Method For Purification Of Nucleic Acids, Particularly From Fixed Tissue

• US 20110245483A1
• Filed: 11/20/2009
• Published: 10/06/2011
• Est. Priority Date: 12/12/2008
• Status: Active Grant

• Current Assignee: Siemens Healthcare Diagnostics Incorporated (Siemens AG)
• Original Assignee: Siemens Healthcare Diagnostics Incorporated (Siemens AG)
• Inventors: Izmailov, Alexandre, Euting, Heike, Hennig, Guido
• Granted Patent: US 8,703,931 B2
• Time in Patent Office: Days
• US Class Current: 536/25.41
• CPC Class Codes: C07B 63/00 Purification; Separation separation of optically-active compounds C07B57/00; Stabilisation; Use of additives
• CPC Class Codes: C12N 15/1013 by using magnetic beads
• CPC Class Codes: C12Q 1/6806 Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay C12Q1/6804 takes precedence
• CPC Class Codes: Y10T 137/8376 Combined

Abstract

The invention relates to a method for purification of nucleic acids, to a kit for performing the method according to the invention and to a new application of magnetic particles for purification of a biological sample. The method according to the invention comprises the following steps: a) accommodating of the sample in a first sample vessel in an aqueous solution and lysing of the sample under non-chaotropic conditions; suspending of first magnetic particles in the solution and inserting of the first sample vessel in a sample vessel holder, wherein the sample vessel is inserted in the annular interior space of a ring magnet associated with the sample vessel holder; separating of the solution from the magnetic particles; and isolating of the nucleic acids from the solution.

Claims

1. A method for purifying nucleic acids from a biological sample, comprising the following steps:

a) collecting the sample in a first sample vessel in an aqueous solution and lysing the sample under nonchaotropic conditions;

b) suspending first magnetic particles in the solution;

c) placing the first sample vessel into a sample vessel holder, wherein the sample vessel is placed into the annular interior space of a ring magnet associated with the sample vessel holder;

d) separating the solution from the magnetic particles; and

e) isolating the nucleic acids from the solution.

2. The method as claimed in claim 1, wherein the magnetic particles have a silicon-containing coating.

3. The method as claimed in claim 2, wherein the sample is a blood sample.

4. The method as claimed in claim 2, wherein the biological sample is a paraffin-embedded sample and/or a fixed sample.

5. The method as claimed in claim 4, wherein the solution is heated prior to step (d) to at least 50°

C., preferably at least 60°

C., more preferably 60-95°

C.

6. The method as claimed in claim 5, wherein the sample is cooled again to below 50°

C. after the heating prior to step W).

7. The method as claimed in claim 6, wherein the sample is contacted with a hydrophobic matrix in at least one of steps a), b), and c).

8. The method as claimed in claim 7, wherein the first sample vessel is made of a hydrophobic material and acts as a hydrophobic matrix.

9. The method as claimed in claim 8, wherein the solution in step (d) is separated from the magnetic particles by aspiration.

10. The method as claimed in claim 9, wherein step (e) comprises the addition of a chaotropic compound to the sample.

11. The method as claimed in claim 10, wherein step (e) further comprises suspending unused second magnetic particles having a silicon-containing coating to the solution.

12. The method as claimed in claim 11, wherein step (e) further comprises:

i) collecting the sample in a second sample vessel and adding a chaotropic compound;

ii) suspending second magnetic particles in the solution and binding the nucleic acids to the second magnetic particles;

iii) placing the second sample vessel into the sample vessel holder, wherein the second sample vessel is placed into the annular interior space of the ring magnet associated with the sample vessel holder; and

iv) separating the solution from the magnetic particles and eluting the nucleic acids from the magnetic particles.

13. The method as claimed in claim 12, wherein, when placing the first and/or second sample vessel into the annular interior space of the ring magnet associated with the sample vessel holder, the respective sample vessel is moved at least once out of the annular space and back in again by means of movement in the direction of the ring axis.

14. An automated system comprising:

a rack for at least one sample vessel;

a temperature-controlled device having at least one slot for the sample vessel;

a sample vessel holder for accommodating at least one sample vessel;

a device for transporting the sample vessel holder from the rack to the temperature-controlled device;

a device for transferring liquid from a sample vessel and/or into a sample vessel;

a control for controlling the transport device, the liquid transfer device, and for controlling the temperature of the temperature-controlled device;

wherein the sample vessel holder for accommodating the at least one sample vessel comprises at least one annular magnet, in the annular interior space of which the sample vessel can be accommodated.

15. A sample vessel holder for accommodating at least one sample vessel, comprising at least one annular magnet, in the annular interior space of which at least one sample vessel can be accommodated.

16. The sample vessel holder as claimed in claim 15, wherein the ring inner diameter of the annular magnet is, at its narrowest point, 4 to 20 mm, preferably 6-12, in particular 8 mm.

17. The sample vessel holder as claimed in claim 15, wherein the annular interior space has a conical region.

18. The sample vessel holder as claimed in claim 15, comprising a multiplicity of slots for sample vessels.