Retentate chromatography and protein chip arrays with applications in biology and medicine

• US 7,575,935 B2
• Filed: 09/25/2006
• Issued: 08/18/2009
• Est. Priority Date: 06/20/1997
• Status: Expired due to Term

• Current Assignee: Bio-Rad Laboratories Incorporated
• Original Assignee: Bio-Rad Laboratories Incorporated
• Inventors: Hutchens, T. William, Yip, Tai-Tung
• Primary Examiner(s): Chin; Christopher L
• Application Number:
  US11/527,266
• Publication Number:
  US 20090181414A1
• Time in Patent Office: 1,058 Days
• Field of Search: 435/7.4, 435/176, 435/180, 435/181, 436/173, 436/518, 436/524
• US Class Current: 436/173
• CPC Class Codes: B01D 15/1878 for multi-dimensional chromatography
• CPC Class Codes: B01D 15/26 characterised by the separation mechanism
• CPC Class Codes: B01D 15/305 Hydrophilic interaction chromatography [HILIC]
• CPC Class Codes: B01D 15/322 Normal bonded phase
• CPC Class Codes: B01D 15/327 with hydrophobic interaction
• CPC Class Codes: B01D 15/362 Cation-exchange
• CPC Class Codes: B01D 15/363 Anion-exchange
• CPC Class Codes: B01D 15/38 involving specific interaction not covered by one or more of groups B01D15/265 - B01D15/36
• CPC Class Codes: B01D 15/3804 Affinity chromatography
• CPC Class Codes: B01D 15/3809 of the antigen-antibody type, e.g. protein A, G, L chromatography
• CPC Class Codes: B01J 20/281 Sorbents specially adapted for preparative, analytical or investigative chromatography
• CPC Class Codes: B01J 20/286 Phases chemically bonded to a substrate, e.g. to silica or to polymers
• CPC Class Codes: B01J 20/287 Non-polar phases; Reversed phases
• CPC Class Codes: B01J 20/288 Polar phases
• CPC Class Codes: B01J 20/289 bonded via a spacer
• CPC Class Codes: B01J 20/3204 Inorganic carriers, supports or substrates
• CPC Class Codes: B01J 20/3212 consisting of a polymer obtained by reactions otherwise than involving only carbon to carbon unsaturated bonds
• CPC Class Codes: B01J 20/3219 involving a particular spacer or linking group, e.g. for attaching an active group
• CPC Class Codes: B01J 20/3236 containing metal, other than zeolites, e.g. oxides, hydroxides, sulphides or salts
• CPC Class Codes: B01J 20/3248 the functional group or the linking, spacer or anchoring group as a whole comprising at least one type of heteroatom selected from a nitrogen, oxygen or sulfur, these atoms not being part of the carrier as such
• CPC Class Codes: B01J 20/3253 : comprising a cyclic structure not containing any of the heteroatoms nitrogen, oxygen or sulfur, e.g. aromatic structures
• CPC Class Codes: B01J 20/3265 : with an organic functional group containing a metal, e.g. a metal affinity ligand
• CPC Class Codes: B01J 20/3274 : Proteins, nucleic acids, polysaccharides, antibodies or antigens
• CPC Class Codes: B01J 20/3285 : Coating or impregnation layers comprising different type of functional groups or interactions, e.g. different ligands in various parts of the sorbent, mixed mode, dual zone, bimodal, multimodal, ionic or hydrophobic, cationic or anionic, hydrophilic or hydrophobic
• CPC Class Codes: B01J 20/3289 : Coatings involving more than one layer of same or different nature
• CPC Class Codes: B01J 2219/00605 : the compounds being directly bound or immobilised to solid supports
• CPC Class Codes: B01J 2219/0061 : The surface being organic
• CPC Class Codes: B01J 2219/00612 : the surface being inorganic
• CPC Class Codes: B01J 2219/00617 : by chemical means
• CPC Class Codes: B01J 2219/00619 : using hydrophilic or hydrophobic regions
• CPC Class Codes: B01J 2219/00621 : by physical means, e.g. trenches, raised areas
• CPC Class Codes: B01J 2219/00626 : Covalent
• CPC Class Codes: B01J 2219/00628 : Ionic
• CPC Class Codes: B01J 2219/0063 : Other, e.g. van der Waals forces, hydrogen bonding
• CPC Class Codes: B01J 2219/00637 : by coating it with another layer
• CPC Class Codes: B01J 2219/00641 : the porous medium being continuous, e.g. porous oxide substrates
• CPC Class Codes: B01J 2219/00644 : the porous medium being present in discrete locations, e.g. gel pads
• CPC Class Codes: B01J 2219/00659 : Two-dimensional arrays
• CPC Class Codes: B01J 2219/00702 : Processes involving means for analysing and characterising the products
• CPC Class Codes: B01J 2219/00725 : Peptides
• CPC Class Codes: B01J 2220/54 : Sorbents specially adapted for analytical or investigative chromatography
• CPC Class Codes: C12Q 1/00 : Measuring or testing processes involving enzymes, nucleic acids or microorganisms measuring or testing apparatus with condition measuring or sensing means, e.g. colony counters, C12M1/34; Compositions therefor; Processes of preparing such compositions
• CPC Class Codes: C40B 30/04 : by measuring the ability to specifically bind a target molecule, e.g. antibody-antigen binding, receptor-ligand binding
• CPC Class Codes: C40B 40/10 : Libraries containing peptides or polypeptides, or derivatives thereof
• CPC Class Codes: G01N 2030/008 : Thermal desorption
• CPC Class Codes: G01N 2030/009 : Extraction
• CPC Class Codes: G01N 30/00 : Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation G01N3/00, G01N5/00, G01N7/00, G01N9/00, G01N11/00, G01N13/00, G01N15/00, G01N17/00, G01N19/00, G01N21/00, G01N22/00, G01N23/00, G01N24/00, G01N25/00, G01N27/00, G01N29/00 take precedence
• CPC Class Codes: G01N 30/02 : Column chromatography
• CPC Class Codes: G01N 30/461 : with serial coupling of separation columns
• CPC Class Codes: G01N 30/463 : for multidimensional chromatography
• CPC Class Codes: G01N 30/466 : with separation columns in parallel
• CPC Class Codes: G01N 30/90 : Plate chromatography, e.g. thin layer or paper chromatography
• CPC Class Codes: G01N 30/95 : Detectors specially adapted therefor; Signal analysis
• CPC Class Codes: G01N 33/531 : Production of immunochemical test materials
• CPC Class Codes: G01N 33/538 : by sorbent column, particles or resin strip , i.e. sorbent materials
• CPC Class Codes: G01N 33/543 : with an insoluble carrier for immobilising immunochemicals
• CPC Class Codes: G01N 33/54306 : Solid-phase reaction mechanisms
• CPC Class Codes: G01N 33/54353 : with ligand attached to the carrier via a chemical coupling agent coatings G01N33/54393
• CPC Class Codes: G01N 33/54373 : involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings
• CPC Class Codes: G01N 33/54393 : Improving reaction conditions or stability, e.g. by coating or irradiation of surface, by reduction of non-specific binding, by promotion of specific binding
• CPC Class Codes: G01N 33/566 : using specific carrier or receptor proteins as ligand binding reagents where possible specific carrier or receptor proteins are classified with their target compounds
• CPC Class Codes: G01N 33/6803 : General methods of protein analysis not limited to specific proteins or families of proteins
• CPC Class Codes: G01N 33/6845 : Methods of identifying protein-protein interactions in protein mixtures
• CPC Class Codes: G01N 33/6851 : Methods of protein analysis involving laser desorption ionisation mass spectrometry
• CPC Class Codes: Y02A 90/10 : Information and communication technologies [ICT] supporting adaptation to climate change, e.g. for weather forecasting or climate simulation
• CPC Class Codes: Y10S 422/906 : Plasma or ion generation means
• CPC Class Codes: Y10S 435/803 : Physical recovery methods, e.g. chromatography, grinding
• CPC Class Codes: Y10S 435/81 : Packaged device or kit
• CPC Class Codes: Y10S 435/814 : Enzyme separation or purification
• CPC Class Codes: Y10S 435/815 : by sorption
• CPC Class Codes: Y10S 435/97 : Test strip or test slide
• CPC Class Codes: Y10S 435/973 : Simultaneous determination of more than one analyte
• CPC Class Codes: Y10S 435/975 : Kit
• CPC Class Codes: Y10S 436/808 : Automated or kit
• CPC Class Codes: Y10S 436/809 : Multifield plates or multicontainer arrays
• CPC Class Codes: Y10S 436/811 : Test for named disease, body condition or organ function
• CPC Class Codes: Y10S 436/813 : Cancer
• CPC Class Codes: Y10S 436/824 : Immunological separation techniques
• CPC Class Codes: Y10S 436/825 : Pretreatment for removal of interfering factors from sample
• CPC Class Codes: Y10T 436/24 : Nuclear magnetic resonance, electron spin resonance or other spin effects or mass spectrometry
• CPC Class Codes: Y10T 436/25 : including sample preparation
• CPC Class Codes: Y10T 436/25125 : Digestion or removing interfering materials
• CPC Class Codes: Y10T 436/2525 : Stabilizing or preserving
• CPC Class Codes: Y10T 436/25375 : Liberation or purification of sample or separation of material from a sample [e.g., filtering, centrifuging, etc.]
• CPC Class Codes: Y10T 436/255 : including use of a solid sorbent, semipermeable membrane, or liquid extraction

Abstract

This invention provides methods of retentate chromatography for resolving analytes in a sample. The methods involve adsorbing the analytes to a substrate under a plurality of different selectivity conditions, and detecting the analytes retained on the substrate by desorption spectrometry. The methods are useful in biology and medicine, including clinical diagnostics and drug discovery.

Claims

1. A method of detecting an enzyme in a sample, comprising:

a) providing a mass spectrometry probe comprising an adsorbent and an enzyme substrate bound to the adsorbent, wherein the activity of the enzyme on the enzyme substrate produces a product having a characteristic molecular mass;

b) exposing the substrate to the sample; and

c) detecting the product by desorption spectrometry, wherein detecting the product provides a detection of the enzyme, thereby detecting the enzyme in the sample.

2. The method of claim 1, further comprising determining the amount of product.

3. The method of claim 1, further comprising comparing to a control.

4. The method of claim 1, wherein the enzyme is selected from the group consisting of a kinase, a phosphatase, a protease, and a nuclease.

5. The method of claim 1, wherein the substrate is a peptide comprising a lysine-lysine or a lysine-arginine pair.

6. The method of claim 1, wherein the adsorbent is a bifunctional linker.

7. The method of claim 6, wherein the bifunctional linker is selected from the group consisting of triethoxysilane, aminoethyl disulfide, carbodiimide and N-hydroxysuccinimide.

8. The method of claim 6, wherein the bifunctional linker comprises a reactive functional group selected from the group consisting of silicon dioxide, a C₁₈ aliphatic hydrocarbon, a quaternary amine and sulphonate.

9. The method of claim 1, wherein the enzyme substrate is non-covalently bound to the adsorbent.

10. The method of claim 1, wherein the enzyme substrate is covalently bound to the adsorbent.

11. The method of claim 1, wherein the enzyme substrate is bound to the adsorbent by a basis of attraction selected from the group consisting of a hydrophobic interaction, a thiophilic interaction, an immobilized dye interaction, a van der Waals force interaction, an anionic interaction, a cationic interaction, a coordinate complex interaction, a glycoprotein interaction and a biospecific interaction.