Technical Report Ames Assay Proficiency Study · served as initial Study Trial Coordinator, followed by Betsy Bombick. 4. Study Protocol The protocol, summarized below, was reviewed

In Vitro Toxicity Testing of Tobacco Smoke Sub-Group

Technical Report

Ames Assay

Proficiency Study

March 2016

Author:

Wendy Wagstaff

Labstat International ULC, Canada

Trial Coordinators:

Betsy Bombick

R.J. Reynolds, USA

T.S. Kumaravel

Advanced Technologies (Cambridge) Ltd, UK

Task Force Coordinator:

Kei Yoshino

Japan Tobacco Inc., Japan

Table of Contents

1. Summary ........................................................................................................................... 3

2. Introduction ....................................................................................................................... 3

3. Participants ........................................................................................................................ 4

4. Study Protocol ................................................................................................................... 4

5. Results – Raw Data ........................................................................................................... 5

6. Statistical Analysis and Interpretations ............................................................................. 6

6.1 Reported Dose Ranges and Solvent Controls ............................................................ 6

6.2 Specific Activity ........................................................................................................ 6

6.3 Effect of S9 Percentage ............................................................................................. 7

6.4 Differentiating Test Brands ....................................................................................... 8

6.5 Comparison among Labs ........................................................................................... 9

7. Discussion of Results ...................................................................................................... 12

7.1 Data Audit ................................................................................................................ 12

7.2 Audit Findings ......................................................................................................... 12

8. Conclusions ..................................................................................................................... 12

9. References ....................................................................................................................... 13

Appendix A – Raw Data .......................................................................................................... 14

A-1 Raw Revertant Colony Count Data .................................................................. 14

Appendix B – Calculated Specific Activities (revertants/mg NFDPM) .................................. 32

IVT-077-CTR Ames Assay Proficiency Study – March 2016 3/33

1. Summary

An Ames Assay proficiency study was conducted by member laboratories of the CORESTA

In Vitro Toxicity Testing Task Force (a Sub-Group since 2015). Given that different

procedures (internal or regulatory) are followed by the participating laboratories, the purpose

was not to develop a CORESTA Recommended Method but to evaluate the mutagenicity of a

mainstream smoke extract from two cigarettes using a common Study Plan. This enabled

laboratories to evaluate method proficiency, compare results with those of other laboratories,

and obtain an external audit of their documentation procedures that might identify potential

areas for improvement. Each laboratory therefore has been able to benefit both technically

(scientific methods) and administratively (documentation procedures) from their participation.

Twelve laboratories participated in the study and the protocol was approved by each

laboratory manager. Ames assays were conducted using smoke condensates (particulate

matter collected on Cambridge filter pads and extracted in dimethylsulfoxide) on two

cigarettes.

The cigarettes were coded as follows:

A, C, E: Canadian Monitor 8 (CM8)

B, D, F: Kentucky Reference 3R4F (3R4F)

Smoke condensates were prepared using the ISO smoking regimen at one laboratory, these

were frozen at approximately -70°C, and shipped on dry ice to each laboratory. Ames assays

were conducted according to the protocol, using bacterial strains of TA98 and TA100 ± S9

metabolic activation. All laboratories used 5% S9 mix; some laboratories also used 10% S9

mix. At least six treatment concentrations were evaluated, with at least three replicate plates

per concentration. Raw data, worksheets and summary conclusions were submitted to the

Study Coordinator. Worksheets were assessed by the Task Force auditor for quality

assurance. Raw data was coded, and a common statistical analysis was conducted by the

Study Statistician.

Participants generally followed the prescribed protocol. Deviations were minor and were

considered unlikely to have a significant impact on the study outcome.

All the laboratories detected concentration-related increases in revertant numbers in each

strain that were sufficient to be considered mutagenic responses by the assessment criteria

applied by each laboratory. Based on the results of the common statistical analysis, eleven of

twelve labs were capable of differentiating the response of Canadian Monitor 8 from that of

Kentucky Reference 3R4F (KR 3R4F > Canadian Monitor 8) using TA98 with S9 activation;

six labs showed a significant difference with TA100 +S9 (KR 3R4F > Canadian Monitor 8).

2. Introduction

At the 2007 meeting of the CORESTA In Vitro Toxicity Testing Task Force (Jeju, 2007) the

Task Force confirmed its intention to conduct an Ames Assay Proficiency study with the

following purpose:

To conduct a proficiency testing programme to evaluate cigarette smoke particulate

matter using a common experimental design for Ames assay. This will be achieved by

evaluating the potential mutagenic activity of two smoke condensates by examining its

ability to revert two histidine requiring strains of Salmonella typhimurium (TA98 and

TA100) in the absence and presence of a rat liver metabolising (S-9) system.


The objective of this study was to evaluate the Ames assay ability to discriminate the

mutagenic potential of mainstream smoke particulate matter extracts from two cigarette

samples (cigarette smoke condensate, CSC) by investigating effects on histidine requiring

strains of Salmonella typhimurium in the absence and presence of a liver metabolising system

(Ames et al., 1975, OECD Test Guidelines number 471). The Ames assay is a rapid, reliable

and economical method for screening compounds for potential genetic activity at the

nucleotide level (Ames et al., 1975). When exposed to a mutagen, some of the bacteria in the

treated population undergo genetic changes which cause them to revert to a non-histidine

requiring state and thus grow in the absence of exogenous histidine. Different tester strains

are used because each is mutated by particular chemical classes of compound. A large

database has been accumulated with this assay, confirming its ability to detect genetically

active compounds of most chemical classes with around 80 to 90% sensitivity and specificity

(Gatehouse et al., 1990).

The following bacterial strains were used in this trial:

Strain Organism Type of Mutation

TA98 S. typhimurium frame-shift

TA100 S. typhimurium base-pair substitution

3. Participants

Table 1 - List of Participating Laboratories

Participants Study contribution

Altadis

Arista Laboratories Collecting and providing condensate

British American Tobacco (ATC) QA Review

Imperial Tobacco Canada Limited

Japan Tobacco Inc

KT&G Central Research Institute

Labstat International ULC Statistical Analysis

Lorillard Tobacco Company (A W Spears)

Ökolab/JTI

ITG/Reemtsma GmbH

RJ Reynolds

Zhengzhou Tobacco Research Institute of CNTC

During the course of this study, Eian Massey (BAT), Betsy Bombick (RJR) and Bernhard

Majer (Ökolab/JTI) served respectively as Co-coordinators of the Task Force. T.S. Kumaravel

served as initial Study Trial Coordinator, followed by Betsy Bombick.

4. Study Protocol

The protocol, summarized below, was reviewed and approved by a Study Manager from each

participating laboratory. The approved Study Plan is available upon request.


1. Samples:

- Kentucky Reference 3R4F (3R4F)

- Canadian Monitor 8 (CM8)

The cigarettes were provided by Arista Laboratories.

2. Cigarettes were conditioned and smoked under ISO conditions (ISO 3308).

3. Three independent smoking sessions were performed (3 separate days). Smoke

chemistry results are presented in table 2 below:

Table 2 - Smoke chemistry results (mean ± 1 standard deviation)

Code Sample

ID

Weight

(mg/cig)

Puff Count

(per cig)

TPM

(mg/cig)

Water

(mg/cig)

Nicotine

(mg/cig)

Tar

(mg/cig)

A CM8 967.0±3.3 8.3±0.1 14.7±0.5 0.64±0.17 1.02±0.03 13.01±0.45

C CM8 968.6±7.2 8.4±0.1 16.4±0.4 0.96 ±0.25 1.08±0.03 14.37±0.45

E CM8 969.9±5.5 8.4±0.1 16.1±0.4 2.28±0.46 1.04±0.03 12.82±0.49

B 3R4F 1043.0±5.2 8.7±0.1 11.0±0.5 0.24±0.12 0.71±0.04 10.09±0.45

D 3R4F 1047.1±6.0 8.8±0.2 11.4±0.7 0.99± 0.16 0.73±0.03 9.67±0.53

F 3R4F 1045.1±6.7 8.7±0.2 11.4±0.5 0.56± 0.18 0.73±0.04 10.05±0.47

4. Pads of each replicate were extracted to provide 10mg Nicotine Free Dry Particulate

Matter (NFDPM)/mL in DMSO.

5. Each participant received 2 aliquots of each condensate extract (4mL each), for a total of

12 vials (2 aliquots x triplicate smokings x 2 cigarettes).

6. Smoke condensates were coded as follows:

A: CM8 Replicate 1

B: 3R4F Replicate 1

C: CM8 Replicate 2

D: 3R4F Replicate 2

E: CM8 Replicate 3

F: 3R4F Replicate 3

7. Assays were conducted using TA98 and TA100 ± S9 using at least 6 non-zero test sample

concentrations and 3 plates per concentration. Concurrent negative and positive controls

were also assayed.

8. Work sheets were sent to each participating laboratory for recording experimental

information and data.

5. Results – Raw Data

Laboratories provided experimental information, raw data and summary data using the

provided worksheets. Summary results as provided by each individual laboratory are reported

in APPENDIX A.

Each participating laboratory used 5% metabolic activation; five laboratories also used 10%.


6. Statistical Analysis and Interpretations

Data analysis was performed with consideration to the following aspects of the study data:

• Reported dose ranges

• Solvent control data

• Average dose-response data

• Specific Activity: reported vs. determined using single methodology (statistician)

• Effect of S9 percentage (5% vs. 10%)

• Ability of individual labs to differentiate between the two test brands (CM8, 3R4F)

• Comparison of assay results among labs

• Determination of “outliers”

• Repeatability (r) and Reproducibility (R) estimates

Further details, including the presentation “Results and Statistical Analysis of Collaborative

Ames Assay Proficiency Data: In Vitro Toxicology Task Force, February 2009” given by

Wendy Wagstaff at the February 2009 Task Force meeting, are available upon request.

6.1 Reported Dose Ranges and Solvent Controls

The dose ranges reported as being used by the participating labs ranged from a low of 10µg

TPM (or NFDPM)/plate to a high of 1000 µg/plate, with the majority of labs reporting

maximum doses of between 250 and 500 µg/plate.

The dose range reported for Lab 12 could not be converted to units that were consistent with

other labs. Lab 12 was therefore excluded from any analysis in which data is compared

amongst laboratories.

The solvent control results were fairly consistent among labs, with all labs except one (Lab 5)

showing comparable abilities to reproduce solvent control results.

6.2 Specific Activity

6.2.1 Methodology

A common methodology for determining specific activity was applied to all results submitted

by participating labs as per Margolin et al., 1981 by fitting the quadratic model y=𝛽0 + 𝛽1𝑥 +𝛽2𝑥

2 to the tobacco smoke condensate sample dose X (e.g. mg NFDPM/plate) and revertant

colony count Y (revertant colonies/plate) for each individual replicate assay and removing the

highest dose data until the quadratic term was no longer significant (α = 0.05). In fitting the

quadratic model however, only doses contributing to downward curvature were considered for

exclusion.

Specific activity results determined using the above methodology for each individual

laboratory are reported in Appendix B.

6.2.2 Self-Reported vs. Determined Specific Activity

For the majority of labs, there were no large differences in self-reported specific activity

results and those determined by the statistician using the above methodology. For labs where

a large difference existed, there may have been issues with the methodology applied by the

individual lab or lack of clear units associated with the specific activity results reported.


6.3 Effect of S9 Percentage

6.3.1 Methodology

For each individual lab and test system, the difference in assay results between the two

different concentrations of S9 (5%, 10%) was evaluated by determining whether the linear

regression model coefficient β1 was significantly different between the two assays for each

test brand (CM8, 3R4F). Data for triplicate test samples was pooled for each test brand when

creating the linear regression models.

6.3.2 Results

Of the 5 labs that submitted data with the two different concentrations of S9, two labs (1 and

11) showed consistent results in which the 10% S9 assays showed lower levels of

mutagenicity (i.e. specific activity) than the 5% S9 assays. Two other labs (4 and 6)

demonstrated no significant effect of the S9 concentration while the third lab (12) appeared to

have mixed results. No specific conclusion regarding the impact of the concentration of S9

can be drawn from this data set since there is no overwhelming evidence either way.

Canadian Monitor #8 (CM8) Kentucky Reference 3R4F

Fig 1: Comparison of S9 Percentage (TA98 +S9, Mean Specific Activities ± 95% C.I.)


Fig 2: Comparison of S9 Percentage (TA100 +S9, Mean Specific Activities ± 95% C.I.)


6.4 Differentiating Test Brands

6.4.1 Methodology

For each individual lab and test system, the ability of a lab to differentiate the assay results of

the two test brands (CM8, 3R4F) was evaluated by determining whether the linear regression

model coefficient β1 was significantly different between the two test brands. Data for

triplicate test samples was pooled for each test brand when creating the linear regression

models.

6.4.2 Results

Eleven of twelve laboratories determined that the mutagenicity of Kentucky Reference 3R4F

was significantly greater than that of Canadian Monitor 8 in the TA98 +S9 test system. Only

six of eleven laboratories were able to differentiate between 3R4F and CM8 in the TA100

+S9 test system, also finding 3R4F more mutagenic than CM8. No predominant patterns, in

regards to specific activity differentiation, were observed in assays without S9 activation with

either TA98 or TA100.

Fig 3: Differentiating 3R4F and CM8: TA98 +S9 (5%) [Mean Specific Activities ± 95% C.I.]

Fig 4: Differentiating 3R4F and CM8: TA100 +S9 (5%) [Mean Specific Activities ± 95% C.I.]


Fig 5: Differentiating 3R4F and CM8: TA98 (-S9) [Mean Specific Activities ± 95% C.I.]

Fig 6: Differentiating 3R4F and CM8: TA100 (-S9) [Mean Specific Activities ± 95% C.I.]

6.5 Comparison among Labs

6.5.1 Methodology

In comparing specific activities among laboratories for the same test brand (CM8, 3R4F) and

test system, analysis of variance (ANOVA) of the specific activity estimates among

laboratories (3 replicate specific activity estimates per lab for each brand in the form of

samples {A, C, E} and {B, D, F}) was applied to determine whether the mean specific

activity estimate is the same for all labs. (The dose ranges reported for Lab 12 could not be

converted to units that were consistent with other labs, thus Lab 12 was excluded from all

analysis presented in these subsections.) In cases where the ANOVA was significant (α =

0.05), paired comparisons were made using the Student-Newman-Keuls (S-N-K) multiple

range test.


6.5.2 Results

With the Student-Newman-Keuls (S-N-K) multiple range test, significantly positive pairs

were connected by green lines in the following tables.


Fig 7: S-N-K Multiple Range Test: TA98 +S9 (5%) [Mean Specific Activities ± 95% C.I.]


Fig 8: S-N-K Multiple Range Test: TA100 +S9 (5%) Mean Specific Activities ± 95% C.I.


Fig 9: S-N-K Multiple Range Test: TA98 (-S9) [Mean Specific Activities ± 95% C.I.]



Fig 10: S-N-K Multiple Range Test: TA100 -S9 Mean Specific Activities ± 95% C.I.

6.5.3 Multiple Range Test ‘Outliers’

In instances where the ANOVA was significant and the S-N-K multiple range test identified a

single lab whose mean specific activity results were significantly different from those of any

other lab, that lab was determined to be an ‘outlier’ for the test brand and test system. All lab

results determined to be ‘outliers’ are listed in the table below.

Table 3 – ‘Outliers’

Test system Sample ID Outlying

Lab

Specific activity

(rev/mg NFDPM) Range of other labs

TA98 +5% S9 3R4F 6 4757 1998 - 3520

CM8 6 2226 1263 - 1862

TA100 +5% S9 3R4F 1 1515 518 - 1086

CM8 none - -

TA98 -S9 3R4F none - -

CM8 none - -

TA100 -S9 3R4F none - -

CM8 none - -

6.5.4 Specific Activity Repeatability and Reproducibility Estimates

Following the removal of the ‘outlying’ lab data identified in the above table, the replicate

specific activity determinations from all remaining labs were pooled to determine estimates of

repeatability (r) and reproducibility (R) for each combination of test brand (CM8, 3R4F) and

test system, which are displayed in the table below.


Table 4 - Specific Activity Repeatability and Reproducibility

Specific Activity (rev/mg NFDPM)

Test

System

Sample

ID

Number of

Labs

Grand

Mean

Repeatability Reproducibility

r C.V. R C.V.

TA98 +S9 (5%) 3R4F 10 2794 586 11% 1148 21%

CM8 10 1487 262 9% 474 16%

TA100 +S9 (5%) 3R4F 9 884 298 17% 495 29%

CM8 10 678 244 18% 423 32%

TA98 -S9 3R4F 11 89 88 51% 132 76%

CM8 11 72 107 76% 138 98%

TA100 -S9 3R4F 10 362 246 35% 472 67%

CM8 10 344 373 55% 431 64%

7. Discussion of Results

7.1 Data Audit

Study data was audited for

Evidence of following the agreed Study Plan

Evidence of “fair testing”

Accuracy of the reported results

7.2 Audit Findings

Some differences in methodology were observed. These differences included technical

details (i.e., concentrations) and unknown analytical details. The specific impact of these

differences is unknown, but the majority of the audit findings were minor and considered

unlikely to impact the final results overall. Further details of the audit findings are available

upon request.

8. Conclusions

Participants generally followed the overall guidance prescribed in the study protocol.

However, some laboratories did not submit requested information or full detailed worksheets,

and the Study Auditor found clear room for improvement in documentation procedures.

Eleven of twelve laboratories were able to differentiate the response of CM8 from that of

3R4F (3R4F > CM8) in the TA98 +S9 test system. The differences were much less

pronounced, and in many cases insignificant, in all other evaluated test systems (e.g. TA100

+S9 or TA98/TA100 -S9).

This Proficiency Trial was designed to:


Evaluate the mutagenicity of two cigarettes using a common study protocol

Enable laboratories to evaluate method proficiency

Enable laboratories to evaluate their documentation procedures via an independent limited

audit of study documents

The goals of the proficiency study were accomplished. The Ames assay is sufficiently

sensitive to distinguish significant differences between the two test cigarettes in at least one of

the applied test systems (TA98 +S9), and each laboratory benefitted from comparing their

results/methodology with that of the other participating laboratories.

9. References

1. Ames, B.N., McCann, J. & Yamasaki, E. (1975) Methods for detecting carcinogens and

mutagens with Salmonella/mammalian-microsome mutagenicity test. Mutation Res. 31,

347-364.

2. Gatehouse, D.G., Wilcox, P., Forster, R., Rowland, I. & Callender, R.D. (1990) Bacterial

mutation assays. In "Basic Mutagenicity Tests. UKEMS Recommended Procedures."

Report of the UKEMS Sub-committee on Guidelines for Mutagenicity Testing. Ed D.J.

Kirkland. Cambridge University Press.

3. OECD Guideline for testing of chemicals, Bacterial Reverse Mutation Test, number 471

(adopted 21 July 1997).


Appendix A – Raw Data

A-1 Raw Revertant Colony Count Data

A-1.1 Sample A (TA98)

Participating CSC TA98 (+S9) with 5% S9 TA98 (-S9) TA98 (+S9) with 10% S9

Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

1

0 19 18 17

13 13 8

15 22 17

10 32 27 30

20 15 9

17 21 30

25 48 59 57

19 32 19

33 30 34

50 71 87 93

21 29 21

54 58 56

75 144 113 137

20 19 20

69 73 82

125 164 178 166

18 16 20

143 155 117

250 309 259 276

39 (C) 30 43

247 247 261

2

0 30 32

30 32

50 118 91 141

4 18 24

100 218 182 229

20 0 26

150 227 290 248

3 0

200 257 298 295

5

21

250 243 259 285

19 27 16

300 285 270 299

15 14 17

3

0 138 112 81

99 134 122

25 91 113 106

101 116 143

50 135 129 104

71 112 109

75 169 180 131

129 143 152

100 202 177 180

127 81 80

125 217 235 227

91 87 88

250 306 303 284

175 134 138

500 302 292 256

106 165 121

4

0 53 45 40 50

43 38 36 38

43 49 40 41

50 148 125 109 136

35 31 47 31

99 112 107 99

100 184 206 212 234

56 113 (c) 39 48

149 137 157 160

200 342 314 280 232

77 85 81 113

316 284 316 265

300 463 471 426 458

44 93 103 84

550 398 501 405

500 416 334 396 C

158 145 141 164

617 458 436 413

1000 301 229 221 256

150 136 190 168

385 417 329 305

5

0 394 379 391

26 23 42

50 497 439 454

34 32 28

100 613 501 497

32 24 29

150 570 635 612

43 36 33

200 766 686 664

30 48 58

250 809 738 696

40 54 42

300 (541)* (672)* 765

22 33 36

6

0 27 27 30

16 20 23

31 33 29

25 69 81 76

22 28 19

63 63 58

50 154 142 151

22 28 18

116 92 111

75 185 199 228

20 23 36

189 198 162

100 274 249 281

23 26 23

241 229 234

150 313 339 336

43 30 25

375 354 342

200 363 357 358

49 46 49

506 461 507

400 336 305 327

tox tox tox

527 578 623

800

tox tox tox

7

0 34 31 30

19 16 13

25 49 51 56

26 20 33

50 95 80 71

25 23 29

75 104 133 99

24 23 18

100 124 160 150

26 20 24

125 168 174 168

38 16 25

200 264 206 239

31 36 33

8

0 27 29 40

19 23 20

25 43 40 37

21 20 23

50 84 79 67

19 28 25

75 123 120 129

24 27 29

100 160 167 145

18 23 20

125 196 180 204

22 24 36

250 244 243 232

32 37 32

500 367 348 349

62 75 69

9

0 33 22 35 24 34

35 33 22 24 29

1000 168 159 200

tox tox tox

500 289 236 267

66 69 51



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

250 299 265 325

44 46 36

150 272 265 272

35 48 51

100 192 166 189

42 30 26

50 120 119 116

36 35 33

25 86 61 71

21 37 27

10

0 73 51 51

28 36 38

25 86 61 71

21 37 27

100 192 166 189

42 30 26

150 272 265 272

35 48 51

250 299 265 325

44 46 36

500 289 236 267

66 69 51

1000 168 159 200

tox tox tox

11

0 18 27 24 39 37 22 15 11 9

28 35 18

25 64 53 71 52 70 48 23 25 25

34 42 51

50 82 117 103 84 84 68 18 19 22

48 58 56

75 99 108 118 99 114 99 43 31 31

66 82 85

100 169 140 180 125 144 118 18 42 36

84 114 104

125 154 178 188 166 195 136 24 42 38

103 128 133

250 252 242 255 206 229 252 50 56 45

248 218 250

12

0 16 27 19

8 7 15

13 22 10

156 16 12 8

23 11 15

19 26 18

313 26 24 27

27 19 32

22 47 68

625 46 45 40

56 87 61

63 76 73

1250 41 42 47

88 73 89

93 96 83

2500 75 67 72

56 74 66

187 174 127

5000 80 67 72

57 60 98

152 168 114

c: Contamination, tox: Growth Inhibition, *: Plate contaminated or plate skewed)

A-1.2 Sample A (TA100)


Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

1

0 98 99 106

100 96 91

91 92 100

10 107 125 117

110 100 141

111 118 112

25 120 117 125

135 153 99

111 96 78

50 156 113 136

139 152 142

135 135 112

75 188 180 165

144 100 113

167 123 112

125 215 200 194

161 135 170

169 167 154

250 247 310 246

120 144 148

273 234 204

2

0

50

100

150

200

250

300

3

0 200 147 162

159 166 164

25 196 245 188

152 164 285

50 180 188 213

167 168 174

75 241 245 213

182 196 205

100 197 244 246

199 179 205

125 231 221 207

183 179 155

250 261 260 351

164 160 130

500 323 333 350

144 144 95

4

0 173 175 197 184

171 160 163 214

196 194 191 196

50 213 200 217 221

218 213 190 226

178 192 192 224

100 240 210 232 220

236 218 226 230

212 182 196 212

200 237 230 269 278

265 286 313 274

261 234 221 236

300 241 242 274 240

248 173 257 273

278 249 272 290

500 229 226 252 257

340 312 318 352

C 269 257 262

1000 266 269 269 250

349 354 417 449

C 286 318 293

5

0 465 459 453

127 97 102

50 498 489 511

106 97 128

100 511 491 525

116 114 121

150 595 553 552

136 104 131

200 562 600 620

115 109 123

250 642 619 599

130 151 153

300 671 603 577

171 150 139

6 0 92 111 90

63 77 81

93 93 75



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

25 134 129 105

87 96 108

118 112 114

50 128 133 123

122 97 108

122 128 131

75 154 115 161

126 115 108

160 166 131

100 147 180 175

120 111 113

144 168 167

150 203 180 C

142 133 129

197 174 171

200 203 220 182

175 146 150

203 211 221

400 201 244 225

211 214 241

251 255 269

800

tox tox tox

7

0 73 103 83

89 81 84

25 90 78 98

108 91 90

50 80 104 80

109 76 106

75 111 110 121

99 104 110

100 90 95 108

95 98 106

125 133 149 135

103 104 96

200 150 188 154

99 111 111

8

0 123 107 117

125 131 132

25 139 144 137

136 128 139

50 141 157 130

128 135 137

75 176 159 157

165 167 164

100 179 181 180

148 135 154

125 204 173 190

171 161 157

250 201 237 208

168 172 188

500 292 277 315

100 120 112

9

0 127 136 154 137 157

106 101 112 118 101

1000 368 334 374

tox tox tox

500 330 361 353

212 189 203

250 241 283 256

247 234 215

150 258 274 288

188 201 175

100 227 225 218

179 159 140

50 192 194 193

138 102 118

25 161 170 157

137 126 120

10

0 144 125 149

112 123 140

50 203 175 186

162 172 140

100 209 224 221

168 173 164

150 270 253 231

199 197 198

200 285 243 263

221 177 202

250 167 264 286

203 203 215

300 325 301 313

221 203 179

11

0 145 134 118 154 113 135 143 110 125

107 122 133

25 141 134 131 145 * 149 157 161 157

125 141 143

50 131 169 147 177 182 177 171 184 168

147 169 136

75 166 163 147 179 178 200 169 182 156

148 150 155

100 178 152 198 223 197 217 175 171 183

170 169 203

125 189 197 193 211 226 216 194 229 180

207 212 168

250 223 209 207 291 252 281 151 169 189

225 239 243

12

0 75 56 77

41 39 51

66 70 53

156 50 80 70

79 64 87

85 83 71

313 63 72 88

76 80 93

60 97 69

625 62 106 99

99 85 79

75 77 64

1250 52 48 42

77 83 82

44 85 134

2500 51 59 73

113 106 93

85 71 63

5000 72 63 102

84 78 73

76 91 88

(c: Contamination, tox: Growth Inhibition, *: Plate contaminated or plate skewed)

A-1.3 Sample B (TA98)


Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

1

0 19 18 17 13 13 8 15 22 17

10 45 46 29 19 18 5 33 29 33

25 67 99 70 16 22 28 43 45 56

50 168 191 156 19 14 19 94 85 91

75 278 273 257 30 26 28 126 156 157

125 406 380 354 29 26 27 314 351 277

250 511 570 595 58 41 52 512 526 637

2

0 30 32 M 30 32 M

50 157 170 197 0 19 27

100 312 309 314 6 4 M

150 370 399 417 8 M 16

200 438 440 413 15 11 10



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

250 477 509 463 18 15 17

300 437 42 458 12 19 20

3

0 106 76 105 121 68 91

25 237 168 177 95 108 78

50 175 138 259 83 113 126

75 253 231 244 91 105 99

100 293 259 365 138 102 115

125 316 323 289 83 162 112

250 405 428 386 96 161 98

500 432 427 376 105 125 112

4

0 53 45 40 50 43 38 36 38 43 49 40 41

50 210 178 202 189 36 33 32 35 143 178 141 174

100 281 284 284 369 49 59 56 56 316 265 308 270

200 458 493 554 438 63 72 85 127 506 514 537 424

300 725 770 645 786 83 75 77 83 743 738 816 756

500 650 739 538 599 87 127 127 117 974 1032 1022 990

1000 449 362 222 210 133 135 161 188 887 1018 718 554

5

0 48 31 41 35 33 30

50 177 208 177 26 31 39

100 424 415 385 22 24 30

150 503 507 528 25 35 23

200 586 623 593 34 39 40

250 654 717 700 30 34 44

300 779 762 760 37 49 (0)*

6

0 27 27 30 16 20 23 31 33 29

25 102 120 127 22 21 22 103 93 84

50 236 243 273 28 30 31 216 196 228

75 341 360 411 29 25 24 318 306 330

100 494 475 480 26 40 26 454 431 431

150 568 623 651 39 33 38 685 683 720

200 608 612 699 32 40 38 897 886 893

400 514 573 585 62 tox 67 956 1001 1029

800 tox tox tox

7

0 34 31 30 19 16 13

25 73 88 66 18 18 19

50 129 128 145 24 19 43

75 216 190 199 25 36 30

100 261 250 255 29 30 28

125 308 360 325 50 35 30

200 333 340 366 48 44 48

8

0 20 29 22 23 22 17

25 53 56 51 18 20 25

50 163 156 146 20 25 23

75 163 175 165 27 21 21

100 273 268 266 27 28 19

125 281 271 269 21 23 21

250 303 273 278 18 25 32

500 476 435 423 62 60 56

9

0 33 22 35 24 34 35 33 22 24 29

1000 385 426 445 tox tox tox

500 475 576 577 82 99 98

250 513 460 520 58 36 53

150 354 393 330 34 70 47

100 271 300 321 30 34 27

50 238 222 237 28 37 33

25 152 127 127 36 34 22

10

0 73 51 51 28 36 38

30 105 117 85 30 36 40

60 207 184 201 51 27 48

90 290 332 276 40 30 41

120 432 421 470 47 43 56

150 470 505 482 31 35 51

180 636 537 641 34 26 29

11

0 29 19 24 20 33 32 18 20 15 23 33 19

25 107 93 101 84 61 66 18 23 10 57 43 60

50 163 138 125 106 138 162 34 23 28 70 104 103

75 223 196 241 193 208 187 28 33 33 152 123 162

100 255 252 283 250 252 223 29 27 39 167 196 198

125 334 327 326 305 260 275 33 42 47 214 223 215

250 394 408 404 336 423 433 64 61 42 372 418 445



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

12

0 16 27 19 8 7 15 13 22 10

156 48 53 41 16 12 15 34 39 45

313 72 52 64 23 37 45 56 38 63

625 116 125 117 45 28 56 171 123 109

1250 141 153 129 78 81 42 145 153 164

2500 98 81 103 74 69 91 285 236 248

5000 94 110 115 43 78 56 228 279 253

(c: Contamination, tox: Growth Inhibition, *: Plate contaminated or plate skewed, M: Missing

A-1.4 Sample B (TA100)


Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

1

0 98 99 106

100 96 91

91 92 100

10 124 112 124

125 119 121

82 98 81

25 137 143 151

140 124 137

120 97 86

50 165 167 181

152 136 153

137 122 123

75 192 181 174

168 155 164

153 139 151

125 256 286 259

141 152 154

208 207 195

250 342 421 335

155 152 180

326 304 325

2

0

50

100

150

200

250

300

3

0 184 155 188

188 178 195

25 213 198 213

191 189 164

50 248 252 250

205 199 188

75 276 290 239

221 198 232

100 250 290 268

215 213 213

125 263 262 253

229 199 225

250 297 373 364

181 201 152

500 452 440 414

161 157 124

4

0 173 175 197 184

171 160 163 214

196 194 191 196

50 164 232 232 194

170 165 181 161

197 185 225 186

100 214 242 230 222

170 189 172 198

212 216 232 262

200 281 252 278 253

220 236 230 242

206 262 258 278

300 248 254 253 272

210 222 218 213

297 280 309 348

500 270 264 246 242

221 256 242 244

310 317 286 322

1000 254 188 234 222

325 361 338 384

340 349 309 286

5

0 94 98 95

102 94 99

50 147 156 138

118 104 115

100 266 235 231

110 115 109

150 283 271 228

125 102 126

200 312 311 306

127 117 141

250 321 329 323

125 137 133

300 356 345 358

138 142 117

6

0 92 111 90

63 77 81

93 93 75

25 125 110 145

95 105 92

142 121 142

50 151 163 167

103 116 107

131 141 138

75 167 184 176

136 114 125

171 186 171

100 215 206 191

124 140 142

193 208 192

150 228 223 170

145 122 151

261 237 247

200 247 228 224

142 133 142

326 269 272

400 224 232 233

211 227 220

329 306 312

800

tox tox tox

7

0 73 103 83

89 81 84

25 86 103 90

124 79 90

50 121 123 96

96 108 85

75 123 118 104

110 80 110

100 126 125 136

95 111 115

125 140 143 155

105 105 90

200 159 205 175

113 110 91

8

0 111 112 132

119 126 111

25 155 136 137

127 137 128

50 153 147 169

151 144 149

75 192 181 209

156 159 161

100 225 193 197

148 154 167



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

125 224 230 219

149 148 160

250 292 295 331

172 165 161

500 324 310 332

148 133 152

9

0 127 136 154 137 157

106 101 112 118 101

1000 423 38 368

tox tox tox

500 457 460 460

144 279 143

250 316 304 343

211 207 179

150 279 259 243

206 188 183

100 263 267 253

162 134 116

50 192 217 181

145 159 136

25 149 210 174

158 161 118

10

0 144 125 149

112 123 140

50 223 209 190

150 143 165

100 252 250 245

145 152 139

150 302 296 301

176 184 164

200 365 323 296

205 176 193

250 361 333 383

171 170 189

300 370 394 381

186 218 157

11

0 124 120 132 145 135 139 133 138 146

159 * 139

25 142 120 169 167 139 182 152 159 133

131 150 142

50 163 163 168 202 191 207 163 156 212

127 175 148

75 183 200 191 200 229 187 154 146 155

175 175 137

100 253 214 229 206 226 233 192 178 155

186 205 226

125 239 253 201 244 244 295 201 219 197

193 203 186

250 206 280 247 322 296 279 177 163 166

308 265 305

12

0 75 56 77

41 39 51

66 70 53

156 85 93 87

50 24 32

88 90 72

313 97 122 131

41 50 40

128 135 119

625 120 116 84

38 57 31

148 156 113

1250 117 134 105

50 56 47

167 174 149

2500 121 148 167

79 65 59

185 165 195

5000 104 108 151

81 79 95

218 222 167

(tox: Growth Inhibition, *: Plate contaminated or plate skewed)

A-1.5 Sample C (TA98)


Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

1

0 19 18 17

13 13 8

15 22 17

10 32 35 33

9 18 18

26 39 28

25 54 47 34

22 16 19

22 34 31

50 95 72 80

26 15 18

71 53 42

75 135 106 110

26 18 28

70 86 61

125 170 141 209

29 28 21

112 111 125

250 275 216 244

27 29 32

259 229 286

2

30 32 M

30 32 M

50 103 83 102

M 29 20

100 177 165 161

11 M 18

150 197 252 241

18 12 24

200 248 257 277

12 14 26

250 277 263 322

13 13 23

300 272 315 294

22 13 17

3

0 105 112 111

83 93 68

25 180 132 148

99 96 84

50 173 156 144

111 111 89

75 176 225 176

115 80 138

100 259 254 279

134 111 124

125 227 263 257

90 86 118

250 393 310 334

105 105 148

500 308 358 320

131 164 96

4

0 49 53 54 58

39 39 28 36

35 49 47 50

50 116 121 157 116

29 35 40 36

111 100 85 96

100 294 258 265 185

28 17 29 37

178 172 149 197

200 497 404 389 393

39 49 31 31

378 317 208 334

300 583 325 384 426

49 60 72 32

169 362 406 495

500 438 469 433 414

49 45 53 47

506 639 579 529

1000 317 386 450 446

75 75 89 79

730 698 811 683

5 0 153 151 174

153 173 165

50 209 216 184

174 163 146



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

100 150 323 306

141 139 161

150 374 318 314

163 120 115

200 463 413 449

136 154 134

250 440 497 520

136 133 136

300 523 464 559

161 136 155

6

0 27 27 30

16 20 23

31 33 29

25 80 79 75

24 27 20

60 74 57

50 133 114 138

21 30 31

114 119 109

75 227 188 175

21 35 29

173 164 173

100 238 224 256

32 37 28

216 219 223

150 368 345 328

33 30 36

321 379 C

200 378 348 383

38 37 34

457 452 459

400 325 307 321

tox tox tox

578 549 572

800

tox tox tox

7

0 25 16 21

20 10 5

25 34 44 36

19 26 28

50 56 65 69

14 40 28

75 88 80 106

29 41 34

100 95 104 106

24 38 43

125 139 128 149

41 58 46

200 178 163 174

51 46 50

8

0 24 25 25

21 24 25

25 55 51 53

24 19 22

50 87 85 97

21 23 24

75 113 123 116

27 21 23

100 148 147 143

22 21 21

125 159 159 163

20 31 27

250 243 256 236

22 20 32

500 275 264 263

46 50 48

9

0 33 22 35 24 34

35 33 22 24 29

1000 206 248 206

tox tox tox

500 286 237 286

54 43 51

250 219 220 219

31 30 38

150 256 265 259

48 38 29

100 157 172 208

35 45 28

50 108 112 125

43 30 27

25 73 63 65

34 36 25

10

0 63 60 56

20 26 31

30 85 99 86

22 41 29

60 117 119 96

29 42 24

90 190 167 188

56 27 34

120 235 215 256

20 28 15

150 242 241 301

19 23 24

180 324 360 377

14 15 17

11

0 24 27 11 23 29 24 14 17 23

27 * 33

25 45 46 53 51 49 70 22 27 27

32 29 38

50 96 75 84 89 84 82 24 27 39

73 39 60

75 108 106 120 99 109 99 29 18 31

82 84 73

100 148 155 111 119 120 115 43 33 37

103 99 89

125 174 143 171 151 166 161 31 28 45

103 123 134

250 265 222 236 228 189 205 55 58 64

* 215 233

12

0 18 22 23

9 11 19

15 23 13

156 21 25 20

13 14 18

9 31 14

313 38 39 38

22 19 28

41 25 37

625 42 39 41

18 24 24

31 35 37

1250 47 28 52

40 37 71

78 58 81

2500 60 63 78

59 36 71

183 164 152

5000 74 86 108

64 100 98

95 126 117

(c: Contamination, tox: Growth Inhibition, *: Plate contaminated or plate skewed, M: Missing)

A-1.6 Sample C (TA100)


Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

1

0 98 99 106

100 96 91

91 92 100

10 122 117 139

105 109 94

86 97 106

25 126 139 109

136 110 135

122 99 97

50 167 152 155

139 113 150

99 113 79

75 180 174 168

163 161 157

175 124 126

125 192 188 216

163 141 192

141 161 174



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

250 283 278 284

165 142 166

228 219 202

2

0

50

100

150

200

250

300

3

0 189 200 177

197 186 190

25 231 217 203

196 147 211

50 244 227 180

217 241 224

75 283 273 274

221 207 240

100 274 264 281

161 204 217

125 300 259 243

174 197 204

250 334 306 377

177 183 209

500 420 281 356

191 166 129

4

0 222 183 193 187

144 155 143 146

165 180 202 204

50 225 224 185 246

205 169 177 192

185 238 232 232

100 178 209 238 244

193 181 188 158

201 237 246 237

200 264 222 270 288

157 145 189 170

274 241 306 252

300 276 293 296 244

196 189 220 204

258 260 254 294

500 296 320 324 254

201 197 182 236

308 354 328 321

1000 274 328 318 269

289 234 266 280

360 369 340 330

5

0 235 195 209

228 215 193

50 211 203 256

203 196 230

100 270 241 239

226 220 233

150 264 287 268

232 211 222

200 284 295 312

221 218 229

250 316 306 288

227 214 184

300 345 312 345

242 220 221

6

0 92 111 90

63 77 81

93 93 75

25 106 121 139

98 119 114

104 113 110

50 159 136 134

120 83 105

129 123 137

75 166 148 131

102 95 120

125 158 159

100 192 180 192

137 124 108

177 142 150

150 189 193 184

125 130 172

191 185 203

200 181 201 194

146 132 156

244 208 191

400 206 220 189

218 215 201

320 293 290

800

tox tox tox

7

0 65 79 80

73 55 69

25 71 86 68

83 89 84

50 84 89 71

130 105 118

75 101 108 110

130 128 96

100 99 116 116

103 114 144

125 129 119 131

120 139 114

200 155 188 136

121 114 114

8

0 115 117 128

109 101 97

25 112 110 135

112 105 100

50 120 123 136

133 119 117

75 161 163 179

163 149 153

100 199 197 190

153 161 155

125 198 190 209

149 157 160

250 279 275 260

154 160 172

500 284 280 280

115 112 124

9

0 127 136 154 137 157

106 101 112 118 101

1000 385 375 394

tox tox tox

500 367 370 386

202 202 180

250 284 297 258

232 173 177

150 267 272 244

157 193 175

100 219 237 225

181 173 158

50 198 191 195

153 154 121

25 131 177 164

107 154 148

10

0 156 157 148

140 115 130

50 191 211 184

170 177 204

100 252 244 201

182 208 197

150 270 260 253

231 216 247

200 308 292 321

241 238 208

250 344 324 312

294 254 244

300 370 357 334

262 255 262

11 0 138 125 122 140 111 * 120 127 160

138 146 115



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

25 133 147 147 137 156 156 164 111 127

159 127 150

50 145 143 157 170 165 183 147 151 171

119 127 129

75 207 191 200 220 158 167 187 170 184

168 148 154

100 200 188 188 191 159 169 215 173 183

194 161 183

125 193 209 177 229 224 186 218 218 154

177 197 160

250 230 266 244 249 265 242 161 150 161

283 246 241

12

0 59 81 66

43 40 38

67 73 49

156 76 63 72

58 54 60

57 74 81

313 79 73 88

67 61 43

83 69 97

625 91 88 102

79 67 47

73 76 80

1250 98 96 95

76 71 66

88 86 77

2500 83 94 121

73 71 70

111 103 86

5000 116 86 85

74 79 85

108 117 121

(tox: Growth Inhibition, *: Plate contaminated or plate skewed, M: Missing)

A-1.7 Sample D (TA98)


Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

1

0 19 18 17

13 13 8

15 22 17

10 33 48 42

8 9 8

22 20 26

25 84 84 71

17 27 19

43 47 48

50 190 143 174

18 29 16

120 82 92

75 289 252 242

33 20 26

154 169 196

125 392 425 425

29 34 18

288 272 256

250 623 524 434

39 43 28

529 459 584

2

30 32 M

30 32 M

50 161 171 180

M M 4

100 304 315 395

9 M M

150 370 359 359

19 15 10

200 432 392 507

10 29 18

250 422 414 453

19 18 15

300 372 457 448

19 12 19

3

0 48 37 35

42 35 29

25 119 135 143

65 46 57

50 155 201 157

77 51 57

75 216 249 217

78 36 52

100 272 236 252

46 64 62

125 301 296 200

62 77 59

250 304 293 168

74 68 74

500 313 374 332

64 65 67

4

0 49 53 54 58

39 39 28 36

35 49 47 50

50 206 293 174 176

39 47 41 41

198 160 136 166

100 356 329 297 336

44 43 41 63

388 364 368 401

200 683 547 429 318

53 47 51 43

711 625 735 325

300 976 819 654 951

75 76 71 76

819 842 746 601

500 814 503 546 509

75 73 76 87

803 1003 1201 855

1000 661 521 501 C

115 105 108 91

1137 903 806 M

5

0 29 34 31

38 31 23

50 130 116 170

44 31 28

100 365 269 400

35 35 33

150 502 412 452

29 38 37

200 561 610 578

37 36 31

250 682 598 640

70 31 27

300 619 681 733

42 33 34

6

0 27 27 30

16 20 23

31 33 29

25 113 129 119

25 19 27

79 92 81

50 275 255 247

23 33 21

187 181 200

75 376 347 443

31 28 25

302 318 334

100 522 509 505

27 20 25

450 463 442

150 605 593 605

34 42 39

709 707 742

200 622 602 668

37 35 40

840 843 835

400 542 594 579

80 75 63

998 984 1029

800

tox tox tox

7

0 25 16 21

20 10 5

25 51 59 50

21 20 26

50 98 111 115

25 29 23

75 140 148 166

26 41 45

100 160 200 186

38 30 45

125 234 169 215

34 39 50



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

200 273 279 236

49 53 59

8

0 36 35 33

19 22 21

25 67 55 64

21 20 25

50 144 152 149

20 25 21

75 199 209 221

24 21 19

100 276 275 283

25 29 20

125 267 291 288

20 27 27

250 373 392 372

27 28 35

500 377 431 373

65 73 85

9

0 33 22 35 24 34

35 33 22 24 29

1000 449 447 492

tox tox tox

500 517 509 476

55 48 37

250 451 528 537

49 49 49

150 460 420 430

40 35 42

100 328 318 348

26 33 26

50 240 177 172

29 24 31

25 109 104 102

33 24 31

10

0 63 60 56

20 26 31

30 111 113 125

30 41 51

60 179 198 174

36 26 47

90 327 285 296

43 39 36

120 435 414 442

22 25 26

150 510 517 498

25 24 24

180 593 626 575

21 19 21

11

0 20 15 28 25 27 28 24 22 27

27 * 23

25 92 82 84 75 77 67 20 22 28

43 49 35

50 132 173 177 132 165 128 19 33 20

80 86 92

75 192 252 222 186 184 191 22 28 31

111 138 109

100 211 248 253 218 234 231 36 29 32

165 181 191

125 350 308 349 252 281 246 34 32 39

233 181 195

250 408 576 507 342 329 386 48 69 43

436 421 417

12

0 18 22 23

9 11 19

15 23 13

156 35 35 28

15 19 31

40 38 48

313 25 69 50

25 19 42

48 53 40

625 67 46 67

33 23 45

57 51 41

1250 82 46 62

32 47 33

95 146 100

2500 108 100 91

36 35 54

64 99 60

5000 86 80 96

44 53 77

162 54 35

(c: Contamination, tox: Growth Inhibition, *: Plate contaminated or plate skewed, M: Missing

A-1.8 Sample D (TA100)


Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

1

0 98 99 106

100 96 91

91 92 100

10 106 113 135

112 140 138

100 93 109

25 120 135 148

120 154 149

92 124 84

50 157 164 174

125 143 108

155 142 136

75 195 205 200

156 141 165

149 153 169

125 270 277 285

149 162 154

205 217 209

250 364 365 376

162 183 181

318 331 296

2

0

50

100

150

200

250

300

3

0 135 136 123

116 113 111

25 175 209 218

193 159 181

50 228 181 196

199 190 218

75 221 215 235

189 177 184

100 206 247 331

237 199 199

125 230 216 241

181 202 224

250 268 269 254

203 202 164

500 306 339 338

117 130 135

4

0 222 183 193 187

144 155 143 146

165 180 202 204

50 201 241 241 241

182 160 192 160

248 218 214 221

100 270 273 264 269

161 166 204 164

226 204 245 213



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

200 304 288 280 301

172 156 173 162

281 276 256 252

300 340 317 317 214

174 176 181 194

318 262 365 370

500 324 301 341 228

192 176 189 182

420 368 410 384

1000 336 296 340 358

209 218 229 244

413 378 374 366

5

0 80 114 84

97 94 80

50 126 113 134

101 114 114

100 185 158 206

118 120 130

150 198 139 212

116 131 85

200 229 209 207

135 109 96

250 243 224 305

122 143 118

300 280 279 285

125 106 129

6

0 92 111 90

63 77 81

93 93 75

25 111 123 129

114 92 83

124 115 116

50 189 160 171

93 110 107

153 153 149

75 169 170 167

96 97 115

189 196 196

100 197 193 189

118 118 103

206 211 164

150 222 190 224

135 131 119

236 225 228

200 217 223 241

124 138 127

269 249 275

400 247 225 247

177 206 186

321 351 312

800

tox tox tox

7

0 65 79 80

73 55 69

25 61 68 88

94 95 89

50 88 93 99

113 91 100

75 105 98 130

111 90 130

100 126 123 113

136 130 130

125 143 141 143

144 131 141

200 170 158 195

129 121 140

8

0 133 129 136

112 117 121

25 142 139 133

111 118 127

50 143 145 141

123 139 133

75 157 176 183

137 140 133

100 211 199 223

141 143 155

125 252 255 287

134 135 147

250 291 328 308

136 148 141

500 319 356 343

141 145 160

9

0 127 136 154 137 157

106 101 112 118 101

1000 414 453 455

tox tox tox

500 393 434 380

197 150 186

250 373 327 325

191 222 212

150 274 286 312

179 174 175

100 209 270 265

142 144 133

50 202 191 188

161 176 138

25 161 174 144

142 135 129

10

0 136 149 170

140 115 130

50 224 228 222

184 176 180

100 306 287 253

206 215 201

150 352 337 321

213 186 218

200 382 347 367

220 221 232

250 406 427 418

221 210 217

300 231 357 406

250 211 258

11

0 110 134 100 139 103 146 115 122 138

120 155 129

25 132 132 125 125 153 131 160 155 145

157 136 146

50 183 171 154 163 162 179 159 193 188

177 179 187

75 175 196 178 225 206 219 171 157 169

170 173 168

100 223 233 169 193 233 267 169 173 205

205 201 183

125 212 223 223 233 219 271 203 196 188

246 233 212

250 281 287 246 289 259 296 134 175 187

322 299 321

12

0 59 81 66

43 40 38

67 73 49

156 115 105 108

27 55 31

94 106 140

313 125 145 97

25 38 48

132 162 111

625 151 140 119

49 42 40

119 124 144

1250 109 113 112

80 84 112

137 98 114

2500 152 134 144

91 96 95

123 182 165

5000 108 113 135

111 96 102

185 219 195

(tox: Growth Inhibition)


A-1.9 Sample E (TA98)


Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

1

0 19 18 17

13 13 8

15 22 17

10 31 30 32

17 17 26

18 20 19

25 30 33 53

16 17 18

15 22 29

50 107 84 99

19 27 26

56 66 44

75 104 92 74

31 27 19

61 68 59

125 163 141 125

19 22 29

125 99 119

250 286 253 277

29 27 40

231 220 245

2

30 32 M

30 32 M

50 113 98 91

52 M 6

100 151 137 167

18 11 3

150 257 258 244

13 17 13

200 277 232 272

15 15 14

250 233 253 267

22 19 25

300 264 264 292

25 13 18

3

0 48 37 35

44 40 67

25 101 82 75

41 45 45

50 116 107 118

57 53 69

75 181 140 151

85 73 51

100 175 161 177

86 75 56

125 161 164 151

42 94 101

250 208 195 196

78 82 76

500 211 222 186

72 51 66

4

0 49 53 54 58

39 39 28 36

35 49 47 50

50 131 112 109 140

40 41 36 45

129 124 105 95

100 244 150 174 158

44 48 43 40

196 164 161 161

200 238 262 325 241

41 32 51 51

297 237 230 261

300 542 409 313 420

75 47 56 57

C 397 462 416

500 382 396 369 329

85 48 73 68

577 746 618 525

1000 365 349 336 269

93 117 91 107

788 847 776 834

5

0 40 34 36

25 44 38

50 93 80 83

32 38 29

100 154 147 105

40 23 43

150 231 172 183

35 29 30

200 275 281 240

39 48 36

250 345 271 306

33 31 46

300 350 311 340

46 54 57

6

0 27 27 30

16 20 23

31 33 29

25 74 66 67

28 18 20

59 56 59

50 146 123 118

18 25 25

102 67 97

75 181 181 199

31 15 31

137 140 152

100 232 239 215

28 26 30

167 168 177

150 293 337 320

31 36 29

299 283 323

200 335 366 353

46 41 41

402 393 415

400 401 333 338

55 58 55

545 553 555

800

23 12 23

7

0 18 30 19

10 21 15

25 43 39 50

23 18 8

50 53 48 64

19 20 14

75 89 86 78

16 18 25

100 149 128 161

29 25 29

125 165 160 160

26 30 18

200 233 276 211

20 30 26

8

0 33 35 24

20 19 20

25 39 31 33

27 25 23

50 69 67 63

18 23 25

75 72 73 73

43 20 19

100 121 113 111

28 27 24

125 139 148 145

21 22 28

250 227 220 194

35 25 29

500 227 220 203

45 56 48

9

0 33 22 35 24 34

35 33 22 24 29

1000 172 236 219

60 71 67

500 283 243 271

44 34 60

250 228 274 271

51 35 30

150 232 237 244

29 39 27

100 152 184 153

31 43 13

50 112 100 102

28 26 20

25 76 69 60

29 26 20



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

10

0 61 52 66

53 48 52

30 75 86 80

50 58 64

60 102 106 106

56 34 51

90 155 164 155

53 64 49

120 175 195 208

44 48 50

150 249 241 240

39 28 42

180 310 321 301

52 49 48

11

0 22 29 29 19 19 29 19 23 25

23 22 23

25 70 38 46 39 48 42 28 31 19

34 28 33

50 69 75 82 70 63 68 41 19 17

54 53 42

75 120 83 135 101 92 122 25 24 20

67 63 58

100 130 134 148 137 129 113 33 24 39

95 87 82

125 173 136 173 157 128 182 32 38 27

94 92 123

250 243 283 294 217 222 223 42 38 50

174 184 174

12

0 17 29 15

7 9 21

23 18 21

156 59 29 29

22 25 20

49 26 48

313 52 42 31

60 34 26

45 45 40

625 53 58 69

64 64 85

56 57 55

1250 112 53 72

82 77 86

70 61 58

2500 135 137 138

235 198 190

109 81 87

5000 77 102 85

61 97 43

153 188 148

(c: Contamination, M: Missing)

A-1.10 Sample E (TA100)


Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

1

0 98 99 106

100 96 91

91 92 100

10 111 95 143

98 121 150

100 95 109

25 110 149 120

137 144 124

100 95 111

50 126 144 135

109 118 111

113 125 106

75 166 179 174

164 123 150

141 144 139

125 193 215 176

138 144 136

148 154 162

250 235 257 260

152 157 140

188 204 189

2

0

50

100

150

200

250

300

3

0 135 136 123

125 122 126

25 161 143 232

154 172 165

50 172 199 175

180 154 180

75 200 190 158

152 168 167

100 217 223 236

201 180 226

125 249 229 191

190 178 191

250 259 228 270

154 167 165

500 243 290 296

120 141 148

4

0 222 183 193 187

144 155 143 146

165 180 202 204

50 272 238 208 240

193 162 164 197

224 190 216 193

100 221 213 248 208

158 198 166 170

197 192 158 204

200 266 236 289 234

178 194 190 153

224 177 166 173

300 292 253 269 286

226 205 181 193

213 258 290 285

500 276 269 276 268

188 194 221 257

300 249 314 262

1000 262 233 260 265

258 253 241 241

298 314 302 281

5

0 79 97 109

99 101 102

50 116 115 92

100 77 95

100 114 134 162

98 129 138

150 157 187 185

110 96 136

200 163 201 205

122 121 115

250 179 227 182

124 135 144

300 232 216 176

131 106 145

6

0 92 111 90

63 77 81

93 93 75

25 116 111 108

103 101 90

133 123 107

50 127 C 114

115 113 103

121 133 145

75 143 128 136

93 79 95

133 134 147

100 145 147 143

153 132 104

157 132 155

150 183 201 180

116 124 121

193 198 168

200 200 190 185

106 148 129

205 195 190



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

400 251 221 237

177 187 179

273 230 237

800

192 242 183

7

0 106 104 110

96 110 123

25 101 135 84

136 130 84

50 104 103 99

116 125 114

75 119 118 101

144 94 140

100 155 135 161

139 141 119

125 149 161 151

141 145 118

200 203 173 171

135 150 120

8

0 125 109 108

127 121 123

25 128 123 127

130 134 140

50 172 152 171

144 153 140

75 169 177 185

156 140 156

100 204 171 179

152 160 147

125 216 192 184

151 152 160

250 214 199 219

168 179 163

500 315 317 308

123 119 132

9

0 127 136 154 137 157

106 101 112 118 101

1000 302 327 320

tox tox tox

500 335 338 335

231 192 220

250 229 217 239

155 152 150

150 264 276 297

133 199 161

100 241 192 203

130 154 142

50 167 189 168

140 127 133

25 181 146 150

139 110 144

10

0 198 184 176

220 150 214

50 188 259 282

217 192 173

100 183 280 287

187 213 204

150 193 280 289

231 196 201

200 244 241 347

200 237 432

250 237 262 294

271 243 245

300 234 265 344

263 262 267

11

0 118 132 109 131 104 146 138 127 116

138 127 116

25 154 151 125 139 125 159 177 154 143

177 154 143

50 136 131 155 160 181 169 156 127 151

156 127 151

75 128 147 148 209 205 197 165 166 147

165 166 147

100 179 192 174 188 196 205 147 155 151

147 155 151

125 184 175 186 209 151 221 171 191 173

171 191 173

250 238 225 237 239 233 254 267 258 257

267 258 257

12

0 93 55 67

33 52 47

65 73 61

156 79 108 124

76 87 108

119 92 116

313 102 98 114

92 83 74

143 142 149

625 107 104 109

81 67 77

142 129 127

1250 139 124 148

81 82 64

153 114 126

2500 112 117 143

100 76 97

151 142 125

5000 153 125 140

63 68 92

128 109 129



A-1.11 Sample F (TA98)


Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

1

0 19 18 17 13 13 8 15 22 17

10 48 40 20 21 14 15 35 29 34

25 100 92 73 16 21 13 39 47 60

50 187 208 191 17 27 18 117 105 84

75 231 276 272 28 18 30 205 180 201

125 339 363 433 29 31 20 309 310 326

250 583 605 461 33 40 31 566 500 573

2

30 32 M 30 32 M

50 163 154 199 23 3 5

100 331 255 318 19 M 8

150 407 464 438 28 8 16

200 438 494 461 11 17 17

250 424 446 433 19 12 16

300 451 522 551 21 15 19

3

0 49 65 42 51 32 45

25 104 94 141 46 67 45

50 136 152 128 38 65 45

75 181 164 220 45 48 30

100 254 222 188 36 52 67

125 297 301 304 na 71 59

250 241 207 339 81 57 77

500 257 252 262 71 80 57

4

0 49 53 54 58 39 39 28 36 35 49 47 50

50 213 212 205 154 57 56 48 49 157 178 149 153

100 479 390 426 393 49 47 39 C 372 385 384 402

200 567 625 590 M 59 52 49 83 649 699 629 436

300 1064 944 847 702 69 57 77 51 1047 1112 967 1004

500 888 634 771 707 85 69 48 68 1272 899 1147 1247

1000 686 461 470 517 111 105 92 127 1345 1208 1135 1235

5

0 30 28 34 28 26 22

50 155 176 201 35 23 33

100 330 340 182 30 32 44

150 472 472 472 32 44 32

200 506 576 582 31 43 38

250 561 583 632 27 25 46

300 576 565 780 36 49 43

6

0 27 27 30 16 20 23 31 33 29

25 123 148 121 22 20 24 104 102 94

50 241 270 254 21 39 23 199 197 198

75 392 338 357 33 30 40 359 316 337

100 486 472 529 18 28 29 469 466 480

150 604 641 651 34 28 37 667 701 583

200 662 659 707 39 32 33 855 778 824

400 528 511 570 76 tox 54 1019 982 1053

800 tox tox tox

7

0 18 30 19 10 21 15

25 78 70 88 15 15 13

50 106 120 90 24 21 23

75 166 148 146 18 24 23

100 183 178 209 30 19 21

125 204 199 248 44 31 33

200 308 301 303 31 48 39

8

0 29 36 29 20 22 19

25 71 75 72 23 20 20

50 89 88 110 20 20 21

75 185 193 220 21 22 27

100 233 229 215 28 23 20

125 321 298 295 29 27 29

250 360 339 327 48 36 49

500 412 404 396 80 64 66

9

0 33 22 35 24 34 35 33 22 24 29

1000 462 492 469 57 56 75

500 454 533 531 54 46 49

250 458 439 432 45 29 43

150 443 489 493 26 35 47

100 371 350 378 31 35 35

50 215 212 186 25 47 24

25 118 115 125 29 38 20



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

10

0 61 52 66 53 48 52

30 109 117 105 48 46 41

60 194 202 177 37 48 43

90 264 297 356 54 58 56

120 426 382 362 42 38 48

150 476 468 534 30 47 35

180 557 511 672 38 27 36

11

0 15 20 20 20 23 15 20 11 19 35 19 14

25 125 87 106 77 56 81 24 19 19 52 38 46

50 168 139 160 129 143 139 42 41 42 80 86 73

75 201 224 231 204 162 179 34 33 51 105 115 119

100 285 261 320 206 233 258 39 37 36 147 132 186

125 345 350 281 275 339 281 27 37 34 210 206 223

250 429 394 339 447 342 394 60 43 45 415 398 339

12

0 17 29 15 7 9 21 23 18 21

156 42 35 50 18 17 24 32 43 48

313 48 84 54 21 12 26 77 54 40

625 76 95 76 31 35 21 100 67 102

1250 82 94 107 46 27 31 161 120 161

2500 117 147 150 57 51 43 273 83 129

5000 118 70 61 63 61 62 161 202 125

(c: Contamination, tox: Growth Inhibition, M: Missing)

A-1.12 Sample F (TA100)


Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

1

0 98 99 106

100 96 91

91 92 100

10 111 138 113

113 120 138

109 111 104

25 141 108 123

111 135 123

110 107 92

50 163 169 135

120 164 164

144 142 124

75 234 187 169

178 181 135

177 193 156

125 272 305 248

178 174 216

206 178 230

250 309 313 362

219 216 227

337 344 315

2

0

50

100

150

200

250

300

3

0 135 136 123

97 135 136

25 109 123 145

125 141 117

50 145 152 117

120 128 96

75 167 157 178

123 159 145

100 143 158 203

159 149 203

125 167 157 167

204 158 152

250 170 183 151

136 125 133

500 199 220 232

123 97 97

4

0 222 183 193 187

144 155 143 146

165 180 202 204

50 229 248 228 201

200 185 197 184

210 213 216 205

100 234 254 240 233

182 212 161 178

221 204 249 213

200 285 296 289 302

160 204 180 176

284 290 286 274

300 286 268 340 318

198 200 233 230

344 380 324 318

500 325 326 334 342

233 205 188 201

393 444 410 421

1000 325 286 241 272

220 221 218 246

430 418 438 349

5

0 113 92 74

93 77 94

50 90 134 84

93 100 108

100 166 164 157

115 120 129

150 221 205 239

133 100 104

200 233 242 273

111 132 124

250 234 224 288

100 92 98

300 246 201 287

107 102 117

6

0 92 111 90

63 77 81

93 93 75

25 133 135 149

84 94 121

116 111 108

50 181 146 162

103 115 98

144 126 135

75 179 181 147

111 114 101

190 167 163

100 214 206 202

112 96 102

193 174 183

150 253 277 235

124 129 163

255 244 244

200 223 211 217

129 145 121

336 263 311



Laboratory (µg/mL) 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 4 5 6

400 250 241 241

228 182 202

340 373 337

800

tox 186 tox

7

0 106 104 110

96 110 123

25 78 108 94

121 105 100

50 99 129 105

90 106 65

75 144 140 123

135 126 114

100 143 159 129

125 124 136

125 178 155 173

161 131 144

200 218 186 179

219 169 159

8

0 143 137 133

116 132 127

25 141 160 143

126 133 140

50 171 167 155

130 135 141

75 185 179 200

149 154 144

100 225 216 205

131 129 134

125 236 235 233

143 154 161

250 279 276 251

187 190 196

500 339 332 331

124 119 131

9

0 127 136 154 137 157

106 101 112 118 101

1000 367 356 386

tox tox tox

500 386 375 368

171 192 170

250 320 322 300

180 183 195

150 275 246 254

163 150 164

100 238 254 243

157 163 152

50 182 228 194

154 171 155

25 146 140 135

146 161 127

10

0 198 184 176

220 150 214

50 231 224 207

178 165 176

100 250 287 313

212 192 213

150 358 330 320

218 213 214

200 434 371 393

217 243 231

250 403 408 414

223 218 232

300 432 425 411

213 226 253

11

0 141 145 127 122 122 127 118 123 119

150 132 *

25 143 155 161 189 174 159 159 152 151

132 161 146

50 174 165 150 177 175 160 160 193 169

165 168 166

75 201 193 177 188 210 175 178 182 188

155 161 183

100 226 214 211 230 210 207 178 175 171

184 203 182

125 223 244 261 253 233 282 207 206 174

239 206 284

250 302 287 280 343 355 333 129 142 148

288 334 291

12

0 93 55 67

33 52 47

65 73 61

156 95 101 97

15 27 39

142 123 118

313 111 123 109

51 34 98

74 80 124

625 99 116 101

52 61 49

119 133 114

1250 131 140 109

64 61 59

117 101 109

2500 121 105 113

78 82 57

154 161 147

5000 108 91 148

127 216 87

83 114 114


A-1.13 Positive Control

TA98 TA100

Rep.

Positive Control Revertant/Plate Positive Control Revertant/Plate

S9 Lab# No. Chemical µg/plate 1 2 3 4 Chemical µg/plate 1 2 3 4

5% 1

1 Benzo[a]pyrene 1 370 374 424

Benzo[ a ]pyrene 1 770 862 865

S9 1 2-aminoanthracene 1 1204 1379 1514

2 1 not stated

not stated

2877 3154 2828 2519

3

1

Benzo[a]pyrene 2.5

582 508 499

2-nitrofluorene 1

1630 1574 1525

2 542 639 599

1618 1622 1680

3 614 590 531

1633 1807 1942

4 442 507 561

1920 1930 1862

5 493 525 484

1832 1936 1973

6 564 558 557

1822 1741 1968

4 1

Benzo[a]pyrene 100

µg/mL

148 216 204 208 2-aminoanthracene

50 µg/mL

3673 3765 3497 3121

2 440 542 509 526 5088 5136 4903 4973

5

1

Benzo[a]pyrene 1

511 480 562

2-aminoanthracene

5

612 655 695

2 241 199 211

361 384 442

3 323 281 289

423 343 440

4 146 102 179

251 316 247


TA98 TA100

Rep.

Positive Control Revertant/Plate Positive Control Revertant/Plate

S9 Lab# No. Chemical µg/plate 1 2 3 4 Chemical µg/plate 1 2 3 4

5 124 133 254

191 224 280

6 31 31 34

292 285 446

6 1 Benzo[a]pyrene 5 471 454 447

Benzo[a]pyrene 5 658 661 694

7

1

2-aminoanthracene 2

1515 1534 1455 2-

aminoanthracene 2

1005 1201 1101

2 1419 1399 1308

1755 1660 1479

3 1121 1237 1274

1567 1524 1554

8 1

2-aminoanthracene 2 1110 1130 1240

2-aminoanthracene

2 1600 1620 1730

2 1520 1410 1490

9 1 2-aminoanthracene 2.5 3595 3456 3594

Sodium Azide 1.5 3495 3308 3273

10

1

2-aminoanthracene 2

1150 1008 1067 2-

aminoanthracene 2

1109 1035 1057

2 1168 1372 1281

1684 1533 1529

3 1094 1178 1235

1136 1273 1177

12

1

not stated not

stated

196 147 132

not stated not

stated

936 495 873

2 142 133 154

976 573 835

3 197 142 156

502 864 927

0

1 1 2-nitrofluorene 1 567 605 740

Sodium Azide 1 639 706 680

3

1

Benzo[a]pyrene 2.5

390 407 330

Sodium Azide 2

1693 1092 1041

2 402 412 396

1168 1101 1199

3 445 385 432

1186 1142 1136

4 567 674 603

1123 1075 989

5 585 575 531

883 1091 1106

6 491 487 490

1087 1058 1139

4 1

2-nitrofluorene 50

µg/mL

1366 1251 1851 2159 Sodium Azide

20 µg/mL

1059 1229 1167 1207

2 2628 2006 2975 C 1346 1139 1196 1300

5

1

2-nitrofluorene 5

1117 1033 1149

Sodium Azide 2

2717 2875 2906

2 1339 1176 1191

2391 2367 2459

3 1221 1093 1268

2303 2036 2094

4 951 610 757

2408 2429 2623

5 707 741 682

2427 2667 2774

6 856 805 673

2356 2647 2454

6 1

2-(2-furyl)-3-(5-nitro- 0.1 861 832 919

2-(2-furyl)-3-(5-nitro- 0.01 403 438 453

2-furyl)acrylamide 2-furyl)acrylamide

7

1 4-nitroquinoline-

2

304 309 376

Sodium Azide 1

514 548 568

2 1-oxide 423 421 396

400 525 536

3

531 523 509

476 461 504

8 1 2-nitrofluorene 4 704 740 709

Sodium Azide 1 541 557 506

9 1 2-nitrofluorene 5 580 732 653

2-aminoanthracene

5 704 666 701

10

1

2-nitrofluorene 4

230 330 275

Sodium Azide 1

486 420 460

2 316 312 332

461 511 535

3 419 323 338

504 528 507

12

1

not stated not

stated

3012 1976 2351

not stated not

stated

1632 2357 1938

2 1354 1979 1538

2057 1983 1316

3 1136 2153 1877

2355 1936 1512

+10% S9

1 1 Benzo[a]pyrene 1 155 178 153

Benzo[ a ]pyrene 1 504 496 645

1 2-aminoanthracene 1 755 795 819

4 1

Benzo[a]pyrene 100

µg/mL

352 470 360 249 2-aminoanthracene

50 µg/mL

3190 2898 3006 2929

2 674 678 727 742 2345 2341 2222 2656

6 1 Benzo[a]pyrene 5 480 509 508

Benzo[a]pyrene 5 635 653 652

12

1

not stated not

stated

157 130 186

not stated not

stated

876 587 896

2 175 129 168

897 676 885

3 161 129 178

897 573 881

(c: Contamination)


Appendix B – Calculated Specific Activities (revertants/mg

NFDPM)

B-1 Canadian Monitor 8 (CM8)

LAB Sample TA98 TA98 TA100 TA100 TA98 TA100

# Name +S9 (5%) +S9 (10%) +S9 (5%) +S9 (10%) -S9 (5%) -S9 (5%)

1 A 1418 1084 937 633 82 396

1 C 1434 1088 861 580 125 948

1 E 1256 1106 941 511 81 333

Average 1369 1093 913 575 96 559

Std. Dev. 99 12 45 61 25 339

2 A 1704 0

2 C 1527 0

2 E 1820 0

Average 1684 0

Std. Dev. 147 0

3 A 1117 448 0 0

3 C 1390 810 142 0

3 E 1283 862 172 600

Average 1263 707 104 200

Std. Dev. 137 226 92 346

4 A 1458 1575 382 165 258 297

4 C 1531 1252 347 301 45 101

4 E 1475 1522 314 228 83 148

Average 1488 1450 348 231 129 182

Std. Dev. 38 174 34 68 114 102

5 A 1673 739 0 163

5 C 1580 454 0 0

5 E 1406 570 57 149

Average 1553 588 19 104

Std. Dev. 136 144 33 90

6 A 2435 2660 619 693 126 348

6 C 2165 2501 634 784 78 312

6 E 2077 2408 579 738 108 236

Average 2226 2523 611 738 104 298

Std. Dev. 186 127 29 46 24 57

7 A 1289 467 70 128

7 C 1056 522 210 520

7 E 1485 548 76 261

Average 1276 512 119 303

Std. Dev. 215 42 79 200

8 A 1555 655 57 302

8 C 1285 868 0 586

8 E 1172 870 0 307

Average 1338 798 19 398

Std. Dev. 196 124 33 162

9 A 1774 968 74 592

9 C 1740 924 40 521

9 E 1716 1102 51 428

Average 1743 998 55 514

Std. Dev. 29 93 17 82

10 A 2004 704 0 288

10 C 1834 802 0 480

10 E 1749 359 0 313

Average 1862 621 0 360

Std. Dev. 130 233 0 104

11 A 1268 972 682 527 159 546

11 C 1225 921 672 588 176 650

11 E 1381 803 689 626 106 360

Average 1291 899 681 580 147 519

Std. Dev. 81 87 9 50 37 147


B-2 Kentucky Reference 3R4F (3R4F)

LAB Sample TA98 TA98 TA100 TA100 TA98 TA100

# Name +S9 (5%) +S9 (10%) +S9 (5%) +S9 (10%) -S9 (5%) -S9 (5%)

1 B 3317 2460 1361 1030 158 866

1 D 3823 2456 1633 1069 117 622

1 F 3386 2480 1550 1080 96 947

Average 3509 2465 1515 1059 123 812

Std. Dev. 275 13 140 26 31 169

2 B 2655 0

2 D 2676 0

2 F 3035 0

Average 2789 0

Std. Dev. 214 0

3 B 1820 776 0 338

3 D 2084 932 131 711

3 F 2091 411 126 516

Average 1998 706 86 521

Std. Dev. 155 267 74 187

4 B 2405 2592 455 252 192 159

4 D 2950 2829 566 478 135 64

4 F 3079 3656 532 525 86 109

Average 2812 3026 518 418 138 111

Std. Dev. 358 559 57 146 53 47

5 B 3164 1186 37 142

5 D 3346 716 0 0

5 F 3036 975 48 181

Average 3182 959 28 108

Std. Dev. 155 236 25 95

6 B 4657 4901 1047 1129 89 323

6 D 4945 5162 953 1127 92 263

6 F 4668 4653 997 1201 69 295

Average 4757 4906 999 1152 83 294

Std. Dev. 163 255 47 42 12 30

7 B 2630 519 168 0

7 D 1826 640 236 386

7 F 1751 586 151 462

Average 2069 581 185 283

Std. Dev. 487 61 45 248

8 B 2388 944 0 320

8 D 2620 1222 44 267

8 F 2493 918 113 190

Average 2500 1028 52 259

Std. Dev. 116 169 57 65

9 B 2385 932 138 489

9 D 3254 1201 97 446

9 F 3424 983 47 353

Average 3021 1039 94 429

Std. Dev. 557 143 46 70

10 B 3450 945 113 219

10 D 3710 1214 0 356

10 F 3398 1098 0 190

Average 3520 1086 38 255

Std. Dev. 167 135 65 89

11 B 2454 1720 1006 682 173 486

11 D 2533 1933 1120 855 148 574

11 F 2629 1687 1002 790 123 580

Average 2538 1780 1043 776 148 547

Std. Dev. 88 133 67 88 25 52

Documents

Technical Report Ames Assay Proficiency Study · served as initial Study Trial Coordinator, followed by Betsy Bombick. 4. Study Protocol The protocol, summarized below, was reviewed