 0.5 kDa to about 10 kDa.

[0468] Implementation Scheme 147. The macromolecular structure as described in any one of Implementation Schemes 1-146, wherein the second end of the macromolecular chain is not coupled to the surface.

[0469] Implementation Scheme 148. A macromolecular structure as described in any one of Implementation Schemes 1-147, wherein the surface comprises at a density of 50 nm per 50 nm 2 The density of at least one tethered portion of the tethered portion.

[0470] Implementation Scheme 149. The macromolecular structure as described in any one of Implementation Schemes 1-148, wherein the surface comprises at a density of 50 nm per 50 nm 2 Approximately one tethered portion to every 5 nm 2 The density of the tethered portion is approximately one tethered portion.

[0471] Implementation Scheme 150. A method for preparing a macromolecular structure as described in any one of Implementation Schemes 1-149, the method comprising: (a) Provide a surface; (b) Coupling a polymer initiator to the surface to form an initiator surface; and (c) The surface of the initiator is contacted with a monomer as described in any one of embodiments 1-113 to form the macromolecular structure.

[0472] Implementation Scheme 151. The method as described in Implementation Scheme 150, wherein the surface comprises particles.

[0473] Implementation Scheme 152. The method as described in Implementation Scheme 150 or 151, wherein the particles are nanoparticles or microparticles.

[0474] Implementation Scheme 153. The method of any one of Implementation Schemes 150-152, wherein the particles have a diameter of about 100 nm to about 500 nm.

[0475] Implementation Scheme 154. The method of any one of Implementation Schemes 150-153, wherein the particles have a diameter of about 100 nm to about 300 nm.

[0476] Implementation Scheme 155. The method of any one of Implementation Schemes 150-154, wherein the particles comprise iron oxide.

[0477] Implementation Scheme 156. The method of any one of Implementation Schemes 150-155, wherein the particles have an iron oxide core.

[0478] Implementation Scheme 157. The method of any one of Implementation Schemes 150-156, wherein the particles are superparamagnetic iron oxide (nano) particles.

[0479] Implementation Scheme 158. The method of any one of Implementation Schemes 150-157, wherein the particles comprise silicon dioxide.

[0480] Implementation Scheme 159. The method of any one of Implementation Schemes 150-158, wherein the particles have a core-shell structure.

[0481] Implementation Scheme 160. The method of any one of Implementation Schemes 150-159, wherein the particles comprise an iron oxide core and a silicon dioxide shell.

[0482] Implementation Scheme 161. The method of any one of Implementation Schemes 150-160, wherein the surface is functionalized with alkoxysilane.

[0483] Implementation Scheme 162. The method of any one of Implementation Schemes 150-161, wherein the surface is functionalized with aminopropyltetraethoxysilane (APTES).

[0484] Implementation Scheme 163. The method of any one of Implementation Schemes 150-162, wherein coupling comprises adding a base.

[0485] Implementation Scheme 164. The method as described in Implementation Scheme 163, wherein the base comprises triethylamine.

[0486] Implementation Scheme 165. The method as described in Implementation Scheme 163 or 164, wherein the base is added at about 0°C.

[0487] Implementation Scheme 166. The method of any one of Implementation Schemes 150-165, wherein coupling comprises providing the polymer initiator to the surface at about 0°C.

[0488] Implementation Scheme 167. The method of any one of Implementation Schemes 150-166, wherein the polymer initiator is represented by the following structure:

[0489] in: X is a halogen; R 10 It is an initiator group.

[0490] Implementation Scheme 168. The method of implementation scheme 167, wherein the initiator group is a halogen.

[0491] Implementation Scheme 169. The method of any one of Implementation Schemes 150-168, wherein the contact comprises a temperature of at least 25°C.

[0492] Implementation Scheme 170. The method of any one of Implementation Schemes 150-169, wherein the contact comprises a temperature of about 25°C to about 75°C.

[0493] Implementation Scheme 171. The method of any one of Implementation Schemes 150-170, wherein contacting comprises contacting the initiator surface with a mixture of the monomer and the organic solvent.

[0494] Implementation Scheme 172. The method as described in Implementation Scheme 171, wherein the organic solvent is dimethylformamide.

[0495] Implementation Scheme 173. A method for preparing a macromolecular structure as described in any one of Implementation Schemes 1-149, the method comprising: (a) Provide a surface; (b) Couple vinyl groups to the surface to form a vinyl-functionalized surface; (c) The vinyl-functionalized surface is contacted with a crosslinking monomer and a monomer selected from hydroxyalkyl methacrylate, aminoalkyl methacrylate, alkynyl methacrylate, glycidyl methacrylate, hydroxyalkyl acrylate, aminoalkyl acrylate, alkynyl acrylate, or glycidyl acrylate to form a crosslinked polymer coupled to the surface. (d) Coupling a polymer initiator to the crosslinked polymer to form an initiator surface; (e) The surface of the initiator is contacted with a monomer as described in any one of embodiments 1-113 to form the macromolecular structure.

[0496] Implementation Scheme 174. The method as described in Implementation Scheme 173, wherein the surface comprises particles.

[0497] Implementation Scheme 175. The method as described in Implementation Scheme 173 or 174, wherein the particles are nanoparticles or microparticles.

[0498] Implementation Scheme 176. The method of any one of Implementation Schemes 173-175, wherein the particles have a diameter of about 100 nm to about 500 nm.

[0499] Implementation Scheme 177. The method of any one of Implementation Schemes 173-176, wherein the particles have a diameter of about 100 nm to about 300 nm.

[0500] Implementation Scheme 178. The method of any one of Implementation Schemes 173-177, wherein the particles comprise iron oxide.

[0501] Implementation Scheme 179. The method of any one of Implementation Schemes 173-178, wherein the particles have an iron oxide core.

[0502] Implementation Scheme 180. The method of any one of Implementation Schemes 173-179, wherein the particles are magnetic.

[0503] Implementation Scheme 181. The method of any one of Implementation Schemes 173-180, wherein the particles are superparamagnetic iron oxide (nano) particles.

[0504] Implementation Scheme 182. The method of any one of Implementation Schemes 173-181, wherein the particles have a core-shell structure.

[0505] Implementation Scheme 183. The method of any one of Implementation Schemes 173-182, wherein the particles comprise an iron oxide core and a silicon dioxide shell.

[0506] Implementation Scheme 184. The method of any one of Implementation Schemes 173-183, wherein the vinyl-functionalized surface comprises vinyl acrylate.

[0507] Implementation Scheme 185. The method of any one of Implementation Schemes 173-184, wherein the crosslinking monomer comprises... The structure represented, in: R 1 R 2 R 3 R 1’ R 2’ and R 3’ Each of them is independently selected from hydrogen or -C1-C6 alkyl; Z is -O- or -NH; and R 8 It is a -C1-C6 alkyl, divalent metal, or symmetrical or asymmetrical disulfide.

[0508] Implementation Scheme 186. The method of any one of Implementation Schemes 173-185, wherein the polymer initiator is represented by the following structure:

[0509] in: X is a halogen; R 10 It is an initiator group.

[0510] Implementation Scheme 187. The method of Implementation Scheme 186, wherein the initiator group is selected from halogens, epoxides, or double bonds.

[0511] Implementation Scheme 188. The method of any one of Implementation Schemes 173-187, wherein the contact comprises a temperature of at least 25°C.

[0512] Implementation Scheme 189. The method of any one of Implementation Schemes 173-188, wherein the contact comprises a temperature of about 25°C to about 75°C.

[0513] Implementation Scheme 190. The method of any one of Implementation Schemes 173-189, wherein contacting comprises contacting the initiator surface with a mixture of the monomer, the crosslinking monomer and the organic solvent.

[0514] Implementation Scheme 191. The method as described in Implementation Scheme 190, wherein the organic solvent is dimethylformamide.

[0515] Implementation Scheme 192. The method of any one of Implementation Schemes 173-191, wherein the contact includes an inert condition.

[0516] Implementation scheme 193. The method of any one of implementation schemes 173-192, wherein the coupling includes an inert condition.

[0517] Implementation Scheme 194. A composition comprising a macromolecular structure as described in any one of Implementation Schemes 1-149 and a biomolecule adsorbed onto the macromolecular structure.

[0518] Implementation Scheme 195. The composition as described in Implementation Scheme 194, wherein the biomolecules adsorbed onto the macromolecular structure form a biomolecular crown on the macromolecular structure.

[0519] Implementation Scheme 196. The composition as described in Implementation Scheme 194 or 195, wherein the composition comprises a biological sample in contact with the macromolecular structure.

[0520] Embodiment 197. The composition as described in Embodiment 196, wherein the biological sample comprises plasma, serum, or blood.

[0521] Embodiment 198. The composition as described in Embodiment 196 or 197, wherein the biological sample comprises a plurality of proteins.

[0522] Implementation Scheme 199. The composition of any one of Implementation Schemes 194-198, wherein at least 100 different biomolecules are adsorbed onto the macromolecular structure.

[0523] Implementation Scheme 200. The composition of any one of Implementation Schemes 194-199, wherein about 100 to about 1000 different proteins are adsorbed onto the macromolecular structure.

[0524] Implementation Scheme 201. The composition of any one of Implementation Schemes 194-200, wherein the biomolecule comprises a protein.

[0525] Implementation Scheme 202. A method for identifying proteins in a sample, the method comprising: (a) Incubating one or more macromolecular structures as described in any one of embodiments 1-113 together with a biological sample containing biomolecules to form a biomolecular crown; (b) isolating at least a portion of the biomolecules in the biomolecular crown; and (c) Determine the biomolecule crown.

[0526] Implementation Scheme 203. The method of implementation scheme 202, wherein the assay is capable of identifying 1 to 20,000 protein groups.

[0527] Implementation Scheme 204. The method as described in Implementation Scheme 202 or 203, wherein the assay is capable of identifying 1,000 to 10,000 protein groups.

[0528] Implementation Scheme 205. The method of any one of Implementation Schemes 202-204, wherein the assay is capable of identifying 1,000 to 5,000 protein groups.

[0529] Implementation Scheme 206. The method of any one of Implementation Schemes 202-205, wherein the assay is capable of identifying 1,200 to 2,200 protein groups.

[0530] Implementation Scheme 207. The method of any one of Implementation Schemes 202-206, wherein the protein group comprises a peptide sequence of minimum length having 7 amino acid residues.

[0531] Implementation Scheme 208. The method of any one of Implementation Schemes 202-207, wherein the assay is capable of identifying 1,000 to 10,000 proteins.

[0532] Implementation Scheme 209. The method of any one of Implementation Schemes 202-208, wherein the assay is capable of identifying 1,800 to 5,000 proteins.

[0533] Implementation Scheme 210. The method of any one of Implementation Schemes 202-209, wherein the sample comprises a plurality of samples.

[0534] Implementation Scheme 211. The method of any one of Implementation Schemes 202-210, wherein the plurality of samples comprises at least two or more spatially isolated samples.

[0535] Implementation Scheme 212. The method of any one of Implementation Scheme 211, wherein the incubation comprises simultaneously contacting the at least two or more spatially isolated samples with the one or more macromolecular structures.

[0536] Implementation Scheme 213. The method as described in Implementation Scheme 211 or 212, wherein isolation comprises simultaneously magnetically isolating the one or more macromolecular structures from unbound proteins in at least two or more spatially isolated samples of the plurality of samples.

[0537] Implementation Scheme 214. The method of any one of Implementation Schemes 211-213, wherein the assay comprises measuring a plurality of unique biomolecular crowns to simultaneously identify proteins in the at least two or more spatially isolated samples.

[0538] Implementation Scheme 215. The method of any one of Implementation Schemes 202-214, further comprising repeating wherein, when repeated, the incubation, isolation, and determination produce a 20% or lower percentage quantile normalized coefficient of variation (QNCV), as determined by comparing peptide mass spectrometry characteristics from at least three complete determination repeats of each of the one or more macromolecular structures.

[0539] Implementation Scheme 216. The method of any one of Implementation Schemes 202-215, wherein, when repeated, the incubation, isolation, and determination produce a percentage quantile normalized coefficient of variation (QNCV) of 10% or less, as determined by comparing peptide mass spectrometry characteristics from at least three complete determinations of each of the one or more macromolecular structures.

[0540] Implementation Scheme 217. The method of any one of Implementation Schemes 202-216, wherein the assay is capable of identifying proteins within a dynamic range of at least 6, at least 7, at least 8, at least 9, or at least 10.

[0541] Implementation Scheme 218. The method of any one of Implementation Schemes 202-217, further comprising washing the one or more macromolecular structures at least once or at least twice after isolating the one or more macromolecular structures from the unbound proteins.

[0542] Implementation Scheme 219. The method of any one of Implementation Schemes 202-218, wherein after the determination, the method further comprises cleaving the proteins in the plurality of unique biomolecular crowns.

[0543] Implementation Scheme 220. The method of any one of Implementation Schemes 202-219 further includes digesting the proteins in the plurality of unique biomolecular crowns to generate digested peptides.

[0544] Implementation Scheme 221. The method of any one of Implementation Schemes 202-220, further comprising purifying the digested peptide.

[0545] Implementation Scheme 222. The method of any one of Implementation Schemes 202-221, wherein the determination includes identifying proteins in the sample using mass spectrometry.

[0546] Implementation Scheme 223. The method of any one of Implementation Schemes 202-222, wherein the determination is performed over about 2 to about 4 hours.

[0547] Implementation Scheme 224. The method as described in any one of Implementation Schemes 202-223, wherein the method is performed over an hour of about 1 to about 20 hours.

[0548] Implementation Scheme 225. The method as described in any one of Implementation Schemes 202-224, wherein the method is performed over a period of about 2 to about 10 hours.

[0549] Implementation Scheme 226. The method as described in any one of Implementation Schemes 202-225, wherein the method is performed over a period of about 4 to about 6 hours.

[0550] Implementation Scheme 227. The method of any one of Implementation Schemes 202-226, wherein the isolation takes no more than about 30 minutes, no more than about 15 minutes, no more than about 10 minutes, no more than about 5 minutes, or no more than about 2 minutes.

[0551] Implementation Scheme 228. The method of any one of Implementation Schemes 202-227, wherein the plurality of samples comprises at least 10 spatially isolated samples, at least 50 spatially isolated samples, at least 100 spatially isolated samples, at least 150 spatially isolated samples, at least 200 spatially isolated samples, at least 250 spatially isolated samples, or at least 300 spatially isolated samples.

[0552] Implementation Scheme 229. The method of any one of Implementation Schemes 202-228, wherein the plurality of samples comprises at least 96 samples.

[0553] Implementation Scheme 230. The method of any one of Implementation Schemes 202-229, wherein the one or more macromolecular structures comprise at least 2 unique macromolecular structures, at least 3 unique macromolecular structures, at least 4 unique macromolecular structures, at least 5 unique macromolecular structures, at least 6 unique macromolecular structures, at least 7 unique macromolecular structures, at least 8 unique macromolecular structures, at least 9 unique macromolecular structures, at least 10 unique macromolecular structures, at least 11 unique macromolecular structures, at least 12 unique macromolecular structures, at least 13 unique macromolecular structures, at least 14 unique macromolecular structures, at least 15 unique macromolecular structures, at least 20 unique macromolecular structures, at least 25 macromolecular structures, or at least 30 unique macromolecular structures.

[0554] Implementation Scheme 231. The method of any one of Implementation Schemes 202-230, wherein the one or more macromolecular structures comprise at least 10 unique macromolecular structures.

[0555] Implementation Scheme 232. The method of any one of Implementation Schemes 211-231, wherein the at least two spatially isolated samples are different in at least one physicochemical property.

[0556] Implementation Scheme 233. The method of any one of Implementation Schemes 202-232, wherein the one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure share at least one physicochemical property and are different in at least one physicochemical property, such that the first unique macromolecular structure and the second unique macromolecular structure are distinct.

[0557] Implementation Scheme 234. The method of any one of Implementation Schemes 202-233, wherein the one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure share at least two physicochemical properties and are different in at least two physicochemical properties, such that the first unique macromolecular structure and the second unique macromolecular structure are distinct.

[0558] Implementation Scheme 235. The method of any one of Implementation Schemes 202-233, wherein the one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure share at least one physicochemical property and are different in at least two physicochemical properties, such that the first unique macromolecular structure and the second unique macromolecular structure are distinct.

[0559] Implementation Scheme 236. The method of any one of Implementation Schemes 202-233, wherein the one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure share at least two physicochemical properties and are different in at least one physicochemical property, such that the first unique macromolecular structure and the second unique macromolecular structure are distinct.

[0560] Implementation Scheme 237. The method of any one of Implementation Schemes 202-233, wherein the one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure comprise a carboxylic acid ester material, wherein the first unique particle is a microparticle, and wherein the second unique macromolecular structure is a nanoparticle.

[0561] Implementation Scheme 238. The method of any one of Implementation Schemes 202-233, wherein the one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure have a surface charge from 0 mV to -50 mV, wherein the first unique macromolecular structure has a diameter of less than 200 nm, and wherein the second unique macromolecular structure has a diameter of greater than 200 nm.

[0562] Implementation Scheme 239. The method of any one of Implementation Schemes 202-233, wherein the one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure have a diameter of 100 to 400 nm, wherein the first unique macromolecular structure has a positive surface charge, and wherein the second unique macromolecular structure has a neutral surface charge.

[0563] Implementation Scheme 240. The method of any one of Implementation Schemes 202-233, wherein the one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure are nanoparticles, wherein the first unique macromolecular structure has a surface charge of less than -20 mV and the second unique macromolecular structure has a surface charge of greater than -20 mV.

[0564] Implementation Scheme 241. The method of any one of Implementation Schemes 202-233, wherein the one or more macromolecular structures comprise a first unique macromolecular structure and a second unique macromolecular structure, wherein the first unique macromolecular structure and the second unique macromolecular structure are microparticles, wherein the first unique macromolecular structure has a negative surface charge, and wherein the second unique macromolecular structure has a positive surface charge.

[0565] Implementation Scheme 242. The method of any one of Implementation Schemes 202-241, wherein the one or more macromolecular structures comprise a subset of negatively charged nanoparticles, wherein each type of particle in the subset is distinct on at least one surface chemical group.

[0566] Implementation Scheme 243. The method of any one of Implementation Schemes 202-242, wherein the one or more macromolecular structures comprise a first unique macromolecular structure, a second particle, and a third unique macromolecular structure, wherein the first unique macromolecular structure, the second unique macromolecular structure, and the third unique macromolecular structure comprise an iron oxide core, a polymer shell, and have a diameter less than about 500 nm, wherein the first unique macromolecular structure has a negative charge, the second unique macromolecular structure has a positive charge, and the third unique macromolecular structure has a neutral charge, wherein the diameter is an average diameter as measured by dynamic light scattering.

[0567] Implementation Scheme 244. The method of any one of Implementation Schemes 202-243, wherein at least one unique macromolecular structure of the one or more macromolecular structures is a nanoparticle.

[0568] Implementation Scheme 245. The method of any one of Implementation Schemes 202-244, wherein at least one unique macromolecular structure of the one or more macromolecular structures is a microparticle.

[0569] Implementation Scheme 246. The method of any one of Implementation Schemes 202-245, wherein at least one unique macromolecular structure of the one or more macromolecular structures is superparamagnetic iron oxide particles.

[0570] Implementation Scheme 247. The method of any one of Implementation Schemes 202-246, wherein each particle of the one or more macromolecular structures comprises an iron oxide material.

[0571] Implementation Scheme 248. The method of any one of Implementation Schemes 202-247, wherein at least one unique macromolecular structure of the one or more macromolecular structures has an iron oxide core.

[0572] Implementation Scheme 249. The method of any one of Implementation Schemes 202-248, wherein at least one unique macromolecular structure of the one or more macromolecular structures has iron oxide crystals embedded in a polystyrene core.

[0573] Implementation Scheme 250. The method of any one of Implementation Schemes 202-249, wherein each of the unique macromolecular structures of the one or more macromolecular structures is a superparamagnetic iron oxide particle.

[0574] Implementation Scheme 251. The method of any one of Implementation Schemes 202-250, wherein each of the one or more unique macromolecular structures comprises an iron oxide core.

[0575] Implementation Scheme 252. The method of any one of Implementation Schemes 202-251, wherein each of the one or more unique macromolecular structures has an iron oxide crystal embedded in a polystyrene core.

[0576] Implementation Scheme 253. The method of any one of Implementation Schemes 202-252, wherein at least one unique macromolecular structure of one or more macromolecular structures comprises a carboxylated polymer, an amination polymer, a zwitterionic polymer, or any combination thereof.

[0577] Implementation Scheme 254. The method of any one of Implementation Schemes 202-253, wherein at least one macromolecular structure of the one or more macromolecular structures comprises an iron oxide core and a silicon dioxide outer shell coating.

[0578] Implementation Scheme 255. The method of any one of Implementation Schemes 202-254, wherein at least one unique macromolecular structure of the one or more macromolecular structures has a negative surface charge.

[0579] Implementation Scheme 256. The method of any one of Implementation Schemes 202-255, wherein at least one unique macromolecular structure of the one or more macromolecular structures has a positive surface charge.

[0580] Implementation Scheme 257. The method of any one of Implementation Schemes 202-256, wherein at least one unique macromolecular structure of the one or more macromolecular structures has a neutral surface charge.

[0581] Implementation Scheme 258. A kit for identifying molecules in biological samples, the kit comprising one or more macromolecular structures as described in Implementation Schemes 1-113.

[0582] Implementation Scheme 259. The kit as described in Implementation Scheme 258, wherein the kit further comprises a lysis agent.

[0583] Implementation Scheme 260. The kit as described in Implementation Scheme 258 or 259, wherein the kit further comprises a digestive agent.

[0584] Implementation Scheme 261. The kit as described in any one of Implementation Schemes 258-260, wherein the kit further comprises a detergent.

[0585] Implementation Scheme 262. The kit as described in any one of Implementation Schemes 258-261, wherein the kit further comprises an elution buffer.

[0586] Implementation Scheme 263. The kit as described in any one of Implementation Schemes 258-262, wherein the kit further comprises a solid support for solid-phase extraction.

[0587] Implementation Scheme 264. The kit as described in any one of Implementation Schemes 258-263, wherein the kit further comprises a multiwell plate.

[0588] Implementation Scheme 265. The kit as described in any one of Implementation Schemes 258-264, wherein the kit further comprises a diluent.

[0589] Implementation Scheme 266. A system for identifying biomolecules in a biological sample, the system comprising: (a) The macromolecular structure as described in any one of embodiments 1-113; (b) Suspension solution; (c) Biological samples containing a certain concentration of protein; and (d) An automated system comprising a network of differentiated functional units for isolating biomolecules adsorbed to the macromolecular structure, wherein the automated system is programmed to perform a series of steps.

[0590] Implementation Scheme 267. The system as described in Implementation Scheme 266, wherein the network of said unit includes: (a) A first unit, comprising a multi-channel fluid transfer device for transferring fluid between units within the system; (b) A second unit, which includes a support for storing multiple biological samples; (c) A third unit comprising a support for a sensor array plate having partitions, the partitions comprising the macromolecular structures for binding and interacting with a population of analytes from the biological sample; (d) The fourth unit includes a support for storing a variety of reagents; (e) A fifth unit, comprising a support for storing reagents to be disposed of; and (f) A sixth unit, which includes a support for storing consumables used by the multichannel fluid transfer device.

[0591] Implementation Scheme 268. The system as described in Implementation Scheme 266 or 267, wherein the series of steps includes: (a) Contact the biological sample with a designated section of the sensor array; (b) Incubate the biological sample together with the macromolecular structure contained within a partition of the sensor array plate; (c) Remove the supernatant from the sensor array plate; and (d) Preparation of biomolecules adsorbed onto the macromolecular structure for mass spectrometry.

[0592] Implementation Scheme 269. The system as described in any one of Implementation Schemes 266-268, wherein i.-iii. is incubated at a temperature of about 20 degrees Celsius to about 80 degrees Celsius.

[0593] Implementation Scheme 270. The system of any one of Implementation Schemes 266-269, wherein the suspension comprises Tris EDTA 150 mM KCl and 0.05% CHAPS buffer.

[0594] Implementation Scheme 271. The system of any one of Implementation Schemes 266-270, wherein the suspension comprises 10 mM Tris HCl pH 7.4 and 1 mM EDTA.

[0595] While preferred embodiments of this disclosure have been shown and described herein, it will be apparent to those skilled in the art that these embodiments are provided by way of example only. Various changes, modifications, and substitutions will now occur to those skilled in the art without departing from this disclosure. It should be understood that various alternatives to the embodiments of this disclosure described herein may be employed in the practice of the disclosure. The appended claims are intended to define the scope of this disclosure and to cover methods and structures within the scope of these claims and their equivalents. Example

[0596] Example 1: Synthesis of SPION@SiO2-APTES

[0597] Silica-coated superparamagnetic iron oxide nanoparticles (SPION@SiO2) were suspended in DMF and sonicated for 15 minutes. The nanoparticle solution was purged with N2 gas for 20 minutes and then heated to 120°C for 4 hours. After cooling, the SPION@SiO2 was washed three times with DMF and then isolated. The isolated particles were dispersed in N,N-dimethylacetamide (DMAc) (2 L). 4.50 g of (3-aminopropyltriethoxysilane) (APTES) was added to the solution, and the resulting solution was reacted at 120°C for 4 hours to obtain the SPION@SiO2-APTES product.

[0598] Example 2: Synthesis of SPION@SiO2-APTES-Br

[0599] SPION@SiO2-APTES in DMF was washed twice with tetrahydrofuran (THF) and then resuspended in THF (801 mL) while sonicating for 15 min. Triethylamine (10.40 g) was added under N2 in an ice bath (0 °C). After the addition of triethylamine, 2-bromoisobutyryl bromide (2.509 g) was added dropwise at 0 °C. The reaction was stirred overnight (16 h) at room temperature. The resulting material was washed once with THF, once with ethanol, once with water, and then once with THF. The final product was dried over N2.

[0600] Example 3: Synthesis of Compound 2 (SPION@SiO2-SIP-POEGMA)

[0601] Initiator particles SPION@SiO2-APTES-Br (1.000 g), CuBr2 (0.010 g), N,N,N',N'',N''-pentamethyldiethylenetriamine (PMDETA) (0.240 g), monomer oligo(ethylene glycol) methyl ether methacrylate (OEGMA, MW500) (5.000 g) and 25 mL of dimethylformamide (DMF) were added to a three-necked flask. The mixture was sonicated for 15 min and purged with N2 for another 15 min. 0.500 g of L-ascorbic acid was dissolved separately in 10 mL of DMF and purged with N2 for at least 15 min. The mixture containing the monomer and nanoparticle suspension was heated to 35 °C under N2, and the L-ascorbic acid solution was added via a syringe pump at 0.05 mL / min. The reaction mixture was maintained at 35 °C for 16 h to obtain compound 2.

[0602] Example 4: Characterization of macromolecular structure The synthesis and according to the methods described in Examples 1-3 Figure 1 An improved version of the scheme described herein synthesizes compounds 1-3. An exemplary synthetic scheme for compound 1 is described in... Figure 4 As shown in [the image]. An exemplary synthetic scheme for compound 2 is [described in the image]. Figure 5 As shown in [the image]. An exemplary synthetic scheme for compound 3 is shown in [the image]. Figure 6 The results are shown in Table 4. Particle size was characterized by DLS, surface charge by zeta potential, and macromolecular chain percentage by TGA (weight loss %). Particle size was compared with that before macromolecular chain functionalization.

[0603] Table 4

[0604] Compounds 5, 6, and 7 were also synthesized using the general procedures of Examples 1-3. The percentage of organic matter in compounds 1, 2, 5, 6, and 7 was analyzed by thermogravimetric analysis. The results are shown in Table 5. The percentage of organic matter in the particles ranged from 9.14% to 15.88%, with the increase originating from the percentage of organic matter found on the initiator surface prior to further functionalization.

[0605] Table 5

[0606] Scanning electron microscope (SEM) images of compounds 2, 5, 6, and 7 are shown in [the image]. Figure 3A The results show that compound 5 has a size of 296 nm according to SEM, compound 6 has a size of 230 nm according to SEM, compound 2 has a size of 311 nm according to SEM, and compound 7 has a size of 302 nm according to SEM. Figure 3B Images of compound 2 obtained by transmission electron microscopy are shown, highlighting the polymer brushes formed on the surface of the particles.

Claims

1. A macromolecular structure, said macromolecular structure comprising: (I) Surface; (II) The tethered portion coupled to the surface; and (III) A macromolecular chain, wherein the first end of the macromolecular chain is covalently attached to the tethered portion, and wherein the macromolecular chain comprises two or more distinct repeating units derived from monomers selected from the following structures: , , , , , , , and Each of X and Y is independently -C-, -O-, or -N-; Each Z is independently -O- or -NH-; Q is -CH2- or ethylene glycol; A is a polymer side chain comprising repeating units derived from monomers represented by the following structures: ； m is an integer selected from 1 to 20; Is it a single bond or a double bond? R 1 R 2 R 1’ R 2’ and R 3’ Each of them is independently selected from hydrogen or -C1-C6 alkyl; R 3 It is a hydrogen, a C1-C6 alkyl group, or a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl groups; R 4 It is absent, hydrogen, sulfonate, carboxylic acid ester, C1-C4 alkylene, amine, quaternary ammonium cation or C1-C6 alkyl optionally substituted with halogen; R 5 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, an amine, an azide, a sulfonate, a carbamate, an asymmetric disulfide, or a C1-C8 alkyl group or a C1-C6 heterocyclic group substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines optionally further substituted with amine, hydroxyl, aryl or sulfonate, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogens, C1-C8 alkyl groups optionally substituted with one or more pyrene, two or more fused 5-6 membered rings optionally further substituted, -C1-C3 alkyl groups optionally substituted with benzyl, trimethoxysilane or phosphocholine, C1-C 12 Alkylamines or C1-C4 alkylamines; R 6 It is hydrogen or C1-C6 alkyl. R 7 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, amines, azides, sulfonates, carbamates, or asymmetric disulfides, or a 3-, 5-, or 6-membered heterocyclic or C1-C alkyl group optionally substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines optionally further substituted with amines or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogens, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted, optionally substituted benzyl groups, -C1-C3 alkyl or C1-C4 alkylyl groups substituted with trimethoxysilane or phosphocholine; R 8 It is a C1-C6 alkyl group, a divalent metal, or a symmetrical or asymmetrical disulfide; R 9 It is hydrogen or oxygen; and n 1 It is an integer selected from 1 to 100.

2. The macromolecular structure as described in claim 1, wherein R 3 It is a methyl group.

3. The macromolecular structure as described in claim 2, wherein R 5 It is C1-C n1 Ethylene glycol.

4. A macromolecular structure, said macromolecular structure comprising: (I) Surface; (II) The tethered portion coupled to the surface; as well as (III) A macromolecular chain, wherein the first end of the macromolecular chain is covalently attached to the tethered portion, and wherein the macromolecular chain comprises repeating units derived from monomers selected from the following structures: , , , , , , , and Each of X and Y is independently -C-, -O-, or -N-; Each Z is independently -O- or -NH; Q is -CH2- or ethylene glycol; A is a polymer side chain comprising repeating units derived from monomers represented by the following structures: ； m is 1-6; R 1 R 2 R 1’ R 2’ and R 3’ Each of them is independently selected from hydrogen or C1-C6 alkyl. R 3 It is a hydrogen, a C1-C6 alkyl group, or a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl groups; R 4 It is absent, hydrogen, C1-C6 alkyl, C1-C4 alkylene, C1-C6 alkyl, sulfonate, amine, quaternary ammonium cation or carboxylic acid ester; R 5 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, an amine, an azide, a sulfonate, a carbamate, an asymmetric disulfide, or a C1-C8 alkyl group or a C1-C6 heterocyclic group substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines further substituted with amines, hydroxyl groups, aryl groups, or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogen groups, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted (e.g., two or more fused 6 membered rings optionally further substituted), optionally substituted benzyl groups, trimethoxysilanes, or phosphoric acid choline-substituted -C1-C3 alkyl groups, C1-C 12 Alkylamines or C1-C4 alkylamines; R 6 It is hydrogen or a linear C1-C6 alkyl group. R 7 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, amines, azides, sulfonates, carbamates, or asymmetric disulfides, or a 3-, 5-, or 6-membered heterocyclic or C1-C alkyl group optionally substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines further substituted with amines or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogen groups, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted (e.g., two or more fused 6 membered rings optionally further substituted), optionally substituted benzyl groups, -C1-C3 alkyl or C1-C4 alkylyl groups substituted with trimethoxysilane or phosphocholine; and R 8 It is a C1-C6 alkyl or symmetrical disulfide or asymmetrical disulfide; R 9 It is hydrogen or oxygen; n 1 It is an integer selected from 1 to 100; and The condition is when R 3 It is CH3, R 4 When it is CH3 or when R 5 When the C1-C8 alkyl group is substituted with a hydroxyl group, the C1-C8 groups are further substituted.

5. A macromolecular structure, said macromolecular structure comprising: (I) a surface and (II) a macromolecular chain coupled to the surface, wherein the macromolecular chain comprises repeating units of formula (I): （I） Where R 1” R 2” and R 3” Each of them is independently hydrogen or a C1-C6 alkyl group; L represents the connector section; A is a polymer side chain comprising repeating units derived from monomers selected from the following structures: , , , , , , and Each of X and Y is independently -C-, -O-, or -N-; Z is -O- or -NH; R 1 R 2 R 1’ R 2’ and R 3’ Each of them is independently selected from hydrogen or C1-C6 alkyl; R 3 It is a hydrogen, a C1-C6 alkyl group, or a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl groups; R 4 It is absent, hydrogen, sulfonate, carboxylic acid ester, C1-C4 alkylene, amine, quaternary ammonium cation or C1-C6 alkyl optionally substituted with halogen; R 5 It is a C1-C6 alkyl group, a C1-C8 alkyl group substituted with one or more hydroxyl groups, an amine, an azide, a sulfonate, a carbamate, an asymmetric disulfide, or a C1-C8 alkyl group or a C1-C6 heterocyclic group substituted with one or more C1-C6 alkyl groups or oxo groups. n1 Ethylene glycol, C1-C8 alkylamines further substituted with amines, hydroxyl groups, aryl groups, or sulfonates, C1-C8 alkoxy groups optionally substituted with one or more oxo or halogen groups, C1-C8 alkyl groups optionally substituted with one or more pyrene groups, two or more fused 5-6 membered rings optionally further substituted (e.g., two or more fused 6 membered rings optionally further substituted), optionally substituted benzyl groups, trimethoxysilanes, or phosphoric acid choline-substituted -C1-C3 alkyl groups, C1-C 12 Alkylamines or C1-C4 alkylamines; R 6 It is hydrogen or a linear C1-C6 alkyl group. R 7 It is a hydrogen, C1-C6 alkyl, a 3-, 5-, or 6-membered heterocycle optionally substituted with one or more C1-C6 alkyl or oxo compounds, a substituted benzene, a C1-C6 alkyl substituted with a hydroxyl group, or a C1-C8 alkyl sulfonate optionally substituted. R 8 It is a C1-C6 alkyl group, a divalent metal, or a symmetrical or asymmetrical disulfide; R 9 It is hydrogen or oxygen; n 1 It is an integer selected from 1 to 100; and n is an integer selected from 1 to 10,000.

6. The macromolecular structure according to any one of claims 1-5, wherein the macromolecular chain comprises repeating units derived from monomers represented by the following structures: .

7. The macromolecular structure according to any one of claims 1-6, wherein the macromolecular chain comprises repeating units derived from monomers represented by the structures in Table 1.

8. The macromolecular structure according to any one of claims 1-7, wherein the macromolecular chain comprises repeating units derived from monomers represented by the structures in Table 2.

9. The macromolecular structure according to any one of claims 1-8, wherein the macromolecular structure has a structure represented by the following: , , , , , or .

10. A method for preparing a macromolecular structure as described in any one of claims 1-9, the method comprising: (a) Provide a surface; (b) Coupling a polymer initiator to the surface to form an initiator surface; as well as (c) Contacting the surface of the initiator with the monomer as described in any one of claims 7-8 to form the macromolecular structure.

11. The method of claim 10, wherein the surface comprises particles.

12. The method of claim 11, wherein the particles have a diameter of about 100 nm to about 500 nm.

13. A method for preparing a macromolecular structure as described in any one of claims 1-9, the method comprising: (a) Provide a surface; (b) Couple vinyl groups to the surface to form a vinyl-functionalized surface; (c) The vinyl-functionalized surface is contacted with a crosslinking monomer and a monomer selected from hydroxyalkyl methacrylate, aminoalkyl methacrylate, alkynyl methacrylate, glycidyl methacrylate, hydroxyalkyl acrylate, aminoalkyl acrylate, alkynyl acrylate, or glycidyl acrylate to form a crosslinked polymer coupled to the surface. (d) Coupling a polymer initiator to the crosslinked polymer to form an initiator surface; as well as (e) Contact the surface of the initiator with the monomer as described in any one of claims 7-8 to form the macromolecular structure.

14. The method of claim 13, wherein the polymer initiator is represented by the following structure: in: X is a halogen; and R 10 It is an initiator group.

15. A composition comprising a macromolecular structure as described in any one of claims 1-14 and a biomolecule adsorbed onto the macromolecular structure.

16. The composition of claim 15, wherein at least 100 different biomolecules are adsorbed onto the macromolecular structure.

17. A method for identifying proteins in a sample, the method comprising: (a) Incubating one or more macromolecular structures as described in any one of claims 1-9 together with a biological sample containing biomolecules to form a biomolecular crown; (b) Isolating at least a portion of the biomolecule in the biomolecular crown; and (c) Determine the biomolecule crown.

18. The method of claim 17, wherein the assay is capable of identifying 1 to 20,000 protein groups.

19. A kit for identifying molecules in biological samples, the kit comprising one or more macromolecular structures as described in any one of claims 1-9.

20. A system for identifying biomolecules in a biological sample, the system comprising: (a) The macromolecular structure as described in any one of claims 1-9; (b) Suspension solution; (c) Biological samples containing a certain concentration of protein; as well as (d) An automated system comprising a network of differentiated functional units for isolating biomolecules adsorbed to the macromolecular structure, wherein the automated system is programmed to perform a series of steps.

Images