# US 6,720,161 B1 · Immobilized substrate, separation gels and method for detecting enzymatic activity

> Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt

- Number: US 6,720,161 B1
- Document ID: US6720161B1
- Status: Granted
- Kind: B1
- Application number: 09/673,337
- Filed: 2000-12-26
- Granted: 2004-04-13
- Earliest claimed: 1998-04-22
- Assignees: Gesellschaft fuer Biotechnologische Forschung mbH
- Inventors: Ronald Frank; Werner Tegge
- Web page: https://patentgrep.com/patent/US6720161B1

## CPC

- G01N 33/561: Immunoelectrophoresis
- 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
- G01N 30/90: Plate chromatography, e.g. thin layer or paper chromatography
- G01N 2650/00: Assays involving polymers whose constituent monomers bore biological functional groups before polymerization, i.e. vinyl, acryl derivatives of amino acids, sugars
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES

## Abstract

The present invention refers to a method for attaching substrate molecules to gel matrices, characterized in that a substrate is covalently bound to a monomeric or oligomeric constitutional unit of a gel and the resultant product is polymerized. The present invention further refers to gels obtainable by this process.

## Claims

### Claim 1

A method for separating and detecting protein kinase activities by conducting the following steps in the given order:
(a) covalently binding a substrate to a monomeric or oligomeric constitutional unit of a gel;
(b) polymerizing the resultant product to form a gel matrix of a separating gel;
(c) electrophoretically separating the protein kinase activities to be separated using the separating gel, and optionally renaturating; and
(d) detecting, either by labelling with a detectable label or destroying a label present, by reacting the substrate which is covalently bound in the gel with the separate protein kinase activity/activities.

### Claim 2 (depends on 1)

The method according to claim 1, wherein the substrate comprises a natural or synthetic peptide, a natural or synthetic oligonucleotide or a small organic molecule.

### Claim 3 (depends on 1)

The method according to claim 1, wherein before polymerization the product prepared from the constitutional unit of substrate and gel is mixed with constitutional unit comprises acrylic monomers, acrylic acid oligomers, methacrylic acid monomers and/or methacrylic acid oligomers.

### Claim 4 (depends on 1)

The method according to claim 1, wherein the constitutional units of the gel of gel which do not contain substrate molecules.

### Claim 5 (depends on 1)

The method according to claim 1, wherein the polymerization method is solution or bulk polymerization.

### Claim 6 (depends on 1)

The method according to claim 1, wherein a spacer is incorporated between the substrate and the monomeric or oligomeric constitutional unit of the gel.

### Claim 7 (depends on 6)

The method according to claim 6, wherein the spacer comprises a natural or synthetic peptide, a natural or synthetic oligonucleotide or a small organic molecule.

### Claim 8

A method of separating and detecting protein kinase activities by conducting the following steps in the given order:
(a) using a separating gel which is obtained by covalently binding a substrate to a monomeric or oligomeric constitutional unit of a gel and polymerizing the resultant product to form a gel matrix of a separating gel;
(b) electrophoretically separating the protein kinase activities to be separated using the separating gel, optionally renaturating; and
(c) detecting, either by labelling with a detectable label or destroying a label present, by reacting the substrate which is covalently bound in the gel with the separate protein kinase activity/activities.

### Claim 9 (depends on 8)

The method according to claim 8, wherein the substrate comprises a natural or synthetic peptide, a natural or synthetic oligonucleotide or a small organic molecule.

### Claim 10 (depends on 8)

The method according to claim 8, wherein before polymerization the product prepared from the constitutional unit of substrate and gel is mixed with constitutional unit comprises acrylic monomers, acrylic acid oligomers, methacrylic acid monomers and/or methacrylic acid oligomers.

### Claim 11 (depends on 8)

The method according to claim 8, wherein before polymerization the product prepared from the constitutional unit of substrate and gel is mixed with constitutional units of gel which do not contain substrate molecules.

### Claim 12 (depends on 8)

The method according to claim 8, wherein the polymerization method is solution or bulk polymerization.

### Claim 13 (depends on 8)

The method according to claim 8, wherein a spacer is incorporated between the substrate and the monomeric or oligomeric constitutional unit of the gel.

### Claim 14 (depends on 13)

The method according to claim 13 wherein the spacer comprises a natural or synthetic peptide, a natural or synthetic oligonucleotide or a small organic molecule.