Reducing non-covalently bound polysaccharide on supports

• US 8,394,647 B2
• Filed: 02/17/2011
• Issued: 03/12/2013
• Est. Priority Date: 02/17/2011
• Status: Active Grant

• Current Assignee: Siemens Healthcare Diagnostics Incorporated (Siemens AG)
• Original Assignee: Siemens Healthcare Diagnostics Incorporated (Siemens AG)
• Inventors: Lele, Bhalchandra, Singh, Pratap, Chatterjee, Asoke
• Primary Examiner(s): Dunn, Colleen
• Assistant Examiner(s): Soliman, Haytham
• Application Number:
  US13/029,786
• Publication Number:
  US 20120214978A1
• Time in Patent Office: 754 Days
• Field of Search: 436/529, 436/508, 436/509, 436/534, 502/404
• US Class Current: 436/529
• CPC Class Codes: C08B 37/00 Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof cellulose D21; microbiological processes C12P
• CPC Class Codes: C08B 37/0021 Dextran, i.e. (alpha-1,4)-D-glucan; Derivatives thereof, e.g. Sephadex, i.e. crosslinked dextran
• CPC Class Codes: C08L 25/06 Polystyrene
• CPC Class Codes: C08L 5/06 Pectin; Derivatives thereof

Abstract

Methods and reagents are disclosed for reducing an amount of non-covalently bound polysaccharide on a support. The method comprises contacting a support comprising both covalently bound polysaccharide and non-covalently bound polysaccharide with an aqueous solution comprising an amount of a chaotropic agent effective to remove non-covalently bound polysaccharide from the support.

Claims

1. A method of reducing an amount of non-covalently bound polysaccharide on a support, the method comprising:

contacting a support comprising both covalently bound polysaccharide and non-covalently bound polysaccharide with an aqueous solution comprising an amount of a chaotropic agent effective to remove non-covalently bound polysaccharide from the support wherein the amount of chaotropic agent in the aqueous solution is about 0.1 M to about 1 M and

separating the support from the aqueous solution wherein the separating comprises contacting the aqueous solution with a membrane and wherein substantiality all of the non-covalently bound polysaccharide passes through the membrane.

2. The method according to claim 1, wherein the polysaccharide is a dextran derivative.

3. The method according to claim 1, wherein the support is particulate.

4. The method according to claim 1, wherein the support is a latex support.

5. The method according to claim 1, wherein the chaotropic agent is selected from the group consisting of trifluoroacetate salts, trichloroacetate salts, thiocyanate salts, urea salts, thiourea salts, guanidinium salts, perchlorate salts, nitrate salts, bromide salts, iodide salts, trifluoromethane sulfonate salts, hexafluorophosphate salts, and bis(trifluoromethanesulfonyl)imidourea salts.

6. The method according to claim 1, wherein the aqueous solution comprises two or more chaotropic agents.

7. A method of preparing a conjugate of a support and an assay molecule, said method comprising:

(a) treating a support under conditions for covalently binding a polysaccharide to the support such that the support comprises both covalently bound polysaccharide and non-covalently bound polysaccharide,

(b) contacting the support with an aqueous solution comprising an amount of a chaotropic agent effective to remove non-covalently bound polysaccharide from the support wherein the amount of chaotropic agent in the aqueous solution is about 0.1 M to about 1 M,

(c) separating the support from the aqueous solution wherein the separating comprises contacting the aqueous solution with a membrane and wherein substantially all of the non-covalently bound polysaccharide passes through the membrane, and

(d) contacting the support with an assay molecule under conditions for binding of the assay molecule to the covalently bound polysaccharide.

8. The method according to claim 7, wherein the polysaccharide is a dextran derivative.

9. The method according to claim 7, wherein the support is particulate.

10. The method according to claim 7, wherein the support is a latex support.

11. The method according to claim 7, wherein the chaotropic agent is selected from the group consisting of trifluoroacetate salts, trichloroacetate salts, thiocyanate salts, urea salts, thiourea salts, guanidinium salts, perchlorate salts, nitrate salts, bromide salts, iodide salts, trifluoromethane sulfonate salts, hexafluorophosphate salts, and bis(trifluoromethanesulfonyl)imidourea salts.

12. The method according to claim 7, wherein the aqueous solution comprises two or more chaotropic agents.

13. The method according to claim 7, wherein the separating comprises contacting the aqueous solution to a membrane wherein substantially all of the non-covalently bound polysaccharide passes through the membrane.

14. The method according to claim 7, wherein the assay molecule is selected from the group consisting of members of a specific binding pair and members of a signal-producing system.

15. A method of reducing an amount of non-covalently bound dextran or dextran derivative on a latex particle the method comprising:

(a) contacting a latex particle comprising both covalently bound dextran derivative and non-covalently bound dextran derivative with an aqueous solution comprising an amount of one or more chaotropic agents effective to remove non-covalently bound dextran derivative from the support, and

(b) separating the latex particle from the aqueous solution by contacting the aqueous solution with a membrane wherein substantially all of the non-covalently bound dextran derivative passes through the membrane.

16. The method according to claim 15, wherein the chaotropic agent is selected from the group consisting of trifluoroacetate salts, trichloroacetate salts, thiocyanate salts, urea salts, thiourea salts, guanidinium salts, perchlorate salts, nitrate salts, bromide salts, iodide salts, trifluoromethane sulfonate salts, hexafluorophosphate salts, and bis(trifluoromethanesulfonyl)imidourea salts.

17. The method according to claim 15, wherein the dextran derivative is aminodextran, dextran aldehyde, dextran sulfate or carboxydextran.