Device and method of collection for RNA viruses
View Patent ↗The present disclosure generally relates to a method and device for inactivation and dry storage, under ambient conditions, of a biological sample containing RNA virus. Methods for collecting and recovering RNA from a biological sample and subsequent analysis for a virus are also provided.
1. A method of inactivation and dry storage of a biological sample containing an RNA virus, the method comprising:
providing a non-dissolvable, dry, solid matrix incorporated with a composition, the composition comprising at least one protein denaturant, at least one acid or acid-titrated buffer reagent impregnated therein in a dry state to provide an acidic pH on hydration;
contacting the biological sample with the non-dissolvable, dry, solid matrix to effectively lyse cells of the biological sample to extract RNA from the lysed cells and preserve the extracted RNA in an intact state;
drying the biological sample on the solid matrix; and
storing the biological sample on the solid matrix in a dry state under ambient conditions,
wherein the biological sample is rendered non-infectious within 30 minutes of being subjected to the drying step, and wherein the acidic pH is in a range from 2 to 7.
2. The method of claim 1 , wherein the acid comprises acetic acid, citric acid, tartaric acid, phosphoric acid, hydrochloric acid, Tris(2-carboxyethyl) phosphine-hydrochloric acid (TCEP-HCl), oxidized Tris(2-carboxyethyl) phosphine-hydrochloric acid (TCEP-O-HCl), sulfuric acid, nitric acid, vanillic acid, 3-(N-morpholino)propanesulfonic acid or combinations thereof.
3. The method of claim 1 , wherein the composition further comprises a UV protectant, a free-radical scavenger, a chelator, a reducing agent or combinations thereof.
4. The method of claim 1 , wherein the composition further comprises an RNase inhibitor.
5. The method of claim 4 , wherein the RNase inhibitor comprises at least one of a triphosphate salt, pyrophosphate salt, vanadyl ribonucleoside complex (VCR), or sodium pyrophosphate.
6. The method of claim 3 , wherein the reducing agent is selected from the group consisting of dithiothreitol (DTT), 2-mercaptoethanol (2-ME), tris(2-carboxyethyl)phosphine (TCEP), tris(2-carboxyethyl)phosphine hydrochloride (TCEP-HCl), and combinations thereof.
7. The method of claim 1 , wherein the extracted and preserved RNA has an RNA integrity number (RIN) of greater than 5.
8. The method of claim 1 , wherein the RNA virus is a blood-borne virus.
9. The method of claim 8 , wherein the blood-borne virus is Ebolavirus, Hepatitis virus, Arenavirus, Filovirus, Lentivirus, or a related subgroup.
10. The method of claim 1 , wherein the method further comprises recovering the RNA from the non-dissolvable, dry, solid matrix.
11. The method of claim 10 , wherein recovering the RNA comprises extracting the RNA by rehydrating the non-dissolvable, dry, solid matrix, solid phase extraction, electroelution, or combinations thereof.
12. The method of claim 11 , wherein rehydrating comprises using an aqueous solution, a buffer solution, an organic solution, or combinations thereof.
13. A method for testing a biological sample for presence of an RNA virus, the method comprising:
providing a non-dissolvable, dry, solid matrix incorporated with a composition, the composition comprising at least one protein denaturant, at least one acid or acid-titrated buffer reagent impregnated therein in a dry state to provide an acidic pH on hydration;
applying the biological sample comprising RNA to the non-dissolvable, dry, solid matrix;
drying the biological sample on the non-dissolvable, dry, solid matrix to preserve the RNA in an intact state;
recovering the RNA of the biological sample from the non-dissolvable, dry, solid matrix; and
analyzing the recovered RNA for the presence of a viral RNA to determine the presence of the RNA virus,
wherein the biological sample is rendered non-infectious within 30 minutes of being subjected to the drying step, and wherein the acidic pH is in a range from 2 to 7.
14. The method of claim 13 , wherein the recovering step comprises extracting the RNA by rehydrating the non-dissolvable, dry, solid matrix, electroelution, solid phase extraction, adding a portion of the solid matrix directly into an amplification reaction, or combinations thereof.
15. The method of claim 14 , wherein rehydrating comprises using an aqueous solution, a buffer solution, an organic solution, or combinations thereof.
16. The method of claim 13 , wherein the acid comprises acetic acid, citric acid, tartaric acid, phosphoric acid, hydrochloric acid, Tris(2-carboxyethyl) phosphine-hydrochloric acid (TCEP-HCl), oxidized Tris(2-carboxyethyl) phosphine-hydrochloric acid (TCEP-O-HCl), sulfuric acid, nitric acid, vanillic acid, 3-(N-morpholino)propanesulfonic acid, or combinations thereof.
17. The method of claim 13 , wherein the composition further comprises a UV protectant, a free-radical scavenger, a chelator, a reducing agent or combinations thereof.
18. The method of claim 13 , wherein the composition further comprises an RNase inhibitor.
19. The method of claim 18 , wherein the RNase inhibitor comprises at least one of a triphosphate salt, pyrophosphate salt, vanadyl ribonucleoside complex (VCR), or sodium pyrophosphate.
20. The method of claim 17 , wherein the reducing agent is selected from the group consisting of dithiothreitol (DTT), 2-mercaptoethanol (2-ME), tris(2-carboxyethyl)phosphine (TCEP), tris(2-carboxyethyl)phosphine hydrochloride (TCEP-HCl) and combinations thereof.
21. The method of claim 13 , wherein the biological sample is blood.
22. The method of claim 13 , wherein the RNA virus is a blood-borne virus.
23. The method of claim 22 , wherein the blood-borne virus is Ebolavirus, Hepatitis virus, Arenavirus, Filovirus, Lentivirus, or a related subgroup.
24. The method of claim 13 , further comprising prolonged storing the RNA in an intact form under a dry format and ambient conditions, wherein any virus present in the biological sample is rendered inactive.