Biological Specimen Collection and Transport System

Patent metadata

Field	Value
Publication number	US-2016108463-A1
Application number	US-201514969339-A
Country	US
Kind code	A1
Filing date	Dec 15, 2015
Priority date	Oct 1, 2007
Publication date	Apr 21, 2016
Grant date	—

Abstract

Disclosed are compositions for isolating populations of nucleic acids from biological, forensic, and environmental samples. Also disclosed are methods for using these compositions as one-step formulations for killing pathogens, inactivating nucleases, and releasing polynucleotides from other cellular components within the sample, and stabilizing the nucleic acids prior to further processing or assay. The disclosed compositions safely facilitate rapid sample collection, and provide extended storage and transport of the samples at ambient or elevated temperature without contamination of the sample or degradation of the nucleic acids contained therein. This process particularly facilitates the collection of specimens from remote locations, and under conditions previously considered hostile for preserving the integrity of nucleic acids released from lysed biological samples without the need of refrigeration or freezing prior to molecular analysis.

First claim

1 . A stock solution which, when combined with a biological sample suspected of containing a pathogen forms a composition comprising components with concentrations as follows:
a chaotrope present in an amount from 0.5 M to 6M; a detergent present in an amount from 0.1% to 1% (wt./vol.); a reducing agent in an amount from 0.5 mM to 0.3 M; a chelator present in an amount from 0.5 mM to 50 mM; a surfactant present in an amount from 0.0001% to 0.3% (wt./vol.); a short-chain alkanol present in an amount from 1 to 25% (vol./vol.); a buffer present in an amount from 1 mM to 1 M; an acid or base present in an amount that provides the stock solution with a pH of from 5 to 7; and nuclease-free water, wherein the components of the composition denature proteins, inactivate nucleases, and kill pathogens all without degrading nucleic acids of the sample.

2 . The stock solution of claim 1 , wherein the chaotrope comprises guanidine thiocyanate, guanidine isocyanate, guanidine hydrochloride or a combination thereof.

3 . The stock solution of claim 1 , wherein the detergent comprises sodium dodecyl sulfate (SDS), lithium dodecyl sulfate (LDS), sodium taurodeoxycholate (NaTDC), sodium taurocholate (NaTC), sodium glycocholate (NaGC), sodium deoxycholate (NaDC), sodium cholate, sodium alkylbenzene sulfonate (NaABS), N-lauroyl sarcosine (NLS), salts of carboxylic acids, salts of sulfonic acids, salts of sulfuric acid, phosphoric and polyphosphoric acid esters, alkylphosphates, monoalkyl phosphate (MAP), salts of perfluorocarboxylic acids, anionic detergents or a combination thereof.

4 . The stock solution of claim 1 , wherein the reducing agent is (β-ME, DTT, DMSO or formamide present in an amount from 0.05 M to 0.3 M, or wherein the reducing agent is TCEP present in an amount from 0.5 mM to 30 mM.

5 . The stock solution of claim 1 , wherein the chelator comprises ethylene glycol tetra acetic acid, hydroxyethylethylenediaminetriacetic acid, diethylene triamine penta acetic acid, N,N-bis(carboxymethyl)glycine, ethylenediaminetetraacetic, citrate anhydrous, sodium citrate, calcium citrate, ammonium citrate, ammonium bicitrate, citric acid, diammonium citrate, ferric ammonium citrate, lithium citrate or a combination thereof.

6 . The stock solution of claim 1 , wherein the surfactant comprises cocoamidopropyl hydroxysultaine, alkylaminopropionic acids, imidazoline carboxylates, betaines, sulfobetaines, sultaines, alkylphenol ethoxylates, alcohol ethoxylates, polyoxyethylenated polyoxypropylene glycols, polyoxyethylenated mercaptans, long-chain carboxylic acid esters, alkonolamides, tertiary acetylenic glycols, polyoxyethylenated silicones, N-alkylpyrrolidones, alkylpolyglycosidases, silicone polymers such as Antifoam A®, or polysorbates such as Tween®, or a combination thereof.

7 . The stock solution of claim 1 , wherein the short-chain alkanol comprises methanol, ethanol, propanol, butanol, pentanol, hexanol or a combination thereof.

8 . The stock solution of claim 1 , wherein the buffer comprises tris(hydroxymethyl) aminomethane, citrate, 2-(N-morpholino)ethanesulfonic acid, N,N-Bis(2-hydroxyethyl)-2-aminoethanesulfonic acid, 1,3-bis(tris(hydroxymethyl)methyl amino)propane, 4-(2-hydroxyethyl)-1-piperazine ethanesulfonic acid, 3-(N-morpholino) propanesulfonic acid, bicarbonate, phosphate or a combination thereof.

9 . The stock solution of claim 1 , wherein the pH is 6.7±0.25.

10 . The stock solution of claim 1 , wherein the pathogen comprises a virus or a bacterium.

11 . The stock solution of claim 1 , wherein the pathogen is hepatitis virus, papillomavirus, HIV, biological agent of SARS, corona virus, rotavirus, Influenza virus, Ebola virus, methicillin-resistant Staphylococcus , or M. tuberculosis.

12 . (canceled)

13 . The stock solution of claim 1 , further comprising an internal positive control.

14 . The stock solution of claim 1 , further comprising an anti-microbial agent, an anti-viral, agent, an antifungal agent, or a combination thereof.

15 . The stock solution of claim 1 , further comprising betadine, bovine serum albumin, an osmolyte or a combination thereof.

16 . A method for detecting a nucleic acid sequence that is indicative of the presence of the pathogen in the sample comprising, at an ambient temperature, contacting the biological sample with an amount of the stock solution of claim 1 forming a composition which is effective to kill pathogens and inactivate nucleases in the sample without degrading or modifying nucleic acid of the sample, so that the nucleic acid sequence of the pathogen is detectable by PCR amplification.

17 . The method of claim 16 , wherein the pathogen comprises bacteria, virus, or a fungus; optionally, wherein the bacteria is tuberculosis or the virus is influenza.

18 . A diagnostic kit containing the stock solution of claim 1 .

19 . The diagnostic kit of claim 18 , which does not require refrigeration.

20 . The diagnostic kit of claim 18 , further comprising one or more nucleic acid extraction devices.

21 . The diagnostic kit of claim 18 , wherein the stock solution is contained in one vessel and further comprising a separate vessel containing one or more additional reagents, buffers, or compounds.

Assignees

• Longhorn Vaccines & Diagnostics Llc

Inventors

• Fischer Gerald W
• Daum Luke T

Classifications

• C12Q1/6851

  Quantitative amplification · CPC title

• C12Q1/6848

  characterised by the means for preventing contamination or increasing the specificity or sensitivity of an amplification reaction · CPC title

• C12Q2600/166

  Oligonucleotides used as internal standards, controls or normalisation probes · CPC title

• C12Q1/6806 Primary

  Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay (C12Q1/6804 takes precedence) · CPC title

• C12N15/1003

  Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor · CPC title