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TEST REPORT
Send to:
Neogen Corporation
620 Lesher Place
Lansing, Michigan 48912
Result: COMPLETE
Customer Name:
Location of Testing:
Description:
Test Type:
Job Number:
Project Number:
10009584
NSF Corporate:
C0245040
Project Manager:
J. Vantine
Executive Summary:
Neogen Corporation contracted the Applied Research Center at NSF International to perform comparison
performance testing of Neogens ATP system, AccuPoint Advanced, against four other commercially available
systems. The recovery efficiencies and consistency of each system were evaluated against an ATP standard
solution and orange juice at different dilutions inoculated onto stainless steel carriers. This study utilized several
methods to determine the effectiveness of each system. Both a directly pipetted ATP standard solution and
commodity onto swab surfaces and surface swabbing of stainless steel coupons were employed with the test
systems.
Neogens AccuPoint Advanced ATP system consistently yielded the highest percent recoveries and the most
consistent readings of the target analytes, when compared to the other four test systems.
ATP devices are utilized to detect the presence of bacteria and organic/food residue on surfaces. ATP has been
incorporated as a key monitoring parameter for the food, beverage and healthcare industries. It is essential that
these devices provide precise and consistent readings so that the hygiene practices of these industries can be
accurately evaluated.
Thank you for having your product tested by NSF International.
Please contact your Project Manager if you have any questions or concerns pertaining to this report.
Digitally signed by Dr. Robert Donofrio /jv
DN: cn=Dr. Robert Donofrio /jv, o=NSF
International, ou=Director - Applied Research
Center, email=Donofrio@NSF.ORG, c=US
Date: 2015.07.13 10:21:51 -04'00'
Report Authorization:____________________________________________
Robert Donofrio Director, Applied Research Center
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TEST REPORT
Neogen Corporation provided its system and the reference standard ATP, all other devices were purchased
independently through normal commercial channels by NSF International.
Each system was tested using 4 differing approaches. Details provided in Methodology.
Section 1: ATP standard solutions were pipetted directly onto sample swabs. Data obtained here was used as a
reference for the calculation of ATP recovery in sections 2 and 3. (Data recorded under NSF J-00170817)
Section 2: ATP standards were deposited over a 4x4 stainless steel surfaces and the above referenced
monitoring systems were used to sample the entire surface under a real world approach*.
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TEST REPORT
Section 3: ATP was recovered from a concentrated spot randomly located on 4x4 stainless steel surface
utilizing all 5 test systems detailed above with a real world swabbing approach. In the real world, residue is not
homogeneously distributed across a surface after cleaning. The spot test attempted to recreate this phenomenon by
randomly placing a small, invisible dot of ATP on the stainless steel surfaces to determine the extent and
consistency of each systems ability to locate and accurately identify its presence. 10 replicates of each test were
completed on a single test day for each reader.
Section 4: Orange juice was used as a standard commodity to measure the detection ability of each of the test
units. Orange juice was deposited over a 4x4 stainless steel surface that was sampled utilizing all 5 test systems
detailed above with a real world swabbing approach.. 10 replicates of each test were completed on a single test
day for each reader.
1:1000(1partorangejuiceto999partssterilewater),
1:5000(1partorangejuiceto4999partssterilewater)and
1:10000(1partorangejuiceto9999partssterilewater).
*Real world approach is defined wherein each 4X4 surface was sampled in a cross-hatch pattern with five
seconds in each direction. This measurement was adapted to replicate real-world operation of these systems that
most often occurs between tightly scheduled production runs where sampling speed is important.
Methodology
These following methods were derived from document Sanitation Sampler Assay Protocol, provided by Brent
Steiner and Ron Sarver of Neogen Biochemistry Laboratory on December 30, 2014. (See Addendum B for
complete text)
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tested.
TEST REPORT
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tested.
TEST REPORT
Sterile water and isopropanol used for cleaning was dispensed from a spray bottle which was sterilized using a
UV lamp (20 minute exposure) and 10% Contrad 70 in water. Periodically a 4x4 plate was checked for
cleanliness using an AccuPoint sampler to ensure the reading was at background levels (below 25 RLU).
100 femtomoles of ATP was spread over the 4x4 surface and air dried for 1 hour at room temperature (1825C) to measure the amount of ATP that could be recovered utilizing each monitoring system. This was
accomplished by pipetting 20 L of a 5.0 nM ATP solution onto the stainless steel surface. The tip of a pipette
was used to distribute the solution over the surface. Ten 4x4 stainless steel squares were covered with 100
femtomoles of ATP for each sanitation system. After the ATP was deposited homogenously across and dried, the
surface was sampled using the sanitation system sampler in the manner recommended by the manufacturers
instructions. The amount of ATP was recovered was determined by comparing the mean response from the
surface recovery to the mean response obtained in Section1. Results are reported in Table 2.
Section 3. The goal of this experimental section was to determine the efficiency of the five test systems in
recovering an ATP standard that had been spot inoculated at a random location on a stainless steel carrier. A 20L
of the 5.0 nM ATP solution (100 femtomoles) was pipetted at a random spot on a 4x4 stainless steel surface to
determine the recovery capability of each monitoring system. The spot was allowed to dry for 1 hour and the plate
was sampled according to the manufacturers sampling instructions. This was repeated 10 times for each sanitation
monitoring system to determine the mean response, standard deviation and the coefficient of variation (CV) for
the recovered ATP from the surface. The percentage recovered from the surface was determined by comparing the
mean response from the surface spot recovery to the mean response obtained in Section 1. Results are reported in
Table 3.
Section 4. The goal of this experimental section was to determine the efficiency and detection limit of the five test
systems in recovering a standard commodity, orange juice, which had been evenly spread across a stainless steel
carrier. This experiment was designed to replicate a typical situation that would be encountered in the field. For
this evaluation, 10mL of orange juice was diluted 1:1000 (1 part orange juice to 999 parts sterile water), 1:5000 (1
part orange juice to 4999 parts sterile water), and 1:10000 (1 part orange juice to 9999 parts sterile water).
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TEST REPORT
Surfaces for each dilution level were prepared by dispensing 50 L of a given dilution level across the surface of
a 4x4 section of stainless steel plate and allowing the samples to dry for 1 hour before sampling the surface
according to the prescribed method for each brand of system. Ten surfaces were prepared and sampled at each
dilution for each brand of sanitation systems. The mean, standard deviation and coefficient of variation was
determined for each dilution and each brand of sanitation sampler.
To determine recovery, 50 L of each orange juice dilution was pipetted directly onto the swab or sample pad and
the response measured using each brand of sanitation system. This was repeated ten times to determine the mean
response for directly pipetting the orange juice dilution onto the sampler. The percentage recovered from the
surface was determined by comparing the mean response from the surface recovery to the mean response obtained
from directly pipetting 50 L onto the samplers. Results are reported in Table 4.
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tested.
TEST REPORT
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tested.
TEST REPORT
real world situational sampling method utilizing each of the 5 monitoring systems to determine the mean response
of each unit.
The percentage recovery was calculated by comparing the mean response from the surface spot recovery to the
mean response of direct swab inoculation observed in Section 1. The results for Section 3 can be found in Table 3.
The Neogen AccuPoint Advanced had the highest percentage recovery of all 5 monitoring systems at 40.50%
recovery of the ATP solution from the unit surface. The Neogen AccuPoint Advanced system exhibited a percent
ATP recovery that was 2xs greater than the next most efficient monitoring system. The Neogen Accupoint
system exhibited the greatest consistency in readings (with a CV of 21.11%), indicating that the system is very
precise. The next closest system was BioControl MVP system at 17.93% recovery.
In Section 4 the experimental protocol was designed to mimic real world contamination scenarios. This study
involved contaminating stainless steel surfaces with orange juice at 3 dilutions: 1:1,000, 1:5,000 and 1:10,000.
RLU reference values for each dilution were first generated by direct inoculation onto the ATP monitoring system
swabs. Recovery sampling using a real world approach, as previously described, was performed on
homogenously inoculated stainless steel surfaces. The percentage recovered from each surface was determined
by comparing the RLU of the surface reading with the RLUs observed from direct swab inoculation. Table 4
provides the results for the RLUs observed from direct inoculation (4a), RLUs from stainless steel recovery (4b)
and calculated percent ATP recovery (4c). Once again, the Neogen AccuPoint Advanced had the highest
observed percentage recovery of all 5 monitoring systems. For each of the orange juice dilutions evaluated, the
percent recovery of ATP by the Neogen AccuPoint Advanced was significantly higher than that of the other four
ATP monitoring systems evaluated. Once again, the Neogen Accupoint system proved to be the most consistent
of the devices evaluated (with a CV of 40.58%). The next closest system for recovery at 1:1000 and 1:5000
dilution factors was BioControl MVP. For the 1:10000 dilution factor the 2nd highest recovery was the 3M
CleanTrace.
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tested.
TEST REPORT
Scope of Work
Disclosure of Deviations
On 5/08/2015 the stainless steel coupon cleaning was validated by 2 Neogen AP swab readings. The readings
were below the 25 RLU background thresholds (4 RLU, 11 RLU) and testing proceeded for the 10 replicate
samples of the AccupPoint 3.04, EnSure and MVP readers. However, each of the three readers had 1 or 2 outlier
samples indicating coupon contamination. Outlier is defined as a sample with an RLU value outside the range
observed when pipetting the orange juice directly on the swab.) After this day of testing, coupons were re-cleaned,
sampled for cleanliness (0 RLU, 0 RLU), and re-tested. Summary of results reported in Table 4 are the data
recovered on the retest date.
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tested.
TEST REPORT
Appendix A
Result Tables
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TEST REPORT
Table 1. Mean RLU response of five ATP monitoring systems against an ATP standard of 100 femtomoles.
ATP standard solutions were pipetted directly onto sample swabs. Mean was taken of 25 replicates (n=25) and
reported. Data obtained was used as a reference for the calculation of ATP recovery in Sections 2 and 3, Tables 2
and 3 respectively. (Data recorded under NSF J-00170817)
Reader
Neogen
AccuPoint 3.04
3M
NG
Hygiena
EnSure
Charm
NovaLUM
BioControl
MVP
AccuPoint
Advanced
CleanTrace
UltraSnap
PocketSwab Plus
Lightning
593.32
871.56
206.64
29,809.04
594.12
Sampler
Mean RLU
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tested.
TEST REPORT
Table 2. Recovery of ATP standards from a homogenously contaminated stainless steel surface. A 100
femtomole ATP standard solution was homogenously deposited over a 4x4 stainless steel surface. The surface
was then sampled with the referenced monitoring systems utilizing a real world approach swabbing technique (5
seconds in one direction, and then swabbing 5 more seconds perpendicular to the first). The mean RLU was
calculated of 10 replicates (n=10) and reported. To calculate percent ATP recovery, the direct inoculation mean
RLU recovery values (Table 1) were used as a reference.
Reader
FI20150713100356
3M
Hygiena
Charm
BioControl
NG
EnSure
UltraSnap
NovaLUM
MVP
PocketSwab
Plus
Lightning
AccuPoint
Advanced
CleanTrace
Average
165.2
62.8
31.1
8,618.10
123.7
Std Dev
34.87
20.08
18.62
5,236.99
47.34
%CV
21.11%
31.98%
59.86%
60.77%
38.27%
Average
27.84%
7.21%
15.05%
28.91%
20.82%
Samplers
Mean RLU
Recovered
from
Surface
% ATP
Recovery
from
Surface
Neogen
AccuPoint
3.04
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tested.
TEST REPORT
%Recovery
35
30
25
20
%Recovery
15
10
5
0
Neogen
3M
Hygenia
Charm
Biocontrol
Figure 1: provides a pictorial representation of Table 2: Recovery of ATP standards from a homogenously
contaminated stainless steel surface.
%CoefficientVariance
70.00%
60.00%
50.00%
40.00%
%CoefficientVariance
30.00%
20.00%
10.00%
0.00%
Neogen
3M
Hygenia
Charm
Biocontrol
Figure 2: provides a pictorial representation of Table 2. Coefficient of Variance (%) was calculated and lowest %
indicates the most consistent (least variable) readings.
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tested.
TEST REPORT
Table 3. Recovery of an ATP standard from a single contamination spot on stainless steel surfaces. A 100
femtomole ATP standard solution was deposited randomly as a drop on a 4x4 stainless steel surface. The
surface was then sampled with the referenced monitoring systems utilizing a real world swabbing approach
technique (5 seconds in one direction, and then swabbing 5 more seconds perpendicular to the first). The mean
RLU was calculated of 10 replicates (n=10) and reported. To calculate percent ATP recovery, the direct
inoculation mean RLU recovery values (Table 1) were used as a reference.
Reader
FI20150713100356
3M
Hygiena
Charm
BioControl
NG
EnSure
UltraSnap
NovaLUM
MVP
PocketSwab
Plus
Lightning
AccuPoint
Advanced
CleanTrace
Average
240.3
71.3
20
3,435.40
106.5
Std Dev
97.51
45.34
13.25
2,900.68
88.79
%CV
40.58%
63.58%
66.25%
84.43%
83.37%
Average
40.50%
8.18%
9.68%
11.52%
17.93%
Samplers
Mean RLU
Recovered
from
Surface
% ATP
Recovery
from
Surface
Neogen
AccuPoint
3.04
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tested.
TEST REPORT
%Recovery
45
40
35
30
25
%Recovery
20
15
10
5
0
Neogen
3M
Hygenia
Charm
Biocontrol
Figure 3: provides a pictorial representation of Table 3: Recovery of an ATP standard from a single
contamination spot on stainless steel surfaces.
%Recovery
45
40
35
30
25
%Recovery
20
15
10
5
0
Neogen
3M
Hygenia
Charm
Biocontrol
Figure 4: provides a pictorial representation of Table 3: Recovery of an ATP standard from a single
contamination spot on stainless steel surfaces. Coefficient of Variance (%) was calculated and lowest %
indicates the most consistent (least variable) readings. Table 4. Recovery of ATP from stainless steel surfaces
inoculated with varying concentrations of orange juice. Orange juice was selected as an ATP source since it is
a standard commodity product. Three dilutions of orange juice were utilized: 1:1,000, 1:5,000 and 1:10,000.
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tested.
TEST REPORT
Table 4a. RLU values observed when the dilutions were amended directly to the ATP monitoring systems
sample pad/swab. These RLU values were used as a reference for the calculation of percent ATP recovery in
Table 4c. The average RLU reading from 10 replicates (n=10) is reported.
Mean RLU for Recovery of Orange Juice Pipetted onto the Sample Pad/ Swab
Samplers
Average
Orange
Juice
Dilution
1:1000
Neogen
AP
Advanced
1,783.4
3M
CleanTrace
3,629.1
Hygiena
UltraSnap
639.6
Charm
PocketSwab
Plus
145,735.9
BioControl
Lightning
2,071.9
Average
1:5000
418.5
832.6
165.6
34,517.6
582.4
Average
1:10000
90.7
217.5
34.0
6,394.1
139.9
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tested.
TEST REPORT
Table 4b. RLU values observed from sampling 4x4 stainless steel surfaces amended with three dilutions of
orange juice. Percent recovery was calculated by dividing the mean RLU values below (homogenous stainless
steel coupon inoculation) by the mean RLU values in Table 4a (direct swab inoculation).
The dilutions were applied homogenously and the surfaces were sampled utilizing with a real world swabbing
approach (5 seconds in one direction, and then swabbing 5 more seconds perpendicular to the first). The average
RLU reading from 10 replicates (n=10) is reported.
Recovery of Orange Juice from 4"x4" Stainless Steel
Neogen AP
Advanced
3M
CleanTrace
Hygiena
UltraSnap
Charm
PocketSwab
Plus
BioControl
Lightning
553.3
71.4
65.7
14,468.30
271
31.03%
1.97%
10.27%
9.93%
13.08%
%CV
33.1%
74.3%
37.2%
47.6%
55.3%
Average
119.8
48.2
27.9
2,115.40
148.1
28.63%
5.79%
16.85%
6.13%
25.43%
46.7%
32.0%
43.3%
36.5%
38.8%
14
26.6
14.4
10.5
15.44%
12.23%
0.00%
0.23%
7.51%
146.7
111.52
NA
316.2
31.2
Samplers
Orange Juice
Dilution
Average
% ATP
Recovery
% ATP
Recovery
1:1,000
1:5,000
%CV
Average
% ATP
Recovery
%CV
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1:10,000
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TEST REPORT
Table 5: Stainless steel coupon cleaning readings were completed each test date by 2 Neogen AP Advanced
swabs of 2 representative coupons. All readings were at 0 RLU on days that data was reported from. Data
recorded on 5/8/2015 was dismissed due to outlier readings and repeated.
5/8/2015
5/11/2015
5/12/2015
5/13/2015
5/14/2015
5/15/2015
5/19/2015
5/20/2015
5/21/2015
5/22/2015
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RLU
Replicate 1 Replicate 2
4
11
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
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TEST REPORT
Appendix B
Sanitation Sampler Assay Protocol
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TEST REPORT
J-00175137
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TEST REPORT
To determine the amount of ATP recovered from a stainless steel surface 100 femtomoles of ATP should be
spread over the 4x4 surface and air dried for 1 hour at room temperature (18-25C). This is accomplished by
pipetting 20 L of a 5.0 nM ATP solution onto the stainless steel surface. The tip of the pipette can be used to
distribute the solution over the surface. Ten 4x4 stainless steel squares should be covered with 100 femtomoles
of ATP for each sanitation system being evaluated. Care should be taken to not spread the solution with
something other than the pipette tip since other materials may remove the deposited ATP from the surface. After
the ATP is deposited and dried, the surface should be sampled using the sanitation system sampler in the manner
recommended by the manufacturer. The amount of ATP recovered is determined by comparing the mean
response from the surface recovery to the mean response obtained in Section 1 from directly pipetting 100
femtomoles of ATP onto the samplers.
Section 3). For recovery of a concentrated dot of ATP randomly spotted on a
4x4 stainless steel surface, 20L of the 5.0 nM ATP solution (100 femtomoles) is pipetted at a random spot on
each plate. The spot is then allowed to dry for 1 hour and the plate sampled according to the sampling method
prescribed by the manufacturer. This is repeated 10 times for each sanitation monitoring system to determine the
mean response, standard deviation and coefficient of variation (CV) for the recovered ATP from the surface. The
percentage recovered from the surface is determined by comparing the mean response from the surface spot
recovery to the mean response obtained in Section 1 from directly pipetting 100 femtomoles of ATP onto the
samplers..
Section 4). Commodity testing with orange juice is completed to determine recovery from a stainless steel surface
and determine the limit of detection in a more typical situation that would be encountered in the field. For this
evaluation, 10mL of orange juice is diluted 1:1000 (1 part orange juice to 999 parts sterile water), 1:5000 (1 part
orange juice to 4999 parts sterile water), and 1:10000 (1 part orange juice to 9999 parts sterile water). Surfaces
for each dilution level were prepared by dispensing 50 L of a given dilution level across the surface of a 4x4
section of stainless steel plate and allowing the samples to dry for 1 hour before sampling the surface according to
the prescribed method for each brand of sampler. Care should be taken to not spread the solution with something
other than the pipette tip since other materials may remove the deposited orange juice from the surface. Ten
surfaces should be prepared and sampled at each dilution for each brand of sanitation systems. The mean,
standard deviation and coefficient of variation should be determined for each dilution and each brand of sanitation
sampler.
To determine recovery, 50 L of each orange juice dilution should be pipetted directly onto the swab or sample
pad and the response measured using each brand of sanitation sampler. This is repeated ten times to determine
the mean response for directly pipetting the orange juice dilution onto the sampler. The percentage recovered
from the surface is determined by comparing the mean response from the surface recovery to the mean response
obtained from directly pipetting 50 L onto the samplers.
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tested.