Method for reducing the presence of amplification inhibitors in a reaction receptacle
View Patent ↗An automated analyzer for performing multiple diagnostic assays simultaneously includes multiple stations in which discrete aspects of the assay are performed on fluid samples contained in sample vessels. The analyzer includes stations for automatically preparing a sample, incubating the sample, preforming an analyte isolation procedure, ascertaining the presence of a target analyte, and analyzing the amount of a target analyte. An automated receptacle transporting system moves the sample vessels from one station to the next. A method for performing an automated diagnostic assay includes an automated process for isolating and amplifying a target analyte, and, in one embodiment, a method for real-time monitoring of the amplification process.
1. A method for reducing the presence of a nucleic acid amplification inhibitor in a reaction vessel, said method comprising the steps of:
(a) providing a nucleic acid containing material to a reaction vessel having an inner surface;
(b) after step (a), coating that portion of the inner surface of the reaction vessel that is in contact with the nucleic acid containing material with a surface treating agent, the surface treating agent being an oil that reduces the amount of an amplification inhibitor that can adhere to the inner surface of the reaction vessel;
(c) after step (b), exposing a first volume of the reaction vessel comprising the nucleic acid containing material to reagents and conditions sufficient to isolate a target nucleic acid;
(d) removing at least a portion of the first volume from the reaction vessel;
(e) after step (d), providing to the reaction vessel a wash solution;
(f) exposing a second volume of the reaction vessel comprising the wash solution to reagents and conditions sufficient to isolate the target nucleic acid;
(g) removing at least a portion of the second volume from the reaction vessel; and
(h) after step (g), providing to the reaction vessel reagents sufficient to amplify a target sequence of the target nucleic acid, and exposing the reaction vessel to amplification conditions, wherein the surface treating agent will not inhibit amplification of the target sequence.
2. The method of claim 1 , wherein the surface treating agent is provided to the reaction vessel prior to step (a).
3. The method of claim 1 , wherein the surface treating agent is provided to the reaction vessel after step (a).
4. The method of claim 1 , wherein the surface treating agent is a silicone oil.
5. The method of claim 1 , wherein step (b) comprises agitating the reaction vessel.
6. The method of claim 1 , wherein the reagents sufficient to isolate the target nucleic acid comprise a solid support material for immobilizing the target nucleic acid.
7. The method of claim 6 , wherein the solid support material comprises a magnetically-responsive particle.
8. The method of claim 7 , wherein the first and second volumes are exposed to a magnetic field in steps (c) and (f).
9. The method of claim 6 , wherein the surface treating agent is provided to the reaction vessel before the solid support material.
10. The method of claim 6 , wherein the solid support material is provided to the reaction vessel before the surface treating agent.
11. The method of claim 1 , wherein the reaction vessel comprises a hydrophobic material.
12. The method of claim 11 , wherein the reaction vessel comprises polypropylene.
13. The method of claim 1 , wherein the amplification inhibitor is provided by the nucleic acid containing material.
14. The method of claim 13 , wherein the amplification inhibitor is hemoglobin.
15. The method of claim 1 , wherein the amplification inhibitor is provided by the wash solution.
16. The method of claim 15 , wherein the amplification inhibitor is an anionic detergent.
17. The method of claim 1 , wherein the reagents for amplifying the target sequence include a primer, nucleoside triphosphates, and a nucleic acid polymerase capable of extending the 3′ end of the primer hybridized to the target nucleic acid.
18. The method of claim 1 , wherein the steps of the method are automated.
19. The method of claim 18 , wherein the steps of the method are performed within a housing of an analyzer.
20. A method for reducing the presence of a nucleic acid amplification inhibitor in a reaction vessel, the method comprising:
(a) removing at least a portion of a first volume of a reaction vessel having an inner surface, the first volume comprising a nucleic acid containing material;
(b) after step (a), providing to the reaction vessel a wash solution and a surface treating agent, wherein the surface treating agent is not an inhibitor of a nucleic acid amplification reaction;
(c) coating that portion of the inner surface of the reaction vessel that is in contact with the wash solution with the surface treating agent, the surface treating agent being an oil that reduces the amount of an amplification inhibitor that can adhere to the inner surface of the reaction vessel;
(d) after step (c), exposing a second volume of the reaction vessel comprising the wash solution to reagents and conditions sufficient to isolate a target nucleic acid;
(e) removing at least a portion of the second volume from the reaction vessel; and
(f) after step (e), providing to the reaction vessel reagents sufficient to amplify a target sequence of the target nucleic acid, and exposing the reaction vessel to amplification conditions, wherein the surface treating agent will not inhibit amplification of the target sequence.
21. The method of claim 20 , wherein the wash solution and the surface treating agent are simultaneously provided to the reaction vessel.
22. The method of claim 20 , wherein the wash solution and the surface treating agent are separately provided to the reaction vessel.
23. The method of claim 20 , wherein the surface treating agent is a silicone oil.
24. The method of claim 20 , wherein step (c) comprises agitating the reaction vessel.
25. The method of claim 20 further comprising providing to the reaction vessel a solid support material for immobilizing the target nucleic acid prior to step (a).
26. The method of claim 25 , wherein the solid support material comprises a magnetically-responsive particle.
27. The method of claim 26 , wherein the second volume is exposed to a magnetic field during step (d).
28. The method of claim 20 , wherein the reaction vessel comprises a hydrophobic material.
29. The method of claim 28 , wherein the reaction vessel comprises polypropylene.
30. The method of claim 20 , wherein the amplification inhibitor is provided by the nucleic acid containing material.
31. The method of claim 30 , wherein the amplification inhibitor is hemoglobin.
32. The method of claim 20 , wherein the amplification inhibitor is provided by the wash solution.
33. The method of claim 32 , wherein the amplification inhibitor is an anionic detergent.
34. The method of claim 20 , wherein the reagents for amplifying the target sequence include a primer, nucleoside triphosphates, and a nucleic acid polymerase capable of extending the 3′ end of the primer hybridized to the target nucleic acid.
35. The method of claim 20 , wherein the steps of the method are automated.
36. The method of claim 35 , wherein the steps of the method are performed within a housing of an analyzer.