Method for producing microparticles loaded with proteins

• US 7,335,401 B2
• Filed: 07/12/2004
• Issued: 02/26/2008
• Est. Priority Date: 08/10/2001
• Status: Expired due to Fees

• Current Assignee: Roche Diagnostics Operations Incorporated (Roche Holding AG)
• Original Assignee: Roche Diagnostics Operations Incorporated (Roche Holding AG)
• Inventors: Klause, Ursula, Finke, Andreas, Jona, Wolfgang, Donie, Federic, Herrmann, Rupert, Von Der Eltz, Herbert, Sluka, Peter
• Primary Examiner(s): Le; Long V.
• Assistant Examiner(s): FOSTER, CHRISTINE E
• Application Number:
  US10/774,325
• Publication Number:
  US 20050079545A1
• Time in Patent Office: 1,324 Days
• Field of Search: 427/414, 427/214, 427/220, 427/222, 427/338, 428/407, 428/403, 424/78.08, 424/460, 424/491, 436533-534, 436/86, 435/7.1, 435/7.5, 435/7.8, 435/177, 435180-181, 435/814, 106/38.4, 530/350, 530/402, 530/421, 530/810, 530/812, 530815-816
• US Class Current: 427/414
• CPC Class Codes: A61K 9/1676 having a drug-free core with discrete complete coating layer containing drug adsorbates of liquid drug formulations on inert powders without simultaneous granulation step A61K9/141; with further drug-free outer coating A61K9/5078; drug conjugated to non-active particles A61K47/6921
• CPC Class Codes: G01N 33/54326 Magnetic particles
• CPC Class Codes: G01N 33/54353 with ligand attached to the carrier via a chemical coupling agent coatings G01N33/54393
• CPC Class Codes: G01N 33/545 Synthetic resin

Abstract

The present invention concerns a method for producing microparticles loaded with proteins which is characterized in that the microparticles are loaded in suspension under strongly alkaline conditions. The invention also concerns microparticles which can be produced using this method and their use in a binding test e.g. in an immunoassay.

Claims

1. A method for producing protein-coated polystyrene microparticles consisting of the steps of:

(a) combining a suspension of uncoated polystyrene microparticles with a protein to form a combination, the protein being a partner of a bioaffinity binding pair and having a size from 10 nm to 300 nm as determined by photon correlation spectroscopy,

(b) coating the protein onto the microparticles by adsorption, wherein said coating step is conducted for a period of 1 to 10 days at a pH selected from a range of 10.5 to 12.5, and

(c) separating the non-adsorbed protein from the protein-coated microparticles.

2. The method of claim 1, wherein the protein is a polymerized protein.

3. The method of claim 1, wherein the protein is a streptavidin which has been polymerized by chemical treatment.

4. The method of claim 1, wherein the microparticles have a magnetizable core.

5. The method of claim 4 wherein the microparticles have a size of 2.8 um and consist of 88% polystyrene and 12% magnetite.

6. The method of claim 1 wherein said coating step is conducted for a period of 4 to 7 days.

7. The method of claim 1 wherein the coating step is conducted at a pH between 11 and 12.

8. A method of producing protein-coated polystyrene microparticles, said method consisting of the steps of:

(a) combining a suspension of polystyrene microparticles with a protein to form a combination, the protein being a partner of a bioaffinity binding pair and having a size from 10 nm to 300 nm as determined by photon correlation spectroscopy,

(b) coating the protein onto the polystyrene microparticles by adsorption, wherein said coating step is conducted using a buffer having a salt content of 0.3 to 1.5 M and a pH selected from a range between 10.5 and 12.5, for a period of 1 to 10 days, and

(c) separating the non-adsorbed protein from the protein-coated microparticles.