US 20070289605A1

(19) United States

(12) Patent Application Publication (10) Pub. No.: US 2007/0289605 A1
Salloom et al. (43) Pub. Date: Dec. 20, 2007
(54) USE OF DILUTE HYDROGEN PEROXIDE Related U.S. Application Data
TO REMOVE DNA CONTAMINATION
(60) Provisional application No. 60/724,302, filed on Oct.
(75) Inventors: Ashlee Salloom, Nottingham, MD 7, 2005.
(US); Matt Collis, Seven Valleys, PA
(US) Publication Classification

Correspondence Address: (51) Int. Cl.

DAVID W. HIGHET, VP & CHIEF A6IL 2/00 (2006.01)
INTELLECTUAL PROPERTY (52) U.S. Cl. ................................................... 134/6; 422/28
COUNSEL BECTON, DICKINSON AND (57) ABSTRACT
COMPANY
BIRCH, STEWART, KOLASCH & BIRCH, The present invention provides a method of reducing nucleic
LLP acid contamination on a Surface, which comprises the steps
1 BECTON DRIVE, MC 110 of contacting a Surface to be decontaminated with a solution
FRANKLIN LAKES, NJ 07417-1880 (US) of hydrogen peroxide and water, and Subsequently wiping
the solution from the surface. Preferably, the solution con
(73) Assignee: BECTON, DICKINSON AND COM sists essentially of hydrogen peroxide and water. The solu
PANY, Franklin Lakes, NJ tion may also be sprayed on a Surface, or applied first to a
paper towel or the like, and then applied to a surface. The
(21) Appl. No.: 111543,781 concentration of the hydrogen peroxide Solution is dilute.
Concentrations of about 0.5% to about 30% are preferred. A
(22) Filed: Oct. 6, 2006 concentration of about 3% is most highly preferred.
US 2007/0289605 A1 Dec. 20, 2007

USE OF DILUTE HYDROGEN PEROXDE TO available from VWR, West Chester, Pa. (Cat. No. VW4540
REMOVE DNA CONTAMINATION 2) may be used. However, the use of any commercial,
generally available, Solution of aqueous hydrogen peroxide
BACKGROUND is contemplated by the present invention. The hydrogen
peroxide solution may also contain additional additives
0001. This Nonprovisional application claims priority (stabilizers, etc.) that are commonly used in commercial
under 35 U.S.C. S 119(e) on U.S. Provisional Application hydrogen peroxide solutions.
No(s). 60/724,302 filed on Oct. 7, 2005, the entire contents 0009. A solution “consisting essentially of hydrogen
of which are hereby incorporated by reference.
peroxide and water may contain further components that are
0002 Decontamination of nucleic acids from surfaces unrelated to the invention, such as, for example, a stabilizer
involved in the Polymerase Chain Reaction (PCR) tech to prevent degradation of the hydrogen peroxide, but will not
nique, and other related DNA amplification techniques, is contain further oxidants such as bleach or Surfactants or
extremely important. PCR, and related techniques, may enzymes.
amplify extraneous nucleic acids that, for example, have
been carried over from a previous amplification. This can 0010) Surfaces
lead to false positive results and mistyping. In prior methods 0011. The surfaces and items that can be treated are any
of Surface decontamination, expensive, difficult to handle that are typically found in a laboratory environment. Pref
Solutions have generally been deployed as the decontami erably this includes any surface that would be present in the
nation agent. In most cases, post-decontamination steps practice of nucleic acid amplification. This would include
involving cleaning decontamination reagent residue are also metal, glass, plastic, and ceramic Surfaces. This would
required. There is a need in the industry for a decontami preferably include surfaces on laboratory benches, instru
nation process that is inexpensive, easy to use, and that ments, and equipment. This would also include Surfaces in
utilizes readily available, user-friendly reagents. pipettors (including automated pipettors) used in nucleic
acid amplification. For example, an instrument Such as the
DESCRIPTION OF THE INVENTION BD ProbeTecTM ET Pipettor manufactured by Becton, Dick
inson and Company, and the like, are in view. This would
0003. The present invention involves treating a surface also include Surfaces in arrays, microarrays, and microwells
with dilute aqueous hydrogen peroxide Solution to remove that are used in nucleic acid amplification.
nucleic acid contamination from the Surface area. Dilute
Solutions of hydrogen peroxide are inexpensive, easy to 0012 Contaminants
handle, and are extremely effective at removing nucleic acid 0013 The contaminants that can be cleaned by the
contamination from a surface. Preferred solutions of this present methods include any nucleic acid based contami
invention consist essentially of hydrogen peroxide and nant. This would preferably include residual contaminants
water. These solutions may be used without the need of that may be present on the Surfaces of laboratory equipment
additional Surface cleaning steps that are generally necessary related to DNA amplification experiments. This especially
with other decontamination solutions. The solutions of the includes any residual contamination that can interfere with
present invention do not leave a residue that can interfere a Subsequent enzymatic reaction. The present methods can
with future amplifications. also decontaminate surfaces and items that are contaminated
0004. Accordingly, the present invention provides a with radioactive contaminants.
method of reducing nucleic acid contamination on a Surface, 0014) Preferred Embodiments
which comprises the steps of contacting a Surface to be
decontaminated with a solution of hydrogen peroxide and 0015. In one preferred embodiment, the hydrogen perox
water, and Subsequently wiping the Solution from the Sur ide solution is contacted with the surface to be decontami
face. Preferably, the solution consists essentially of hydro nated, the Solution is allowed to dry, and is then wiped away
gen peroxide and water. The solution may also be sprayed on in a final step. In this embodiment further wiping or cleaning
a Surface (e.g., a vertical Surface), or applied first to a paper of the surface is not performed. Thus, the present invention
towel or the like, and then applied to a Surface, and then eliminates the need for further cleaning steps that are often
wiped off. required to remove residue left by the decontamination
solution itself.
0005. In a typical embodiment the hydrogen peroxide 0016. In another preferred embodiment, the hydrogen
solution is allowed to dry for at least about three minutes peroxide solution is thoroughly contacted with the Surface to
before wiping. However, the hydrogen peroxide solution be decontaminated and then the surface is rinsed with water
may be allowed to dry for periods of 30 minutes, or an hour, before wiping and drying.
O. O.

0006 The concentration of the hydrogen peroxide solu 0017. In yet another preferred embodiment, an item to be
tion is dilute. Concentrations of about 0.5% to about 30% are decontaminated is soaked in the hydrogen peroxide Solution
preferred. More preferred concentrations are in the approxi and then is drained, rinsed with water, and dried.
mately 2% to approximately 10% range. A concentration of 0018. In yet another preferred embodiment, a wipe or
about 3% is most highly preferred. towel is first soaked in the hydrogen peroxide Solution and
0007 Hydrogen Peroxide then the surface to be cleaned is wiped with the wipe or
towel. If needed or desirable for a particular application, the
0008. The hydrogen peroxide solutions of the present surface can then be further wiped with a dry towel or wipe,
invention may be readily, and inexpensively, obtained from or with a towel soaked with water, or the surface can be
commercial sources. For example, Hydrogen Peroxide 3% rinsed with water.
US 2007/0289605 A1 Dec. 20, 2007

EXAMPLES Additional materials:

General Materials and Methods 0027 VWRTM Hydrogen Peroxide 3%, Stabilized (Cat.
No. VW3540-2) GC Plasmid Stock 10x10 copies/ul con
0019. In a typical test procedure, a surface area contain centration Nuclease-free Water LS Pipette Tips (100
ing 48—1"x1" squares is contaminated using 10x10 copies/ uL-1000 uL).
mL GC plasmid. The plasmid stock is diluted to a 10x10° General Testing Procedure
copies/mL concentration and applied to the Surface. Two
Swabs from each square are taken to ensure the Surface is 0028. Use thin labeling tape to set-up a “grid” containing
contaminated. Dilute hydrogen peroxide is then applied to six 1"x 1" by eight 1"x1" squares creating a total of 48—1"x
the contaminated Surface area and used as a decontamination 1" squares. Dilute a 10x10 GC plasmid stock to a 10x10°
reagent. An additional two Swabs are taken from each square copies/mL concentration. Dispense 100 mL of the dilute GC
and then tested on the BD ProbeTecTM ET. In one test, 93/96 plasmid stock onto each square of the grid. Use the cleaning
Swabs tested negative in both assays demonstrating a reduc swab provided in the BD ProbeTecTM ET Chlamydia tra
tion rate of 97%. chomatis and Neisseria gonorrhoeae (CT/GC) Amplified
0020 All monoplex runs incorporated Amplification DNA Assay Endocervical Specimen Collection and Dry
Transport Kit to spread the 100 mL of GC plasmid evenly
Control (AC) microwells to ensure that the hydrogen per across each individual square. Allow the grid Surface to dry
oxide was not interfering with the Strand Displacement completely. (30 min-1 hr.) Using the endocervical swab
Amplification reaction. There were no AC indeterminates provided in the BD ProbeTecTM ET Chlamydia trachomatis
throughout the study. Therefore, it can be concluded that and Neisseria gonorrhoeae (CT/GC) Amplified DNA Assay
hydrogen peroxide does not cause inhibition in the assay and Endocervical Specimen Collection and Dry Transport Kit
should not be considered a risk factor to the product. take two Swab Samples from each square, totaling 96
0021 Definitions samples. Express each swab into a BD ProbeTecTM ET
CT/GC Swab Diluent Tube. Cap and vortex each tube for 5
0022 GC: Neisseria gonorrhoeae. AC: Amplification seconds. Prepare a negative and positive control tube by
Control. IAC: Internal Amplification Control. Monoplex: adding 2 mL of BD ProbeTecTM CT/GC Sample Diluent to
GC assay with an external amplification control (AC). each tube and Vortexing for 5 seconds. Heatlyse all samples
Diplex (Qx): GC assay with an internal amplification control and controls at 114°C. for 30 minutes. Then allow samples
(IAC). MOTA: Method Other Than Acceleration. PAT: to cool at room temperature for 15 minutes-6 hours.
Passes. After Threshold. HO: Hydrogen Peroxide. SD: Unscrew sample diluent caps and discard. Set up priming
Sample Diluent (potassium phosphate buffer) w/DMSO and amplification plates in accordance to Appendix I. Per
(10%) (CT/GC kit component). form the amplification reactions. Each Swab must test posi
tive in at least one of the two assays (GC monoplex/GC Qx)
0023 Materials to be considered positive.
0024. The BD ProbeTecTM ET System is a robotic, high 0029 Break seal and liberally apply dilute (3%) Hydro
throughput, real-time nucleic acid amplification system, gen Peroxide over entire grid surface and allow to stand for
manufactured by Becton, Dickinson and Company, Franklin three minutes. Wipe surface with Teri-Towel in a one
Lakes, N.J. A set of accessories for the system is available directional motion. Repeat General Testing Procedure
from the manufacturer for the detection of Chlamydia tra described above.
chomatis (CT) and Neisseria gonorrhoeae (GC) in clinical
specimens. The BD ViperTM Sample Processor is a robotic Example 1
system that automates the sample handling associated with
high Volume amplified molecular testing and is also avail Comparison of EliminaseTM vs. Hydrogen Peroxide
able from Becton Dickinson. The BD ViperTM Sample
Processor can be used with the BD ProbeTecTM ET system 0030) A total of 6 1'x2' blocks were measured and taped
for detection of Chlamydia trachomatis and Neisseria gon off on a laboratory counter top. Each cleaning method is
orrhoeae. BD ProbeTecTM ET GC Priming and Amplifica represented by 3 blocks as shown below:
tion Microwells, BD ProbeTecTM ET AC Priming and
Amplification Microwells, BD ProbeTecTM ET GCQx Prim
ing and Amplification Microwells, BD ProbeTecTM ET 1 3 5
CT/GC Positive Control, BD ProbeTecTM ET CTQx/GCQx 2 4 6
Positive Control, BD ProbeTecTM ET Negative Control, BD
ProbeTecTM ET Sample Diluent Tubes, BD ProbeTecTM ET Hydrogen Peroxide
Pipette Tips, BD ProbeTecTM ET Chlamydia trachomatis Eliminase TM
Eliminase TM
and Neisseria gonorrhoeae (CT/GC) are accessories that are Hydrogen Peroxide
useful with the BD ProbeTecTM and BD ViperTM systems. Hydrogen Peroxide
The above products are further described and can be ordered Eliminase TM
through the Becton Dickinson web site.
0.025 Amplified DNA Assay Endocervical Specimen 0.031) 48 Positive Controls were resuspended with 1 mL
Collection and Dry Transport Kit Equipment CT/GCSD, lysed for 30 min. at 114°C., then cooled for 15
min. 8 (8 mL) Positive Controls were “spilled on to each of
0026 BD ProbeTecTM ET Instrument System. BD Pro the 6 blocks and spread using a swab. The blocks were
beTecTM ET Matrix Pipettor. allowed to dry for approx. 30 minutes.
US 2007/0289605 A1 Dec. 20, 2007

0032. Before cleaning, 8 swab samples were taken from 114° C. for 30 minutes and cooled for 15 minutes. 150 uL
each block and expressed into a pre-filled SD tube. The sample was added to the priming wells to be tested, and then
blocks were then cleaned according to the designated clean incubated at room temperature for 20 min.
ing method for each block. After cleaning, an additional 8 0036) 150 uL of positive and negative CT/GC controls
Swab samples were taken from each block and expressed were added to control priming wells. The priming plates
into a pre-filled SD tube. 2 mL of SD was added to CT/GC were placed on a 72° C. heat block and amplification plates
Positive and Negative Controls. The tubes were then lysed were placed on a 54° C. heat block for 10 min. 100 uL of
in lysing block at 114°C. for 30 minutes and cooled for 15 priming mix was transferred to corresponding amplification
minutes. 150 u, sample was added to the priming wells to wells. The plates were sealed and run in ProbeTec(R) instru
be tested, and then incubated at room temp for 20 min. 150 ment for 60 minutes. Both GC monoplex and GC diplex
uL of positive and negative CT/GC controls were added to were tested in this study. Dilute hydrogen peroxide was
control priming wells. liberally poured onto the 48 contaminated squares, allowed
0033. The priming plates were placed on a 72° C. heat to stand for 3 minutes, then wiped away in a one-directional
block and amp plates were placed on a 54°C. heat block for motion. These steps were repeated and then 2 Swabs were
10 min. 100 uL of priming mix was transferred to corre taken from each square, processed and tested. 93/96 swabs
sponding amplification wells. The plates were sealed and run tested negative in both assays after cleaning the Surface with
in the ProbeTec instrument for 60 minutes. Both Eliminase hydrogen peroxide resulting in a reduction rate of 97%.
and hydrogen peroxide were effective decontamination TABLES 3-10 in the RESULTS Section show data obtained
reagents. Both reduced the number of contaminated Swabs from before cleaning. TABLES 11-18 show data from after
by 100%. The results are shown in TABLES 1 and 2 in the cleaning.
RESULTS Section.
Example 3
Example 2
Potency Stability of Hydrogen Peroxide
The Effectiveness of Dilute Hydrogen Peroxide as 0037. The potency stability of the hydrogen peroxide has
a Decontamination Reagent also been verified. The same method used as described
0034). A total of 48 1"x1" blocks were measured and above in Example 2 was used on Days 1, 3, 5, and 8. On day
taped off on the counter top. A GC plasmid stock with a one, the seal on the hydrogen peroxide bottle was broken
concentration of 10.11x10 was used to make the GC and the same bottle was used for the duration of the testing.
plasmid dilution. A 1:10 dilution was used to create a final The positivity reduction rates for each day were as follows:
GC plasmid concentration of 10.11x10°. To create enough Day 1–95%, Day 3–81%, Day 5–88%, and Day
volume to “contaminate' the 48 blocks, 500 uL of the GC 8—81%. Although, there appears to be a slight decline in the
plasmid stock and 4500 uL of deionized water were com potency of the hydrogen peroxide in reducing the contami
bined. nation, the results show that the effectiveness of the solution
0035. The blocks were allowed to dry for approx. 1 hour. remains very high over this length of time. The results are
shown in Tables 19-82 in the RESULTS Section.
Two swab samples were taken from each block and
expressed into a pre-filled SD tube. 2 mL of SD was added Results
to CT/GC Positive and Negative Controls and QX Positive
Control. The tubes were then lysed in the lysing block at 0038
TABLE 1.
Eliminase
2 2 3 3 6 6
Before After Before After Before After

Swabi Sample # CT GC CT GC CT GC CT GC CT GC CT GC

1 2O77O 20282 244 16S 36413 16583 313 S4O 31595 2521 531. 367
2 3172 450 95 27S 24613 14351 1OS 540 282SO 11942 711 555
3 31679 246 25 250 4895 S25 2SO 195 27549 123964. 261 307
4 14981 4378 13 95 19757 7859 14 100 8157 17550 204 717
5 3.0782 672 40 98 2822 6755 271 14O 30287 441 S3O 708
6 32988 4.184 106 14 15368 13670 357 2S2 35619 370 1S 412
7 3O822, 14154 416 253 33O8O 240 4SO 152 26O73 76S 467 493
8 1578O 1978 27O 387 32095 75O1 272 301 5550 491 438 SO7

Total # of Positives (MOTA over 2,000)

Before cleaning: After cleaning: Reduction percentage:
CT 24 CT O 100%
GC 14 GC O 100%
US 2007/0289605 A1 Dec. 20, 2007

0039)
TABLE 2
Hydrogen Peroxide
1 1 4 4 5 5
Before After Before After Before After

Swab/Sample # CT GC CT GC CT GC CT GC CT GC CT GC

1 29212 3585 124 333 17100 722 S64 734 10 8269 713 258
2 30976 2O2 32S 467 26314 414 268 797 2832 23837 411S 728
3 26248 22051 317 408 7228 20329 632 882 S982 2S144 376 244
4 4390 3531 370 286 SS16 14712 373 S23 14536 24993 651 296
5 241 221 S64 407 4908 13976 224. 466 25 19496 763 SO9
6 29537 593 289 455 73.54 490 1112 S93 911 12677 556 254
7 32499 SO18 111 491 8974 16SS6 SO2 342 2858 12808 312 S25
8 30944 592 436 S97 21606 4190 847 399 3961 580 77O 395

Total # of Positives (MOTA over 2,000)

Before cleaning: After cleaning: Reduction percentage:
CT 2O CT O 100%
GC 16 GC O 100%

Conclusions: Both Eliminase and hydrogen peroxide were effective decontamination reagents. Both reduced the num
ber of contaminated Swabs by 100%.

0040
TABLE 4-continued
TABLE 3
Diplex.
Monoplex
- VIOIODICX Swab # GC IAC
Swab # GC AC
4 O 47.7
1 31364 SO394 5 39.4 44.8
2 3S493 42726 6 31.6 45.3
3 33960 4.8992 7 35.1 44.2
4 32278 43363 8 25.6 45.3
5 33599 36893 9 25 45.3
6 2S119 38112 10 16 46.4
7 23628 43983 11 28.9 44.8
8 241.30 4386S 12 O 46.5
9 33329 393O4 13 31.8 44.3
10 27.083 46092 14 38.5 39.9
11 31278 44114 15 26.7 45
12 33784 34758 16 33.5 43
13 298.63 36999 17 30.1 45.2
14 31960 3O135 18 28.8 44.6
15 28277 2S647 19 27.8 45.4
16 21782 30744 2O 22.1 44.8
17 44O78 38812 21 23.9 45
18 41871 38268 22 20.7 44.2
19 3977O 2S318 23 26.9 44
2O 34484 33 183 24 29.3 42.7
21 39340 39S43
22 41163 31320
23 27470 37782
24 32O76 30784 0.042

TABLE 5
0041) Monoplex
TABLE 4 Swab # GC AC
Diplex 25 3.3856 33496
26 27569 37453
Swab # GC IAC 27 31136 34121
28 15777 39507
1 O 48.3 29 34.828 24.126
2 O 47.3 30 31661 281.96
3 O 47.4 31 24737 3O375
US 2007/0289605 A1 Dec. 20, 2007

TABLE 5-continued TABLE 7-continued

Monoplex Monoplex
Swab # GC AC Swab # GC AC

32 19459 26424 57 30474 36143

33 22536 34749 58 4526S 3.6869
34 32804 42O86 59 30729 29.538
35 27411 40577 60 36792 35887
36 42416 39354 61 35771 34762
37 35807 37442 62 36325 43327
38 297.62 34159 63 35065 31751
39 25496 3O281 64 33028 34615
40 20446 3.1083 65 35178 36599
41 3OO29 36878 66 3S010 37790
42 15555 27768 67 35754 47S46
43 26864 32447 68 33550 37351
44 21429 42172 69 35627 32.382
45 24216 33836 70 36684 42521
46 2S102 3S481 71 44,428 34987
47 24764 27891 72 31486 3S386
48 26986 29736

0045
0043)
TABLE 8
TABLE 6
Diplex
Diplex
Swab # GC IAC
Swab # GC IAC
49 32.4 40.7
25 38.1 40.1 SO 32.9 42.6
26 32.7 43 51 30 4.1.8
27 27.7 44.3 52 33.6 42
28 3O.S 43.4 53 28.2 43.5
29 33 43.3 S4 28.1 41
30 33 42 55 31.3 43.2
31 39 41.9 56 35.3 41.1
32 30 43.9 57 37.6 43.3
33 33.9 43.6 58 12.7 43.9
34 36.1 42.9 59 30.3 43.1
35 32.8 44.3 60 28.5 41
36 37.1 44.9 61 25.2 4O.S
37 25.2 43.9 62 32.1 39.5
38 36.9 42.2 63 32.6 36.7
39 34 41.5 64 29.5 36.6
40 8 43 65 27.2 44.1
41 29.8 44.6 66 30 44.5
42 39.3 39.6 67 33.7 4.1.8
43 18.6 43.1 68 34.8 42.4
44 19.4 43.7 69 29.9 42.4
45 18.1 42.4 70 16.7 43.3
46 26.7 42.9 71 6 42.9
47 31.6 42 72 38.2 41.7
48 25 42.4

0046)
0044)
TABLE 9
TABLE 7
Monoplex
Monoplex
Swab # GC AC
Swab # GC AC
73 34515 33506
49 12784 382O2 74 391.51 42.188
50 24748 434.08 75 35729 45.742
51 25867 31233 76 42110 33697
52 32685 36069 77 43.137 3S236
53 34427 4O106 78 38550 29939
S4 31736 311 69 79 34194 34515
55 36900 34232 8O 28716 30208
56 26713 3O842 81 3.1626 37949
US 2007/0289605 A1 Dec. 20, 2007
6

TABLE 9-continued TABLE 11-continued

Monoplex Monoplex
Swab # GC AC Swab # GC AC

82 2O845 32903 11 2O7 39055

83 30901 31340 12 245 36666
84 3O293 25968 13 31 SO1OO
85 26748 19044 14 120 33414
86 24083 14329 15 316 42091
87 224.52 162OS 16 220 30603
88 2O362 13921 17 391 44918
89 31049 32319 18 120 43951
90 25964 36240 19 431 45O14
91 28274 39771 2O 179 44951
92 28831 38660 21 4O6 43.860
93 26130 2.7489 22 563 37072
94 21889 29938 23 568 34083
95 15220 2931.8 24 16840 43604
96 1821.7 2S4O1

0049)
0047
TABLE 12
TABLE 10
Diplex
Diplex
Swab # GC IAC
Swab # GC IAC
1 O 34
73 32.7 44.2 2 O 40.3
74 26.7 43.5 3 O 39
75 31.6 41.6 4. O 40.4
76 28.9 42.8 5 O 40.4
77 33.8 40.9 6 O 42.1
78 34.1 38.9 7 O 39.4
79 31.3 4.O.S 8 O 39.5
8O O 41.4 9 O 42.1
81 31.7 44.9 10 O 40.6
82 29.7 44.4 11 O 40.8
83 33.3 42.5 12 O 39.9
84 36.6 42.9 13 O 40.2
85 31.5 43.1 14 O 42.1
86 29.8 43 15 O 38.2
87 31.8 42 16 O 37.6
88 33.2 38 17 O 36.4
89 38 43.1 18 O 40.1
90 28.8 42.8 19 O 39.5
91 35.4 43 2O O 40.8
92 24.9 43.2 21 O 40.1
93 36.4 41.5 22 O 36.7
94 33.8 41.6 23 O 40.7
95 33.9 40.3 24 O 37
96 26.5 39.9

0050
0048)
TABLE 13
TABLE 11
Monoplex
Monoplex
Swab # GC AC
Swab # GC AC
25 470 37526
1 283 31104 26 274 39090
2 102 3SO19 27 283 43810
3 281 26828 28 410 38.927
4 151 21997 29 585 38668
5 593 22532 30 107 38.429
6 536 21070 31 706 25.856
7 319 2O845 32 212 37936
8 12S6 1948S 33 3O2 44990
9 340 31008 34 463 42995
10 118 36O24 35 315 4985O
US 2007/0289605 A1 Dec. 20, 2007
7

TABLE 13-continued TABLE 15-continued

Monoplex Monoplex
Swab # GC AC Swab # GC AC

36 324 366O1 61 17 31348

37 218 39498 62 250 38853
38 552 34683 63 52 35122
39 488 35960 64 222 33951
40 531 3OSOO 65 551 31394
41 554 271.98 66 871 43460
42 638 31739 67 124 42415
43 224 36759 68 521 34260
44 455 34113 69 722 373.71
45 260 29577 70 277 35209
46 181 351.86 71 188 41689
47 964 3S460 72 374 41145
48 3066 38114

0053)
0051)
TABLE 16
TABLE 1.4
Diplex
Diplex
Swab # GC IAC
Swab # GC IAC
49 O 44.1
25 O 44.4 50 O 43.8
26 O 42.6 51 O 43.5
27 O 42.5 52 O 44.4
28 O 4.O.S 53 O 44
29 O 42.6 S4 O 42.2
30 O 42 55 O 44.5
31 O 44.3 56 O 45
32 O 41.6 57 O 423
33 O 41.3 58 O 42.8
34 O 43.1 59 O 43.6
35 O 42.6 60 O 43.1
36 O 4.1.8 61 O 40.8
37 O 41.1 62 O 41.3
38 O 39.7 63 O 4.1.8
39 O 35.2 64 O 39.2
40 O 37.9 65 O 4.1.8
41 O 40.4 66 O 423
42 O 42.7 67 O 44.1
43 O 38.3 68 O 43.2
44 O 41.7 69 O 43.3
45 O 42.5 70 O 43.7
46 O 37.9 71 O 41.9
47 O 39.9 72 O 38
48 O 40.6

0054)
0052)
TABLE 17
TABLE 1.5
Monoplex
Monoplex
Swab # GC AC
Swab # GC AC
73 478 41807
49 429 24785 74 744 4O794
50 351 32287 75 614 38845
51 252 33821 76 168 36003
52 77 38.974 77 373 32306
53 431 3S841 78 461 31253
S4 242 33858 79 329 35115
55 325 43736 8O 425 32632
56 187 35096 81 582 44919
57 453 394.94 82 641 44467
58 192 36295 83 552 42058
59 16605 4O458 84 42 42134
60 1 31242 85 40S 35646
US 2007/0289605 A1 Dec. 20, 2007
8

TABLE 17-continued TABLE 19-continued

Monoplex Monoplex
Swab # GC AC Swab # GC AC

86 3O8 39475 9 255.26 22095

87 361 33523 10 32223 23.860
88 312 234.08 11 33152 29639
89 288 49240 12 32868 26648
90 52 4.0311 13 31303 39657
91 68 37115 14 33661 28446
92 191 35237 15 35865 26365
93 83 38290 16 32361 34905
94 2 39829 17 23508 42571
95 1 39750 18 27570 45438
96 60 S1462 19 29S46 37838
2O 28973 43077
21 38.210 40570
22 271 77 2777B
0055) 23 3OO73 330S6
24 291.02 32163
TABLE 1.8

Diplex 0057)
Swab # GC IAC
73 O 40 TABLE 20
74 O 39.4 Dipl
75 O 445 DIDICX
is g .. Swab # GC IAC
78 O 41.9 1 36.1 40.4
79 O 43.7 2 38.5 39.6
8O O 35.4 3 39.4 38.7
81 O 39.9 4 38 39.1
82 O 38.7 5 37.7 36.4
83 O 40.4 6 36.2 39.4
84 O 42 7 39 37.3
85 O 423 8 38.8 41.5
86 O 42 9 37.3 39.8
87 O 38.4 10 36.1 37.3
88 O 39.6 11 37.3 34.4
89 O 39.8 12 37.3 35.6
90 O 37.2 13 34.8 38.9
91 O 41.9 14 35.5 38.9
92 O 42.6 15 38.7 33.4
93 O 40 16 32.6 41.9
94 O 40 17 35.9 38.1
95 O 42.9 18 34.7 40.2
96 O 39.7 19 38.4 32.5
2O 36.7 39.8
21 37.3 36.6
22 34.4 38.5
Example 3 23 35.7 39.2
24 39.6 32.7
Day 1— Before Cleaning
0056 All swabs are positive for GC in at least one assay 0058
(either monoplex or diplex)
TABLE 21
TABLE 19
Monoplex
Monoplex
Swab # GC AC
Swab # GC AC
25 35894 466O2
1 29422 23955 26 38841 SO4O6
2 34169 34733 27 39232 S1088
3 34500 34463 28 34687 47266
4 60421 32173 29 391.93 52651
5 36864 35684 30 3.1448 38943
6 37397 32498 31 46369 S162O
7 42297 34.432 32 42591 S4348
8 41069 36655 33 27952 37645
US 2007/0289605 A1 Dec. 20, 2007
9

TABLE 21-continued TABLE 23-continued

Monoplex Monoplex
Swab # GC AC Swab # GC AC

34 295.99 471SO 59 37248 35698

35 33911 49571 60 38765 S2546
36 29796 35096 61 37227 52762
37 31922 40339 62 361.75 48067
38 25925 31743 63 42462 47615
39 25725 26851 64 38529 47503
40 28S42 29505 65 49228 36441
41 32265 24964 66 34716 38208
42 33388 31862 67 42590 35867
43 34.465 41 O20 68 31124 35637
44 361.21 2881 6 69 391.30 35033
45 45894 30O27 70 34766 3O135
46 46652 34509 71 SO648 3O106
47 36485 28747 72 32734 27707
48 34370 41159

0061
0059)
TABLE 24
TABLE 22
Diplex
Diplex
Swab # GC IAC
Swab # GC IAC
49 38.6 36.8
25 38.6 36.6 50 37.9 38.9
26 33.4 35.4 51 36.7 38.5
27 37.9 40.2 52 31.6 35.5
28 37 35.4 53 37.5 37.8
29 35.7 34.6 S4 38.7 34.2
30 38.3 41.5 55 39.9 35.5
31 37.4 39.3 56 33.1 40.1
32 33.8 41.5 57 39.3 33.8
33 36.8 40.4 58 36.8 40.9
34 33.4 38.8 59 39.6 41.1
35 37.9 36 60 38.5 37.5
36 37 39.6 61 38.5 40.7
37 35.7 4.1.8 62 39.2 34.7
38 38.3 36.7 63 38.5 41.1
39 3.4 37.9 64 39.9 35
40 33.8 39.8 65 35.6 423
41 36.6 40.1 66 38.9 39
42 37.4 37.3 67 28.9 39.5
43 35.3 38.6 68 35.9 36.6
44 34.6 41.1 69 33.2 38.2
45 38.7 39.4 70 35 38.5
46 37 36.9 71 37.6 37.8
47 37.7 38.6 72 39.6 34.7
48 38 41.1

0062)
0060)
TABLE 25
TABLE 23
Monoplex
Monoplex
Swab # GC AC
Swab # GC AC
73 32979 41309
49 15975 26558 74 36966 53985
50 2O2O2 32.382 75 34823 411.93
51 21074 26215 76 37498 62391
52 24646 29210 77 37O68 50708
53 22,727 32027 78 33964 44997
S4 24899 27500 79 SO102 46267
55 27316 35641 8O 35765 39053
56 24139 24O61 81 26910 27768
57 3S224 27628 82 31258 32934
58 37746 41.375 83 29923 31891
US 2007/0289605 A1 Dec. 20, 2007
10

TABLE 25-continued TABLE 27

Monoplex Monoplex
Swab # GC AC
Swab # GC AC
1204 42294
84 31396 31515 490 40421
85 32027 37284 533 45944
86 33364 4O736 152 46735
87 34679 42714 498 45032
88 35762 4O487 246 35374
89 3827O 21807 225 353.81
582 37428
90 33906 32713
3O8 23294
91 4349S 34213 1 299 26372
92 46868 3O389 189 30289
93 3O472 2S301 1244 33642
94 46633 3O108 321 3.1469
95 53505 22971 495 29552
96 47014 18459 460 26O23
400 21632
220 S3294
416 38017
317 47082
0063 297 SS486
484 42940
TABLE 26 433 391.94
310 42997
Diplex 360 33393

Swab # GC IAC
73 39.8 40.4 0065
74 37.6 41.7
75 39.6 39.7 TABLE 28
76 37.6 38.4
77 38.8 39.1 LE 30
78 36.9 40.4
79 36.7 37.4 Swab # GC IAC
8O 38.1 38.3
81 38.3 35.5 36.6
82 37.5 36.3 38.7
83 38.6 39.2 40.2
84 37 34.6 39.8
85 38.7 34.2 36.8
86 35 37 39.9
87 38.4 39 37
88 38.1 40.7 36.5
89 37.8 38 39.7
90 38.6 37.3 1 41.3
91 36.4 39.3 39.8
92 34.9 40.2 41.1
93 37 40.7 42.1
94 37.6 37.8 37.2
95 38.8 38.4 37.5
96 35.8 39.9 36.6
36.7
39.2
37.1
35.5
Example 3 39.8
36.3
Day 1 — After Cleaning 23.4
36.8
Comments:

91/96 swabs tested in negative in both assays after cleaning 0.066)

the Surface with hydrogen peroxide resulting in a reduction
rate of 95%. TABLE 29
Monoplex
0064 Dilute Hydrogen Peroxide was liberally poured
onto the 48 squares, allowed to stand for 3 minutes, then Swab # GC AC
wiped away in a one-directional motion. These steps were 25 413 37998
repeated and then 2 Swabs were taken from each square, 26 205 43597
processed and tested.
US 2007/0289605 A1 Dec. 20, 2007
11

TABLE 29-continued TABLE 31-continued

Monoplex Monoplex
Swab # GC AC Swab # GC AC

27 477 S2248 51 1217 3O328

28 241 45243 52 311 34605
29 431 47610 53 196 33636
30 294 51932 S4 838 35845
31 421 43744 55 590 4O157
32 554 48093 56 385 394O1
33 497 47045 57 418 35035
34 237 58197 58 456 43324
35 435 41834 59 22531 31816
36 S16 47229 60 295 37551
37 663 40S10 61 493 39241
38 433 SS668 62 835 36399
39 469 45576 63 862 361.63
40 791 488.33 64 533 26O28
65 614 23176
41 156 24085 66 525 24176
42 324 22507 67 639 20922
43 384 2696S 68 440 2.9317
44 363 32557 69 716 2S602

46 522 29461 71 545 21254

47 494 30017 72 5O1 1984.4
48 149 25616

0069
0067
TABLE 32
TABLE 30
Diplex
Diplex
Swab # GC IAC
Swab # GC IAC
49 O 40.9
25 O 40.3 50 O 37.8
26 O 39.8 51 O 42.8
27 O 39.2 52 O 39.8
28 O 39.5 53 O 40.2
29 O 41 S4 O 39.8
30 O 37.4 55 O 39.9
31 O 37.5 56 O 39.7
32 30.1 31.9 57 O 37
33 O 4.O.S 58 O 38.9
34 O 39.5 59 O 35
35 O 36.6 60 O 36.5
36 O 37.5 61 O 39
37 O 38 62 O 35.8
38 6.4 38.6 63 O 34.4
39 O 39 64 O 36.3
40 O 38.3 65 O 38.3
41 O 42 66 O 40.7
42 O 39.4 67 O 40.9
43 O 41.6 68 O 41.1
44 O 41.3 69 O 423
45 O 39.1 70 O 39.7
46 O 39.1 71 O 39.9
47 O 39 72 O 36.9
48 O 38.2

0070)
0068
TABLE 33
TABLE 31
Monoplex
Monoplex
Swab # GC AC
Swab # GC AC
73 277 58236
49 331 26628 74 364 38829
50 6344 23273 75 8064 45206
US 2007/0289605 A1 Dec. 20, 2007
12

TABLE 33-continued TABLE 35

Monoplex Monoplex
Swab # GC AC
Swab # GC AC
28269 28675
76 887 48376 28713 30598
77 966 48913 321.59 333S4
78 330 S1316 25215 23263
79 346 45.505 27720 23861
8O 442 S1633 30649 24231
3O842 24298
81 3O2 19278
25695 241.14
82 259 19057 26399 22461
83 298 261 S6 1 2S366 27534
84 322 24.191 26871 36.559
85 539 24740 19861 31157
86 S88 26227 17878 29555
24319 28.074
87 734 241.15 2S910 3O255
88 386 160O2 2O197 26469
89 371 27.913 24281 262S3
90 558 331.97 3O381 291.70
91 559 33815 30972 298.32
29912 27532
92 778 30529
3O293 27264
93 579 27800 27895 26693
94 636 35917 284.04 24931
95 418 346O2 2S489 241.25
96 363 25.391

0073)
0.071) TABLE 36
TABLE 34 Diplex
Diplex Swab # GC IAC

Swab # GC IAC 37.5 43

29.6 43.6
73 O 36 33.8 43.2
74 O 39.8 38.5 41.7
75 O 37.9 32.9 39.7
76 O 40.6 32.9 42.9
77 O 4.O.S 23 42.7
78 O 40.3 37.2 42.6
79 O 42.6 32.3 38.8
8O O 42.4 1 25.3 4O.S
81 O 41.2 39.1 40.1
82 O 37.5 38 39.5
83 O 40.4 34.9 39.5
84 O 39.9 32 41
85 O 39.9 32.3 40.8
86 O 40 31.5 38.3
87 O 41.2 27.1 34.7
88 O 39.5 26.1 39.5
89 O 36.2 23 41.9
90 O 36.7 36.4 39.2
91 O 38.1 O 30.9
92 O 36.6 35.5 38.5
93 O 38.2 38.7 40
94 O 34.8 36.1 40.8
95 O 34.5
96 O 36.9
0074)
Example 3 TABLE 37
Monoplex
Day 3— Before Cleaning
Swab # GC AC
0072 All swabs are positive for GC in at least one assay 25 25359 32435
(either monoplex or diplex). Therefore according to AV-17 26 24834 34223
Verification, the study can proceed to step 4.4.2.
US 2007/0289605 A1 Dec. 20, 2007
13

TABLE 37-continued TABLE 39-continued

Monoplex Monoplex
Swab # GC AC Swab # GC AC

27 26929 35910 51 20287 2S482

28 2S380 37983 52 2O328 24307
29 26948 36347 53 20097 23162
30 274.67 31959 S4 21645 244OO
31 29334 39686 55 22045 21234
32 2S386 35578 56 23S18 1961.2
33 23573 1958O 57 27857 2S814
34 227O6 22479 58 294,08 27977
35 28262 26352 59 28918 35343
36 26692 241.12 60 27052 29925

38 24597 26.104 62 31364 31792

39 2S836 32045 63 30503 32883
40 25751 29917 3. t
41 23622 228SO
66 3SO43 421.93
42 26037 22178 67 3.1780 49960
43 26189 23894 68 26957 33457
44 24838 19523 69 30678 37343

46 2977O 23886 71 28.999 33832

47 3O3O8 2406S 72 24542 45846
48 2798O 24366

0.077
0075)
TABLE 40
TABLE 38
Diplex
Diplex
Swab # GC IAC
Swab # GC IAC
49 36.6 37.7
25 33 38.9 50 38.7 40.7
26 32.3 35.7 51 35.9 43.6
27 36 31.6 52 35.4 43.7
28 33 40.1 53 4.O.S 40.9
29 35.4 41.9 S4 38.5 39.5
30 30.9 39.9 55 36.7 43.9
31 33.2 39.4 56 14.1 41.5
32 37.1 39.3 57 32.2 42.4
33 28.3 42.1 58 35.5 37.9
34 28 40 59 34.6 36.4
35 32.8 36.1 60 29.6 39.1
36 32.8 42 61 37.4 39.4
37 31.4 42.4 62 38.4 34.7
38 28.3 37.6 63 23.3 40.4
39 33.1 40.3 64 37.5 40.3
40 38.1 38.5 65 29.8 39.9
41 31.1 41 66 22.9 37.9
42 18.1 37 67 36.3 36.4
43 37.1 4.O.S 68 34.7 39.1
44 29.9 40.4 69 34.1 39.4
45 31.5 40.8 70 36.7 34.7
46 9.3 39.9 71 28.2 40.4
47 35.2 41.4 72 32.7 43.6
48 36 42.5

0078
0076)
TABLE 41
TABLE 39
Monoplex
Monoplex
Swab # GC AC
Swab # GC AC
73 28.107 35932
49 2O781 26675 74 31822 30339
50 23907 31573 75 35960 32628
US 2007/0289605 A1 Dec. 20, 2007

These steps were repeated and then 2 swabs were taken from
TABLE 41-continued each square, processed and tested.
Monoplex 0081 Positive readings=Monoplex>2000: Diplex>0
Swab # GC AC
TABLE 43
76 28681 31094
77 28334 37460 Monoplex
78 32436 43941 o
79 29070 47051
8O 26.192 33008 Swab # GC AC
81 21642 39498
82 24427 4O681 1 5751 39114
83 17199 33291 2 25 495.99
84 21546 49143 3 299 37472
85 23140 60859
86 19817 51050 4 2O3 40416
87 23894 32179 5 17215 34549
88 2SO42 55674 6 293 38176
89 19735 35744 7 18O 47261
90 29270 242S1 8 3O8 34214
91 22414 28916
92 34374 33019 9 2007 37580
93 3.2129 36949 10 132 37678
94 291.36 40456 11 257 40897
95 34883 422O2 12 250 41077
96 38119 36930 13 589 44152
14 530 51009
15 470 43644
0079 16 526 4O733
TABLE 42 17 192 4OOOO
18 324 3962O
Diplex 19 561 46059
2O 319 393OO
Swab # GC IAC 21 327 55.292
73 33.5 42.1 22 341 2553
74 29.6 39.9 23 737 SO389
75 25.6 42.8 24 592 36519
76 36.7 38.7
77 36.7 36.9
78 31.8 38.7
79 9.5 38.9 0082
8O 34.7 42.7
81 28.1 43.8
82 35.8 423 TABLE 44
83 27.9 44.6
84 38.5 38.8 Diplex
85 29.1 42.8
86 32 43.6 Swab # GC IAC
87 18 43.3
88 35.9 41 1 O 43.5
89 38.3 42.8 2 O 43
90 38.2 40.3 3 O 43.9
91 38.6 43.2 4 O 42.6
92 25.6 43.6 5 O 39.4
93 34.8 42.2 6 O 423
94 36.5 43.4 7 O 43.1
95 37.9 42.1 8 O 43.1
96 34.9 41.5 9 O 39.6
10 O 44.2
11 O 42.4
12 O 41.9
Example 3 13 O 44.3
14 O 43.7
15 O 41.3
Day 3— After Cleaning 16 O 44.4
17 O 42
Comments: 18 O 41.4
78/96 swabs tested negative in both assays after cleaning the 5. g t
Surface with hydrogen peroxide resulting in a reduction rate 21 O 42.2
of 81%. 22 O 41.3
23 O 42.5
0080) Dilute Hydrogen Peroxide was liberally poured 24 O 44.1
onto the 48 contaminated squares, allowed to stand for 3
minutes, then wiped away in a one-directional motion.
US 2007/0289605 A1 Dec. 20, 2007
15

0083) 0085
TABLE 45 TABLE 47

Monoplex Monoplex

Swab # GC AC Swab # GC AC

25 294 45861 49 273 421.31

26 12726 36094 50 594 61642
27 S4O 40O88 51 1114 34843
28 357 378O1 52 1097 46640
29 551 43124 53 1042 42126
30 686 45622 S4 13659 46112
31 577 491.16 55 1034 38427
32 323 40229 56 527 40274
33 198 57227 57 183 48220
34 S16 46376 58 500 38.077
35 25 SO839 59 721 46726
36 157 371.83 60 993 4S463
37 751 40349 61 1276 37482
38 559 47310 62 451 46061
39 523 33604 63 430 42818
40 681 361.36 64 821 28208
41 262 33835 65 2525 52564
42 460 31611 66 932 5O197
43 666 32606 67 2498O 41297
244 32778 68 1324 43962
45 298 36770 69 S84 50562
46 287 2871.8 70 409 43O29
47 3279 3.0035 71 577 S1491
48 731 28.123 72 382 40703

0084) 0086)
TABLE 46 TABLE 48
Diplex Diplex
Swab # GC IAC Swab # GC IAC

25 41.6 49 37.6
26 41.5 50 38.9
27 40.3 51 41.1
28 43 52 4.1.8
29 42.6 53 42.6
30 42.5 S4 40.3
31 41.7 55 39.2
32 41.9 56 43.5
33 41.9 57 4O.S
34 42.6 58 40.4
35 42.4 59 34.3
36 42.2 60 38.7
37 41.3 61 35.8
38 43.1 62 37.3
39 42 63 39
40 39.6 64 35.4
41 39.4 65 39.7
42 41.2 66 39.3
43 41.5 67 38.8
42.2 68 39
45 4.1.8 69 38.5
46 40.4 70 33.2
47 42 71 36
48 40.8 72 35
US 2007/0289605 A1 Dec. 20, 2007
16

0087
TABLE 51
TABLE 49
Monoplex
Monoplex
Swab # GC AC
Swab # GC AC
23009 40S62
73 17860 41914 26577 40316
74 88 42132 29876 38775
75 3.162 45307 223OO 33.885
76 8392 47415 27645 38.243
77 935 37677 19933 35072
78 661 43790 19117 37660
79 774 44761 1626S 34.828
8O 24303 31399 26453 42292
81 21888 38229 1 281OS 39623
82 S4O 4.0997 3O862 47444
83 634 42981 31149 47756
84 695 33747 2.9971 48294
85 3217 37516 25929 424.54
86 742 37621 26655 39936
87 938 29094 25.358 52489
88 1339 24077 22344 47246
89 141OO 33408 30948 342S2
90 3.3396 45378 36.560 45075
91 33304 4O631 30724 37856
92 765 4O744 35273 42456
93 897 36348 32790 4O457
94 1591 4O470 34967 37327
95 13982 4O999 27614 45958
96 771 38647

0090)
0088) TABLE 52
TABLE 50 Diplex
Diplex Swab # GC IAC

Swab # GC IAC 29.7 31.2

31.2 37.3
73 O 38.8 35.5 36.8
74 O 36.6 37.7 38.9
75 O 39.8 33.7 39.1
76 O 35.6 21.1 41.4
77 O 37.2 37.3 42.8
78 O 37.1 32.6 42.2
79 O 35.5 34.4 39.5
8O O 38 1 14.5 43.1
81 O 38.2 36 40.4
82 O 37.8 32.2 40.1
83 O 41.4 20.8 39
84 O 42.7 21.9 40.2
85 12.4 41.3 26.8 40.6
86 O 42.9 33.3 37.9
87 O 39.9 22.3 33.2
88 O 43.3 29.2 39.9
89 O 37 28.6 36.9
90 O 39.8 33.1 36.7
91 O 40.6 32.2 37.8
92 O 39.6 34.8 35.8
93 O 41.4 31.4 38.5
94 O 39.5 13.6 40.6
95 O 40.3
96 O 39.2
0091)
Example 3 TABLE 53
Monoplex
Day 5 - Before Cleaning
Swab # GC AC
0089 All swabs are positive for GC in at least one assay 25 41108
(either monoplex or diplex). Therefore according to AV-17 26 S3646
Verification, the stud can proceed to step 4.4.2.
US 2007/0289605 A1 Dec. 20, 2007
17

TABLE 53-continued TABLE 55-continued

Monoplex Monoplex
Swab # GC AC Swab # GC AC

27 31426 46343 51 298.30 48.186

28 28727 34127 52 371.89 51529
29 28.614 3SOO6 53 37999 S612O
30 34448 38076 S4 263SO 45854
31 29779 3OOO7 55 33093 39654
32 28336 22292 56 292OS 33619
33 298.23 444O7 57 20193 25575
34 30570 495.99 58 28386 40288
35 32O82 40517 59 24537 37120
36 26663 57011 60 26.192 40023
37 3O479 S3485 61 222O3 34134
38 29013 4S452 62 28671 44.278
39 27707 43117 63 238S9 3443O
40 29377 40858 64 21967 23673
65 25512 41513
41 26571 46937
66 28647 49577
42 334O2 39512 67 31960 38860
43 311.63 41068 68 32386 45.532
44 30759 37567 69 26973 3708O

46 28432 40295 71 31.456 46548

47 33074 42755 72 24671 36217
48 244.08 26841

0094)
0092)
TABLE 56
TABLE 54
Diplex
Diplex
Swab # GC IAC
Swab # GC IAC
49 O 4O.S
25 14.4 43.3 50 14.3 41.1
26 20.7 38.8 51 4.2 41
27 19.6 42.2 52 O 37.9
28 24.8 40.2 53 2O 40.4
29 24 41.7 S4 23 42
30 23.3 42.4 55 34.1 41
31 3O4 41.1 56 23.2 39.8
32 30.7 41.6 57 O 34.7
33 29 29.5 58 25.8 29.6
34 12.8 37.9 59 O 4O.S
35 28.2 38.2 60 34.8 35.2
36 28.7 40 61 O 39
37 34.8 38.7 62 O 37.2
38 29.9 38.4 63 O 40.2
39 32.2 37.9 64 O 39
40 32.3 38.2 65 O 41
41 35.7 29.9 66 O 38.1
42 32.3 38.5 67 12 35.7
43 29.2 38.5 68 4.1 40.1
44 33.9 37 69 30.6 39.3
45 31.9 40 70 15.2 42.4
46 30.3 38.2 71 3.8 44.4
47 34.8 38.3 72 5.9 41
48 34.9 33.2

0095
0093)
TABLE 57
TABLE 55
Monoplex
Monoplex
Swab # GC AC
Swab # GC AC
73 3O106 15578
49 2SO94 33676 74 31548 4O262
50 31386 44991 75 29591 48.258
US 2007/0289605 A1 Dec. 20, 2007

These steps were repeated and then 2 swabs were taken from
TABLE 57-continued each square, processed and tested.
Monoplex 0099) Positive readings=Monoplex>2000: Diplex>0
Swab # GC AC
TABLE 59
76 41748 38891
77 39939 25659 Monoplex
78 32962 56889 o
79 29.348 56762
8O 301.45 15697 Swab # GC AC
81 2S4OS 51494
82 28531 54127 1 96.19 33237
83 30792 49439 2 472 52156
84 29.030 S8478 3 462 41883
85 31400 SOSO4
86 33641 52312 4 40S 45453
87 26.232 34899 5 42O 38909
88 2SOO1 26889 6 451 41785
89 25151 45733 7 449 40829
90 39127 S4393 8 443 33O16
91 2S2O4 3841S
92 2951O 57O16 9 429 47621
93 2836S 44002 10 440 44767
94 27723 54297 11 383 599.93
95 3OO73 40586 12 339 45096
96 24243 3S190 13 239 52.275
14 360 50065
15 10 44358
0096) 16 336 42785
TABLE 58 17 469 321.93
18 541 30172
Diplex 19 311 33607
2O 454 34474
Swab # GC IAC 21 289 40475
73 13.7 38.6 22 396 31906
74 O 39.9 23 3O8 32182
75 27.8 39.3 24 28O 33715
76 11.4 37.7
77 29.2 38.3
78 28.3 41.6
79 33.2 36.2 0100
8O 29 41.1
81 33.5 31.9
82 33.5 35.2 TABLE 60
83 28.8 37.1
84 33.3 38.9 Diplex
85 36.6 41.4
86 34.5 38.8 Swab # GC IAC
87 35.8 40.8
88 38.5 34.1 1 O 44.6
89 36 34.6 2 O 42.6
90 33.1 41.2 3 O 43.1
91 37.1 31 4 O 42
92 36.8 27.8 5 O 42
93 30.7 41.2 6 O 41.5
94 33.9 40.4 7 O 423
95 33.6 39.5 8 O 39.2
96 38.1 38.7 9 O 42.9
10 O 42.7
11 O 43
12 O 43.4
Example 3 13 O 4.1.8
14 O 4.1.8
15 O 42.6
Day 5— After Cleaning 16 O 42.4
17 O 43.5
Comments: 18 O 39.9
0097 84/96 swabs tested negative in both assays after 5. g 2. 8
cleaning the Surface with hydrogen peroxide resulting in a 21 O 42.8
reduction rate of 88%. 22 O 41.2
23 O 4.1.8
0098. Dilute Hydrogen Peroxide was liberally poured 24 O 41.4
onto the 48 contaminated squares, allowed to stand for 3
minutes, then wiped away in a one-directional motion.
US 2007/0289605 A1 Dec. 20, 2007
19

0101 0103)
TABLE 61 TABLE 63

Monoplex Monoplex

Swab # GC AC Swab # GC AC

25 4O6 43661 49 301 44083

26 688 45628 50 78 58387
27 364 S1911 51 148 42208
28 412 S1949 52 117 S2129
29 602 41483 53 50 S9421
30 612 41653 S4 77 52677
31 359 S10O2 55 171 57628
32 337 43837 56 351 48090
33 464 55.289 57 821 425.18
34 652 56726 58 527 56048
35 828 491.75 59 612 64727
36 574 53555 60 330 56105
37 444 S3683 61 769 56571
38 717 4.0472 62 840 4792O
39 415 38670 63 333 4O764
40 411 35603 64 778 27526
41 1363 34O21 65 419 454.66
42 4O1 34518 66 509 44726
43 11094 29699 67 865 3S466
768 35252 68 806 46479
45 11945 37411 69 1008 56753
46 24287 35033 70 5148 444S5
47 741 3933O 71 338 55593
48 365 32297 72 604 32S14

0102) 0.104)
TABLE 62 TABLE 64
Diplex Diplex
Swab # GC IAC Swab # GC IAC

25 42.9 49 41
26 423 50 33.7
27 43.7 51 42
28 42 52 41.7
29 41.3 53 38.8
30 43.3 S4 38.8
31 41.5 55 41
32 39.7 56 29.3
33 43.1 57 43.7
34 41.1 58 43.7
35 42.7 59 43.9
36 41.5 60 43.2
37 39.6 61 42.8
38 40.8 62 44.4
39 42.5 63 41.9
40 40.9 64 25.5
41 423 65 40.9
42 42.5 66 41
43 41.4 67 43.5
423 68 4O.S
45 42.1 69 43.8
46 39.3 70 39.8
47 40.9 71 35.6
48 41.2 72 32
US 2007/0289605 A1 Dec. 20, 2007
20

01.05
TABLE 67
TABLE 65
Monoplex
Monoplex
Swab # GC AC
Swab # GC AC
34218 28572
73 1010 S3281 34731 37477
74 623 59035 33560 28.999
75 5170 39881 32114 28145
76 263 46845 32970 28804
77 523 S6542 27705 23330
78 264 53710 2S646 26112
79 347 53.738 22720 30697
8O 745 35921 24O61 S2874
81 352 40240 1 23719 422O3
82 398 44022 26768 54382
83 585 55182 24246 33472
84 681 47458 2492O 35321
85 2716 411.83 22810 32819
86 1032 38811 234.72 31461
87 1139 4.5937 24918 32090
88 547 43834 24962 33355
89 440 23338 2S639 395O2
90 745 24383 3.1440 43734
91 561 31253 2S346 3128O
92 547 31604 27893 40488
93 588 34870 25817 35960
94 796 33059 29259 31 616
95 831 29424 25597 34019
96 411 27360

0108)
0106 TABLE 68
TABLE 66 Diplex
Diplex Swab # GC IAC

Swab # GC IAC 33.3 38.5

39.5 35.5
73 O 39.3 37.6 38.5
74 O 23.7 17.6 41.1
75 O 34.6 39.5 31.8
76 2O.S 14.1 35.7 37.5
77 O 40.7 36.1 36.7
78 O 37.9 33.8 38.9
79 32.3 39 34.9 36.1
8O O 36 1 36.5 4O.S
81 O 38.9 39.3 35.5
82 O 41.2 O 41
83 O 44.2 37.3 37.4
84 15.1 22.9 35.9 37
85 O 40.9 36.7 33
86 O 41.9 32.2 37.8
87 O 41.4 8.8 39.2
88 O 4.O.S O 43.9
89 13.9 42.6 41.5 38.2
90 O 39.3 O 43.6
91 O 29.3 38.5 37.2
92 15.9 41.7 37.1 43.8
93 O 41.9 40.7 37.3
94 O 40.8 38.1 32.8
95 O 39.1
96 O 40.9
01.09)
Example 3 TABLE 69
Monoplex
Day 8 Before Cleaning
Swab # GC AC
0107 All swabs are positive for GC in at least one assay 25 28829 35884
(either monoplex or diplex). Therefore according to AV-17 26 2912S 48436
Verification, the study can proceed to step 4.4.2.
US 2007/0289605 A1 Dec. 20, 2007
21

TABLE 69-continued TABLE 71-continued

Monoplex Monoplex
Swab # GC AC Swab # GC AC

27 301.07 4.0476 51 29920 44-381

28 27001 40800 52 28790 47069
29 28841 45752 53 2S462 38O26
30 26529 32369 S4 28008 4663S
31 28649 3871.4 55 32136 43514
32 26641 25595 56 27957 47381
33 241.95 49374 57 27346 30722
34 2.7538 47288 58 33807 45084
35 29941 43319 59 30774 32495
36 26906 SO361 60 31358 3O388
37 28763 46044 61 3O82O 51444
38 2S489 40462 62 28517 469SO
39 3O482 46252 63 29792 39421
40 22100 42740 3. 2.
41 25416 22918 66 29721 474.09
42 28381 411.75 67 2.7028 59023
43 2SO12 3.3289 68 30286 46546
44 28297 41713 69 288.18 51287
45 2996.1 33447 70 24318 31282
46 33681 39634 71 24340 31622
47 294OO 4S541 72 19194 28583
48 2S889 40960

0112
0110
TABLE 72
TABLE 70
Diplex
Diplex
Swab # GC IAC
Swab # GC IAC
49 31.5 39.2
25 39.6 38.7 50 36 36.1
26 37.6 43.2 51 33.6 37.9
27 O O 52 35.4 37.7
28 38.6 39.3 53 24.4 36.6
29 38.7 36.2 S4 31.6 36.3
30 39 39.5 55 37 30.8
31 39.2 35.4 56 21.2 30.8
32 38.4 32.6 57 36.4 34.4
33 34 40.1 58 35.6 3O.S
34 37.6 41 59 36 34
35 O 13.3 60 33.9 30.7
36 35.8 33.6 61 31.2 35.4
37 38.9 40.3 62 35.9 32.5
38 34.7 41.3 63 34.6 36.8
39 27.7 22.5 64 35.8 30.8
40 37.9 34.5 65 28.7 27.9
41 33.7 32.3 66 32.5 28
42 24.1 36.3 67 31.1 33.7
43 32.5 23.6 68 32.6 31.3
44 34.4 30.6 69 16.4 28.4
45 33.2 28.7 70 19.2 23.2
46 24.2 22 71 32.3 33.7
47 32.3 34.8 72 21.4 31.3
48 10.6 30.6

0113)
0111
TABLE 73
TABLE 71
Monoplex
Monoplex
Swab # GC AC
Swab # GC AC
73 26529 39221
49 2.7932 37044 74 28321 SO860
50 285.15 48930 75 32996 41992
US 2007/0289605 A1 Dec. 20, 2007
22

then wiped away in a one-directional motion. These steps

TABLE 73-continued were repeated and then 2 Swabs were taken from each
Monoplex square, processed and tested.
Swab # GC AC 0.116) Positive readings=Monoplex>2000; Diplex>0
76 33,774 S4266 TABLE 75
77 33622 396.13
78 36040 43275
79 31719 54146 Monoplex
8O 291.56 39.048
81 17727 24910 Swab # GC AC
82 17989 26189
83 1946O 393OO 1 42O 35365
84 19777 29916 2 395 35670
2. 3. 3 425 42341
87 22SO4 34760 4 435 34112
88 21937 38570 5 469 34857
89 22723 16069 6 509 28802
90 27638 17416 7 283 35825
3. i. 3. 8 SO8 42276
93 18370 17640 9 323 3.2138
94 2O528 16623 10 392 32658
95 18283 19858 11 164 33870
96 17648 17270 12 173 31391
13 6571 33740
14 611 28683
0114 15 1482 28890
16 624 33873
TABLE 74 17 217 36900
18 665 3461O
Diplex 19 379 35350
2O 567 34898
Swab # GC IAC 21 380 37342
73 11 14.6 22 658 32614
74 36.9 27 23 28O 37890
75 26.8 30.6 24 589 36614
76 29.1 26.5
77 37 20.7
78 35.2 31.6
79 23.9 29.8 0117)
8O 25.9 35.5
81 37.2 35.8
82 36.3 34.7 TABLE 76
83 21.5 39.7
84 19.4 22.2 Diplex
85 25.9 30.3
86 34.7 35.3 Swab # GC IAC
87 15.7 26.2
88 38.9 25.8 1 31.9 40.8
89 35.2 34 2 14.6 35.6
90 28.5 32.1 3 O 36
91 31.4 34.2 4 O 41.3
92 26.4 38.5 5 27.3 38.9
93 O 41.3 6 O 39.9
94 O 39.5 7 O 39.9
95 31.1 11.5 8 O 41
96 33.2 29.2 9 O 423
10 O 38
11 O 38.9
12 O 43
13 O 40.8
Example 3 14 O 37.8
15 O 37.6
Day 8 After Cleaning 16 O 39
17 O 44.1
Comments: 18 31.5 41.7
19 27.1 43.2
78/96 swabs tested negative in both assays after cleaning the 3. g t
Surface with hydrogen peroxide resulting in a reduction rate 22 O 43.1
of 81%. 23 O 40.8
24 O 42.9
0115) Hydrogen Peroxide was liberally poured onto the
48 contaminated squares, allowed to stand for 3 minutes,
US 2007/0289605 A1 Dec. 20, 2007
23

0118 0120
TABLE 77 TABLE 79

Monoplex Monoplex

Swab # GC AC Swab # GC AC

25 345 324O7 49 588 32098

26 60S 35031 50 631 39291
27 563 49398 51 1441 40568
28 12896 33637 52 5856 42414
29 6213 33O46 53 529 33702
30 6O1 27879 S4 490 44.296
31 502 22786 55 2O2 35479
32 715 16557 56 433 55877
33 510 335O2 57 231 26525
34 778 46897 58 496 34.609
35 456 366OS 59 1073 29498
36 458 37934 60 1045 47527
37 505 34226 61 1290 38073
38 579 27248 62 724 34092
39 26S 26513 63 939 34940
40 116 24731 64 648 34841
41 555 19289 65 404 39464
42 252 19429 66 388 421.25
43 223 22409 67 565 446O7
344 2O63S 68 904 47893
45 240 2O295 69 656 3S4O3
46 438 19267 70 319 36.662
47 821 17697 71 10088 2S822
48 381 18837 72 483 17999

0119) 0121)
TABLE 78 TABLE 8O
Diplex Diplex
Swab # GC IAC Swab # GC IAC

25 O 34 49 37.8
26 O 42 50 40.6
27 O 42.7 51 38.7
28 14.5 32.9 52 38.6
29 18.7 40.7 53 36.5
30 31.5 S4 36.2
31 25.4 37.4 55 34.6
32 O 39.1 56 24.9
33 39.5 57 38.7
34 36.4 58 41.2
35 38.6 59 40.8
36 41.5 60 39.7
37 43.4 61 41
38 37.4 62 38.9
39 40.3 63 40.2
40 28.3 64 41.7
41 39.7 65 1 35.6
42 40.9 66 32.6
43 41.3 67 34.2
38.8 68 33.3
45 40.8 69 36.6
46 42.2 70 34.7
47 42.1 71 36
48 31.4 72 2 32.7
US 2007/0289605 A1 Dec. 20, 2007
24

0122) wiping the hydrogen peroxide and water solution from the
Surface.
TABLE 81 2. The method of claim 1 further comprising the step of
Monoplex allowing the hydrogen peroxide and water Solution to stand
for at least about 3 minutes before wiping.
Swab # GC AC 3. The method of claim 1 wherein the concentration of
73 807 42662 hydrogen peroxide in the solution is between about 0.5%
74 378 46540 and about 30%.
75 917 4632O 4. The method of claim 1 wherein the concentration of
76 590 SO889
77 703 32475
hydrogen peroxide in the solution is between about 2% and
78 636 43325 about 10%.
79 269 47888 5. The method of claim 1 wherein the concentration of
8O 556 47134 hydrogen peroxide in the solution is about 3%.
81 409 36989
82 326 42705 6. The method of claim 1, wherein the method comprises
83 622 44882 the additional step of rinsing the surface with water.
84 390 37163
85 573 33.701 7. The method of claim 1, wherein the contamination
86 658 3.2927 comprises radioactive contaminants.
87
88
609
431
43897
46131
8. A method of reducing nucleic acid contamination on an
89 13750 34084
item comprising:
90 442 35375
91 972 38919 soaking the item in a solution consisting essentially of
92 435 39781 hydrogen peroxide and water.
93 837 41371
94 309 48695
9. The method of claim 8, wherein the concentration of
95 396 36321 hydrogen peroxide in the solution is between about 0.5%
96 366 31143 and about 30%.
10. The method of claim 8, wherein the concentration of
hydrogen peroxide in the solution is about 3%.
0123) 11. The method of claim 8, wherein the contamination
comprises radioactive contaminants.
TABLE 82 12. The method of claim 1, in which the contacting step
Diplex
is performed by soaking a towel or a wipe in a solution
consisting essentially of hydrogen peroxide and water; and
Swab # GC IAC
wiping the surface to be cleaned with the soaked wipe or
73 38.4 towel.
74 20.6
75 34.2 13. The method of claim 12, wherein the concentration of
76 1 1 35 hydrogen peroxide in the solution is between about 0.5%
77 38.7 and about 30%.
78 38.3
79 38.2 14. The method of claim 12, wherein the concentration of
8O 35 hydrogen peroxide in the solution is about 3%.
81 38.9
82 38.7
15. The method of claim 12, further comprising the step
83 39.1 of wiping the surface with a dry towel or wipe.
84 35.5 16. The method of claim 13, further comprising the step
85 36.6
86 36.7
of wiping the surface with a towel soaked with water.
87 37.3 17. The method of claim 13, further comprising the step
88 35.7 of rinsing the surface with water.
89 38.5
90 1 4 37.9 18. A method of reducing nucleic acid contamination on
91 41.3 a Surface consisting of
92 39
93 2 37.2 contacting the Surface to be decontaminated with a solu
94 37.3 tion consisting essentially of hydrogen peroxide and
95 35.3
96 35.6 water, and
wiping the Solution from the Surface.
19. The method of claim 18, wherein the concentration of
We claim: hydrogen peroxide in the solution is between about 0.5%
1. A method of reducing nucleic acid contamination on a and about 30%.
Surface comprising: 20. The method of claim 18, wherein the concentration of
contacting the Surface to be decontaminated with a solu hydrogen peroxide in the solution is about 3%.
tion consisting essentially of hydrogen peroxide and
water, and