METHODS AND ASSEMBLIES FOR SAMPLE ANALYSIS

Abstract

A method of preparing for analysis of a sample is shown and described. In one embodiment, the method includes soaking at least one sample collector in an apyrase solution, and removing the apyrase solution. The result is preparing the sample collector by removing adenosine triphosphate from the sample collector.

Claims

1. A method of preparing for analysis of a sample, said method comprising: a. soaking at least one sample collector in an apyrase solution, thereby defining an apyrase-absorbed sample collector; b. mechanically substantially removing said apyrase solution about said apyrase-absorbed sample collector; c. submerging said sample collector in an swab soak solution, thereby defining a swab-soaked sample collector; and d. mechanically substantially removing said swab soak solution about said swab-soaked sample collector.

2. The method of claim 1, whereby preparing said sample collector for analysis of said sample comprises removing adenosine triphosphate from said sample collector.

3. The method of claim 1, wherein mechanically substantially removing said apyrase solution includes spinning said apyrase-absorbed sample collector.

4. The method of claim 1, including substantially destroying said apyrase solution about said sample collector.

5. The method of claim 4, wherein substantially destroying said apyrase solution includes providing a sample collector substantially free of a residual apyrase enzyme.

6. The method of claim 1, wherein substantially removing said apyrase solution includes adjusting a pH for inactivating said apyrase enzyme.

7. The method of claim 1, wherein substantially removing said apyrase solution includes heating said sample collector for a predetermined temperature and time profile.

8. The method of claim 7, wherein substantially removing said apyrase solution includes autoclaving said sample collector.

9. The method of claim 1, wherein substantially removing said swab soak includes spinning said swab-soaked sample collector.

10. The method of claim 1, including soaking a plurality of sample collectors having a sample collector tip in said apyrase solution, and submerging a plurality of sample collectors having a sample collector tip in a swab soak solution.

11. The method of claim 1, including: a. extracting a liquid from an organic matter containing apyrase and mixing said liquid with a first solution; b. removing a first supernatant; c. mixing a second solution into said first supernatant and extracting a precipitate containing apyrase; and d. performing dialysis on said precipitate containing apyrase.

12. The method of claim 11, including mixing a third solution with said precipitate containing apyrase thereby creating an apyrase solution.

13. The method of claim 11, wherein said first solution comprises an organosulfur thiourea and said second solution comprises ammonium sulfate.

14. The method of claim 11, including introducing a buffer fourth solution thereby releasing a pellet formation from a surface.

15. A sample collector provided by the method of claim 1.

16. A method of preparing for analysis of a sample, said method comprising: a. exposing at least one sample collector to an apyrase solution, thereby defining an apyrase-absorbed sample collector; and b. withdrawing said apyrase solution about said apyrase-absorbed sample collector.

17. The method of claim 16, including substantially destroying said apyrase solution about said sample collector.

18. The method of claim 17, wherein substantially destroying said apyrase solution includes providing a sample collector substantially free of a residual apyrase enzyme.

19. The method of claim 16, including exposing said sample collector to a swab soak solution, and withdrawing said swab soak solution about said swab-soaked sample collector.

20. A sample collector provided by the method of claim 16.

Description

EXAMPLE

[0044] First, a 10 mM sodium acetate solution was prepared. 48 liters of reverse osmosis de-ionized water was placed in a carboy. 68.05±0.01 grams of sodium acetate trihydrate was added to a flask containing 1000 mL of reverse osmosis de-ionized water and stirred until completely dissolved. The sodium acetate solution was transferred to the carboy and mixed for 5 minutes. 1M HCl was introduced to adjust the pH of the solution to 6.50±0.05. Then, the total volume of the solution was raised to 50 liters and continue to mix for 15 minutes.

[0045] Second, 30 pounds of Russet Potatoes was stored in 4 degrees Celsius and protected from light.

[0046] Third, 60 mL of 0.7M 1-phenyl-2-thiourea (PTU) was added to the acetone in the flask, and mixed on a stir plate for 15 minutes.

[0047] Fourth, the PTU solution was transferred to a separate carboy from the sodium acetone carboy.

[0048] Fifth, the potatoes were sliced and juice collected into a beaker.

[0049] Sixth, the potato juice was introduced to the PTU flask and mixed, then allowed to settle. The sediment that formed from mixing was waste and was disposed of after juicing.

[0050] Seventh, the supernatant was introduced into a graduated cylinder before placing supernatant into bottles for centrifuge application. The supernatant was spun in 4 degrees Celsius and at 7000 rotations per minute (RPM) for 30 minutes. After cycle, supernatants were poured into a bucket and mixed while discarding the pellets.

[0051] Eighth, ammonium sulfate concentration was introduced to the supernatant to produce a 45% (25.8 g/100 mL) solution. The solution was mixed at 4 degrees Celsius for 1.5 hours before spinning the solution with a centrifuge at 10,000 RPM for 30 minutes.

[0052] The supernatant was pooled into a graduated cylinder while discarding the pellets.

[0053] Ninth, ammonium sulfate concentration to 90% (30.2 g/100 ml) was introduced with the supernatant and mixed at 4 degrees Celsius overnight. Then, the juice was introduced into the centrifuge at 4 degrees Celsius at 10,000 RPM for 30 minutes. Tenth, the supernatant was discarded (the apyrase now being in the pellet). The pellets were reconstituted with sodium acetate buffer until all pellets were reconstituted and the pellet was completely in solution. 5-10 milliliters of sodium acetate buffer was used to rinse each bottle. This rinse was introduced to the first bottle of reconstituted pellets. The bottle was inverted to mix. The solution produced a milk-beige color.

[0054] Eleventh, reconstituted enzyme pellet were dialyzed in approximately 4.0 L-8.0 L of 10 mM Sodium Acetate Buffer at 4 degrees Celsius, leaving headspace in tubing to allow enzyme to swell. The dialysis buffer was replaced after 1 hour and then after every subsequent two hours. Then dialyzed overnight at 4 degrees Celsius in the final buffer change. Upon completion, dialyzed apyrase was pooled and the enzyme centrifuged at 10,000 rpm for 30 minutes and in 4 degrees Celsius.

[0055] Twelfth, the supernatant was transferred to a storage container. The apyrase, which was in the supernatant, was frozen at −20 degrees Celsius to act as unpurified apyrase.

[0056] Thirteenth, a predetermined concentration of apyrase was calculated by the formula:

2000 ml*6.0U/ml=Total amount of units needed:12000units

12000 units/(concentration of Apyrase in U/ml)=(mlneeded of Apyrase)

[0057] Fourteenth, using the autoclaved graduated cylinder, 2000 ml of AG-SS was introduced into a screw top bottle with a cap. Then, the apyrase was introduced and a volume removed in excess of the 2000 ml mark from the bottle, so the final volume remained 2000 mL.

[0058] Fifteenth, with a vacuum pump and two 1000 ml 0.2 um CN membrane filter units, a 3 ml sample was poured into the beaker from each bottle.

[0059] Sixteenth, two aerobic count peel plates and two yeast-mold peel plates containing sample were monitored for colony development.

[0060] Seventeenth, for the soak process, apyrase solution was poured into a beaker and sample collectors were submerged. The AG swab soak solution was poured into a beaker deeper than the apyrase solution.

[0061] Eighteenth, sample collectors were introduced into the apyrase beaker soaked for 10 minutes. Then the sample collectors were mechanically spun in Honey Spinner cups for 10 minutes before removing sample collectors and place in the beaker with the Swab Soak Solution for 10 minutes. Then the sample collectors were mechanically spun in Honey Spinner cups for 10 minutes.

Example

[0062] The following example and related table show efficacy levels for treated swab collectors shown and described herein. The testing involved a 300 uL spiked positive assay, shaken for five seconds and allowed to rest for one minute (to breakdown ATP, if present), with an AG-B positive spike at 1,000,000 RLU with ATP tablets for a 300 uL/assay.

[0063] For testing: in the control there was no apyrase treatment; test A included a 6 units/mL apyrase concentration and analyzed fresh for initial data.

TABLE-US-00001 TABLE 1 Initial baseline Control A Before 1052801 57086 Before 1013271 34056 Before 1027641 52715 After 965279 After 1516114 After 923504 Average: 1083102 47952 % change: −95.6%

[0064] As illustrated in Table 1 above, the initial baseline data shows the control did not degrade during the analysis and the results of testing in Column A reflect the unforeseen advantage of absence of ATP.

[0065] Applicants' unexpectedly discovered the unanticipated advantages under the processes and methodology herein for providing sample collectors being substantially free of adenosine triphosphate, for example for subsequent analysis of a sample. For instance, alternative treatments, such as under cleanroom conditions, irradiation treatment, and gas treatment, failed to provide the efficiency and benefits of sample collectors being substantially free of adenosine triphosphate provided herein. Instead, the teachings herein remove residual adenosine triphosphate, and then in certain examples coupled with an autoclave process, produce a significantly less destructive time and temperature profile, for instance in which raw materials, including but not limited to swab tips and shafts, were not discolored or otherwise degraded. As a result, Applicants' unexpectedly discovered sample collectors being substantially free of adenosine triphosphate, a residual apyrase enzyme, or the like, thereby not negatively impacting any of the additional analysis and testing shown and described herein.

[0066] Numerous characteristics and advantages have been set forth in the foregoing description, together with details of structure and function. Many of the novel features are pointed out in the appended claims. The disclosure, however, is illustrative only, and changes may be made in detail, especially in matters of shape, size, and arrangement of parts, within the principle of the disclosure, to the full extent indicated by the broad general meaning of the terms in which the general claims are expressed. It is further noted that, as used in this application, the singular forms “a,” “an,” and “the” include plural referents unless expressly and unequivocally limited to one referent.