I N S T A N D e. V. Gesellschaft zur Förderung der Qualitätssicherung in medizinischen Laboratorien e. V. (vormals Hämometerprüfstelle) WHO Collaborating Centre for Quality Assurance and Standardization in Laboratory Medicine in cooperation with Deutsche Vereinigung zur Bekämpfung der Viruskrankheiten (DVV) Gesellschaft für Virologie (GfV) Deutsche Gesellschaft für Hygiene und Mikrobiologie (DGHM) INSTAND-Geschäftsstelle Ubierstr. 20 40223 Düsseldorf Telefon: +49 (0)211 1592 13 0 Fax: +49 (0)211 1592 1330 E-mail: instand@instand-ev.de Internet: www.instand-ev.de EQAS Adviser: Assistant EQAS Adviser: Prof. Dr. Heinz Zeichhardt Dr. Hans-Peter Grunert CharitéCentrum für diagnostische und präventive Labormedizin Institut für Biotechnologische Diagnostik in der GBD Institut für Virologie, Campus Benjamin Franklin Potsdamer Chaussee 80, 14129 Berlin Hindenburgdamm 27, 12203 Berlin Tel.: +49-(0)30-688197730; Fax: +49-(0)30-688197741 Tel.: +49-(0)30-84453625/23; Fax: +49-(0)30-84453626 Email: HPGrunert@gmx.de Email: Heinz.Zeichhardt@charite.de August 2011 Final Report External Quality Assessment Scheme (EQAS) - June/July 2011 Virus Genome Detection - Human Papilloma Viruses (373) Differentiation of High Risk and Low Risk Types and Typing (PCR/NAT HPV) INSTAND-Target Value Laboratories: Uniklinik Köln, Institut für Virologie, Nationales Referenzzentrum für Papillom- und Polyomaviren: Prof. Dr. H. Pfister, Prof. Dr. U. Wieland, Dr. R. Kaiser Charité - Universitätsmedizin Berlin, Campus Mitte, Institut für Medizinische Virologie, Nationales Konsiliarlaboratorium für Hantaviren: Prof. Dr. D. H. Krüger, PD Dr. J. Hofmann Klinikum der Johann Wolfgang Goethe-Universität, Institut für Medizinische Virologie, Frankfurt/Main: Prof. Dr. H. W. Doerr, Prof. Dr. H. Rabenau, PD Dr. A. Berger, Dr. R. Allwinn Labor Enders, Institut für Virologie, Infektiologie und Epidemiologie, Stuttgart: Prof. Dr. Gisela Enders & Partner Universitätsklinikum des Saarlandes, Institut für Virologie, Homburg/Saar; Prof. Dr. S. Smola, Prof. Dr. N. Müller-Lantzsch 373 Human Papilloma Viruses Genome June July 2011 Letter 20110831a.doc 1
Dear colleagues, Attached please find the final report for the EQAS "PCR-/NAT-HPV". Your certificate of successful participation, statement of participation and statement of your individual (validity period: 12 months) were sent out independently. The following test categories are individually stated on the documents: 20. Differentiation between "High Risk"- and "Low Risk" HPV - Differenzierung von HPV in "High Risk"- und "Low Risk"-Typen 25. Typing of "High Risk" HPV and detection of "Low Risk" HPV (without typing) - Typisierung von "High Risk"-HPV und Nachweis von "Low Risk"-HPV (ohne Typisierung) 30. Typing of HPV - Typisierung von HPV 35. Typing of "High Risk" HPV (without determination of "Low Risk" HPV) - Typisierung von "High Risk"-HPV (ohne Bestimmung von "Low Risk"-HPV) 60. Determination of "High Risk" HPV (without determination of "Low Risk" HPV) - Bestimmung von "High Risk"-HPV (ohne Bestimmung von "Low Risk"-HPV) The report for this EQA scheme is also available as PDF file on the INSTAND-Homepage under EQAS / Reports / Year and Category (Virus genome detection) in English language (http://www.instandev.de/en/eqas/reports/ ) and German language (http://www.instandev.de/ringversuche/kommentare/ ). Sample properties and evaluation In this EQAS the test are requested as follows for: Differentiation: High Risk, Low Risk, " or "indeterminate ; Typing: HPV (e.g. HPV 6 = 6 = human papilloma virus 6 ) Sample properties were as follows: Sample 373011: positive; lysate of HeLa cells, containing sequences of. Sample 373012: HPV 16 low positive; lysate of SiHa cells, containing sequences of HPV 16. Please see comment to sample 373012. Sample 373013: Biopsy material of a condyloma. The biopsy material is HPV 6 positive as analyzed by the INSTAND-Target Value Laboratories. Sample 373014: positive; lysate of HeLa cells, containing sequences of. Sample 373015: HPV ; lysate of MRC-5 cells (human lung fibroblasts), no HPV genome detectable. The positive samples 373011 and 373014 represented different dilution steps of a dilution series of a lysate of HeLa cells. Please see table 1 for details on sample properties, and success rates. A result was not considered for evaluation in case you had specified that this result should only be taken as additional information and ignored as valid result. For receiving a certificate of successful participation it is required that you analyzed all 5 samples of the sample set ly with the same method in the corresponding test categories (100 ). The evaluation criteria for the of EQA schemes in virology follow the new Guideline of the German Medical Association (Bundesärztekammer/ RiliBÄK = Richtlinie der Bundesärztekammer zur Qualitätssicherung laboratoriumsmedizinischer Untersuchungen). Please see section B2 of the RiliBÄK "qualitative determinations in laboratory medicine = Qualitative laboratoriumsmedizinische Untersuchungen" (effective since 01.07.2011 with a transition period of two years; Deutsches Ärzteblatt, Jg. 108, Heft 30, 29.07.2011, Seite A 1647-1651). Section B3 of the RiliBÄK "direct detection and characterization of infectious diseases pathogens = Direkter Nachweis und Charakterisierung von Infektionserregern" is in progress. Results The are summarized and differentiated for each sample according to test category 20, 25, 30, 35 and 60, respectively, in table 1. Detailed for the detection and differentiation between "High Risk" and "Low Risk" HPVs as well as for typing are shown in the following tables. Comment to sample 373012: Sample 373012 is low positive for HPV 16 due to the adjusted concentration of SiHa cells, which contain sequences of HPV 16. The INSTAND-Target Value Laboratories pre-tested this sample. Laboratories which applied PCR test systems reported HPV positive for this sample whereas a laboratory which used a 373 Human Papilloma Viruses Genome June July 2011 Letter 20110831a.doc 2
hybridization test (e.g. HPV screening test principle Hybrid Capture 2 Technology/Qiagen) reported. It could already be shown in previous EQA schemes that screening by hybridization methods for the detection of "High Risk" HPV s revealed a lower analytical sensitivity in comparison to PCR detection methods. In this EQA scheme those for sample 373012 which were received by application of the HPV-screening test "Hybrid Capture 2 Technology/Qiagen" were not evaluated for test categories 20 and 60 (without disadvantage for the certificate). The manufacturer concerned has already been informed. In recent years samples containing "High Risk" or "Low Risk" HPV s in different concentrations have sporadically been included into the EQA schemes in order to get insight into the analytical sensitivities of the different applied test systems. Objective of the INSTAND EQA schemes is to assess the performance of screening methods in comparison to the more sensitive PCR test systems including consecutive typing. The question of different analytical sensitivities of diverse methods for the detection of HPV was already addressed by the Jointed Diagnostic Council of the Deutsche Vereinigung zur Bekämpfung von Viruskrankheiten (DVV) and the Gesellschaft für Virologie (GfV) in accordance with Prof. Dr. H. Pfister, Director of the Nationales Referenzzentrum für Papillom- und Polyomaviren, Uniklinik Köln, Institut für Virologie, in August 2007. Test systems which show differences in analytical sensitivity are desirable and should be applied depending on the clinical status of the patient. The lower sensitivity of hybrid capture tests can be adequate or even of advantage for screening of cervical carcinoma and early detection of cervical intraepithelial neoplasia (CIN) under the prerequisite that these tests are able to detect not only HPV 16 and but also relevant "High Risk" HPV s. Enhanced sensitivity of PCR/NAT for the detection of HPV genome is desirable for: the detection of adenocarcinoma and for the early detection of adenocarcinoma in situ, the triage of women with cytologic diagnosis ASCUS (atypical squamous cells of undetermined significance), the control of women after therapy. Typing of HPV by PCR/NAT is reasonable for: the differentiation between reactivation of persisting HPV infection and primary infection/reinfection with a new HPV, the detection of multiple HPV s in immunocompromised patients. Comments to test categories Test category 35: Typing of "High Risk" HPV (without determination of "Low Risk" HPV) These tests exclusively detect and "High Risk" HPV and cannot detect "Low Risk" HPV. This means that a test result obtained by these tests can only be interpreted as " for High Risk" HPV. Results representing " for High Risk HPV" will be stated as "" in your statement of individual. Test category 60: Determination of "High Risk" HPV (without determination of "Low Risk" HPV) These tests exclusively detect "High Risk" HPV (without specification of the HPV ) and cannot detect "Low Risk" HPV. This means that a test result obtained by these tests can only be interpreted as " for High Risk HPV". Results representing " for High Risk HPV" will be stated as "" in your statement of individual. Determination of "High Risk HPV E6/E7 mrna One laboratory applied a TMA test for the detection of "High Risk HPV E6/E7 mrna. This test was assigned to test category 60 because this test system only detects E6/E7 mrna of a distinct number of "High Risk" HPV s. We gratefully acknowledge the good cooperation with Prof. Dr. H. Pfister, Prof. Dr. U. Wieland and Dr. R. Kaiser (Nationales Referenzzentrum für Papillom- und Polyomaviren, Institut für Virologie, Universität Köln) as well as Prof. Dr. H.-P. Berlien and Dr. U. Müller (Evangelische Elisabeth Klinik, Berlin). We additionally thank Dr. M. Enders and Dr. G. Schalasta (Labor Prof. Gisela Enders und Partner, Stuttgart) for the studies on suitability of the EQA samples for the detection of "High Risk" HPV E6/E7 mrna by a TMA test system. Thank you very much for your kind cooperation. Prof. Dr. H. Zeichhardt 373 Human Papilloma Viruses Genome June July 2011 Letter 20110831a.doc 3
Properties High Risk or Low Risk Table 1: Summary of sample properties, and success rates Sample 373011* Sample 373012 # Sample 373013 Sample 373014* Sample 373015 HPV positive HPV positive # HPV positive HPV positive HPV High Risk High Risk # Low Risk High Risk Type 18 16 # 6 18 Material lysate of HeLa cells lysate of SiHa cells # biopsy material lysate of HeLa cells lysate of MRC-5 cells Dilution 1 : 50* 1 : 1.6 1 : 25 1 : 100* --- Success rates for all 5 samples of the sample set & considered as for test category 20: differentiation between "High Risk" and "Low Risk" HPV (human papilloma viruses) High Risk High Risk # Low Risk High Risk 98.9 91.9 # 97.8 94.6 98.9 (91/92) (57/62) # (90/92) (87/92) (91/92) considered as for test category 25: typing of "High Risk" HPV and detection of "Low Risk" HPV (without typing) HPV 16 Low Risk / not done 100 (7/7) 100 (7/7) 100 (7/7) 100 (7/7) considered as for test category 30: typing of HPV HPV 16 HPV 6 / not done 98.7 (76/77) 92.2 (71/77) 100 (78/78) 97.4 (75/77) 100 (7/7) 98.7 (76/77) considered as for test category 35: typing of "High Risk" HPV (without determination of "Low Risk" HPV) HPV 16 for High Risk-HPV for High Risk-HPV 100 (8/8) 100 (8/8) 100 (7/7) considered as for test category 60: determination of "High Risk" HPV (without determination of "Low Risk" HPV) High Risk High Risk # for High Risk-HPV High Risk for High Risk-HPV 94.4 100 # 94.4 100 94.4 (17/18) (11/11) # (17/18) (18/18) (17/18) * Samples represent different dilution steps of the same dilution series of a lysate of HeLa cells. # Sample 373012 is low positive for HPV 16 due to the adjusted concentration of SiHa cells, which contain sequences of HPV 16. The for the HPV screening test principle "Hybrid Capture 2 Technology/Qiagen" reported for test categories 20 and 60, respectively, were not evaluated (without disadvantage for the certificate). For details see comment to sample 373012. & The success rates for all 5 samples of the corresponding sample set of test categories 20, 25, 30, 35 and 60, respectively, refer to the number of participating laboratories. Laboratories having reported obtained by several methods are recorded only once in the corresponding test category. 100 (8/8) 100 (8/8) 91.1 &# (82/90) 100 & (7/7) 88.2 & (67/76) 100 & (7/8) 88.9 &# (16/18) 373 Human Papilloma Viruses Genome June July 2011 Letter 20110831a.doc 4
INSTAND e. V., Gesellschaft zur Förderung der Qualitätssicherung in medizinischen Laboratorien e. V. (vormals Hämometerprüfstelle) - www.instand-ev.de in cooperation with Deutsche Vereinigung zur Bekämpfung der Viruskrankheiten e. V. (DVV) Gesellschaft für Virologie e. V. (GfV) Deutsche Gesellschaft für Hygiene und Mikrobiologie e. V. (DGHM) EQAS Virology June/July 2011 Virus genome detection PCR- / NAT-HPV (Human Papilloma Viruses) (373) Results for sample 373011 20. Differentiation between "High Risk" and "Low Risk" HPV Correct result: High Risk test in High Risk Low Risk indeterminate 92 91 1 0 0 98.9 High Risk Low Risk indeterminate hybridization (70) 76 75 1 0 0 98.7 Chipron / Chipron 3 3 100.0 Gen ID (AID) / Gen ID (AID) 12 12 100.0 Genomica / Genomica 1 1 100.0 Greiner / Greiner 4 4 100.0 Innogenetics / Innogenetics 10 10 100.0 Roche / Roche 6 6 100.0 in house / Qiagen 1 1 100.0 in house / Roche 1 1 100.0 in house / manuf. 1 1 100.0 manuf. / manuf. 1 1 100.0 nested PCR / hybridization (1270) in house / in house 1 1 100.0 Abbott / Abbott 4 4 100.0 hybridization / hybridization (3070) Qiagen / Qiagen 30 30 100.0 PCR / hybridization (9870) astra Diag. / astra Diag. 1 1 0.0 sequencing (72) 11 11 0 0 0 100.0 PCR / sequencing (1172) in house / in house 6 6 100.0 nested PCR / sequencing (1272) in house / in house 4 4 100.0 TaqMan / sequencing (1372) manuf. / manuf. 1 1 100.0 test (99) 5 5 0 0 0 100.0 PCR / test (1199) in house / in house 4 4 100.0 nested PCR / test (1299) in house / in house 1 1 100.0 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 1
Results for sample 373011 25. Typing of "High Risk" HPV and detection of "Low Risk" HPV (without typing) Correct result: test in 7 7 0 0 100.0 hybridization (70) 5 5 0 0 100.0 Gen ID (AID) / Gen ID (AID) 5 5 100.0 test (99) 2 2 0 0 100.0 PCR / test (1199) in house / in house 1 1 100.0 LightCycler / test (4099) in house / in house 1 1 100.0 30. Typing of HPV Correct result: test in Results for sample 373011 77 76 1 0 98.7 hybridization (70) 58 57 1 0 98.3 Chipron / Chipron 7 7 100.0 Gen ID (AID) / Gen ID (AID) 8 8 100.0 Genomica / Genomica 1 1 a 100.0 Greiner / Greiner 9 9 100.0 Innogenetics / Innogenetics 15 15 b, c 100.0 Roche / Roche 12 12 100.0 in house / Roche 1 1 100.0 in house / in house 1 1 100.0 in house / manufacturer 1 1 100.0 manufacturer / manufact. 1 1 100.0 nested PCR / hybridization (1270) in house / in house 1 1 100.0 andere PCR / hybridization (9870) astra Diag. / astra Diag. 1 1 d 0.0 a One participant detected additionally HPV 45. b One participant detected additionally HPV 39, HPV 68 and HPV 73. c One participant detected additionally HPV 39. d The sample 373011 comprises exclusively "High Risk" HPV 18. Therefore the reporting of "Low Risk" HPV 42, HPV 43 and HPV 44 was assessed as "in" result. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 2
30. Typing of HPV (continued) Results for sample 373011 sequencing (72) 13 13 0 0 100.0 PCR / sequencing (1172) in house / in house 8 8 100.0 nested PCR / sequencing (1272) in house / in house 4 4 100.0 TaqMan / sequencing (1372) manufacturer / manufact. 1 1 100.0 test (99) 6 6 0 0 100.0 PCR / test (1199) AmpliSens / AmpliSens 1 1 100.0 in house / in house 4 4 100.0 TaqMan / test (1399) Roche / Roche 1 1 100.0 Results for sample 373011 35. Typing of "High Risk" HPV (without determination of "Low Risk" HPV) (Tests for exclusive detection and typing of "High Risk"-HPV )* Correct result: test in * 8 8 0 0 100.0 * hybridization (70) 7 7 0 0 100.0 Abbott / Abbott 7 7 100.0 test (99) 1 1 0 0 100.0 TaqMan / test (1399) Roche / Roche 1 1 100.0 * These tests exclusively detect and "High Risk"-HPV and cannot detect "Low Risk"-HPV. This means that a test result obtained by these tests can only be interpreted as " for High Risk-HPV". Results representing " for High Risk-HPV" will be stated as "" in your statement of individual. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 3
Results for sample 373011 60. Determination of "High Risk" HPV (without determination of "Low Risk" HPV) (Tests which exclusively detect "High Risk"-HPV)* Correct result: High Risk test in High Risk Low Risk indeterminate below level of * 18 17 0 0 1 94.4 High Risk Low Risk indeterminate below level of * hybridization (70) 16 16 0 0 0 100.0 Roche / Roche 2 2 100.0 Abbott / Abbott 7 7 100.0 hybridization / hybridization (3070) Qiagen / Qiagen 7 7 100.0 test (99) 2 1 0 0 1 50.0 PCR / test (1199) manuf. / manuf. 1 1 0.0 TMA / test (6099) Gen-Probe / Gen-Probe 1 1 100.0 * These tests exclusively detect "High Risk"-HPV (without specification of the HPV ) and cannot detect "Low Risk"-HPV. This means that a test result obtained by these tests can only be interpreted as " for High Risk-HPV". Results representing " for High Risk-HPV" will be stated as "" in your statement of individual. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 4
Results for sample 373012 20. Differentiation between "High Risk" and "Low Risk" HPV Correct result: High Risk test in High Risk Low Risk indeterminate evaluated without evaluation # hybridization (70) # hybridization / hybridization (3070) # Qiagen / Qiagen # 62 30 # 30 # High Risk Low Risk indeterminate 57 2 # 2 # 0 0 # 1 2 # 2 # 4 26 # 26 # 91.9 hybridization (70) 46 43 0 0 3 93.5 Chipron / Chipron 3 3 100.0 Gen ID (AID) / Gen ID (AID) 12 10 2 83.3 Genomica / Genomica 1 1 100.0 Greiner / Greiner 4 4 100.0 Innogenetics / Innogenetics 10 10 100.0 Roche / Roche 6 6 100.0 in house / Qiagen 1 1 100.0 in house / Roche 1 1 100.0 in house / manuf. 1 1 100.0 manuf. / manuf. 1 1 100.0 nested PCR / hybridization (1270) in house / in house 1 1 100.0 Abbott / Abbott 4 4 100.0 PCR / hybridization (9870) astra Diag. / astra Diag. 1 1 0.0 sequencing (72) 11 11 0 0 0 100.0 PCR / sequencing (1172) in house / in house 6 6 100.0 nested PCR / sequencing (1272) in house / in house 4 4 100.0 TaqMan / sequencing (1372) manuf. / manuf. 1 1 100.0 test (99) 5 3 0 1 1 60.0 PCR / test (1199) in house / in house 4 2 1 1 50.0 nested PCR / test (1299) in house / in house 1 1 100.0 # Results without evaluation: please see comment to sample 373012. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 5
Results for sample 373012 25. Typing of "High Risk" HPV and detection of "Low Risk" HPV (without typing) Correct result: HPV 16 test in HPV 16 7 7 0 0 100.0 HPV 16 hybridization (70) 5 5 0 0 100.0 Gen ID (AID) / Gen ID (AID) 5 5 100.0 test (99) 2 2 0 0 100.0 PCR / test (1199) in house / in house 1 1 100.0 LightCycler / test (4099) in house / in house 1 1 100.0 30. Typing of HPV Correct result: HPV 16 test in Results for sample 373012 HPV 16 77 71 0 6 92.2 HPV 16 hybridization (70) 58 55 0 3 94.8 Chipron / Chipron 7 7 100.0 Gen ID (AID) / Gen ID (AID) 8 6 2 75.0 Genomica / Genomica 1 1 100.0 Greiner / Greiner 9 9 100.0 Innogenetics / Innogenetics 15 15 100.0 Roche / Roche 12 12 100.0 in house / Roche 1 1 100.0 in house / in house 1 1 100.0 in house / manufacturer 1 1 100.0 manufacturer / manufact. 1 1 100.0 nested PCR / hybridization (1270) in house / in house 1 1 100.0 andere PCR / hybridization (9870) astra Diag. / astra Diag. 1 1 0.0 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 6
30. Typing of HPV (continued) Results for sample 373012 sequencing (72) 13 13 0 0 100.0 PCR / sequencing (1172) 100.0 in house / in house 8 8 100.0 nested PCR / sequencing (1272) in house / in house 4 4 100.0 TaqMan / sequencing (1372) manufacturer / manufact. 1 1 100.0 test (99) 6 3 0 3 50.0 PCR / test (1199) AmpliSens / AmpliSens 1 1 100.0 in house / in house 4 1 3 25.0 TaqMan / test (1399) Roche / Roche 1 1 100.0 Results for sample 373012 35. Typing of "High Risk" HPV (without determination of "Low Risk" HPV) (Tests for exclusive detection and typing of "High Risk"-HPV )* Correct result: HPV 16 test in HPV 16 * 8 8 0 0 100.0 HPV 16 * hybridization (70) 7 7 0 0 100.0 Abbott / Abbott 7 7 100.0 test (99) 1 1 0 0 100.0 TaqMan / test (1399) Roche / Roche 1 1 100.0 * These tests exclusively detect and "High Risk"-HPV and cannot detect "Low Risk"-HPV. This means that a test result obtained by these tests can only be interpreted as " for High Risk-HPV". Results representing " for High Risk-HPV" will be stated as "" in your statement of individual. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 7
Results for sample 373012 60. Determination of "High Risk" HPV (without determination of "Low Risk" HPV) (Tests which exclusively detect "High Risk"-HPV)* Correct result: High Risk test in High Risk Low Risk indeterminate evaluated without evaluation # hybridization (70) # hybridization / hybridization (3070) # Qiagen / Qiagen # 11 High Risk Low Risk indeterminate 7 # 7 # 11 3 # 3 # 0 0 # 0 0 # below level of * 0 4 # 4 # below level of * 100.0 hybridization (70) 9 9 0 0 0 100.0 Roche / Roche 2 2 100.0 Abbott / Abbott 7 7 100.0 test (99) 2 2 0 0 0 100.0 PCR / test (1199) manuf. / manuf. 1 1 100.0 TMA / test (6099) Gen-Probe / Gen-Probe 1 1 100.0 * These tests exclusively detect "High Risk"-HPV (without specification of the HPV ) and cannot detect "Low Risk"-HPV. This means that a test result obtained by these tests can only be interpreted as " for High Risk-HPV". Results representing " for High Risk-HPV" will be stated as "" in your statement of individual. # Results without evaluation: please see comment to sample 373012. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 8
Results for sample 373013 20. Differentiation between "High Risk" and "Low Risk" HPV Correct result: Low Risk test in High Risk Low Risk indeterminate 92 0 90 0 2 97.8 High Risk Low Risk indeterminate hybridization (70) 75 0 73 0 2 97.3 Chipron / Chipron 3 3 100.0 Gen ID (AID) / Gen ID (AID) 12 11 1 91.7 Genomica / Genomica 1 1 100.0 Greiner / Greiner 4 4 100.0 Innogenetics / Innogenetics 10 10 100.0 Roche / Roche 6 6 100.0 in house / Qiagen 1 1 100.0 in house / Roche 1 1 100.0 in house / manuf. 1 1 100.0 manuf. / manuf. 1 1 100.0 nested PCR / hybridization (1270) in house / in house 1 1 100.0 astra Diag. / astra Diag. 1 1 100.0 hybridization / hybridization (3070) Qiagen / Qiagen 30 29 1 96.7 PCR / hybridization (9870) astra Diag. / astra Diag. 3 3 100.0 sequencing (72) 12 0 12 0 0 100.0 PCR / sequencing (1172) in house / in house 7 7 100.0 nested PCR / sequencing (1272) in house / in house 4 4 100.0 TaqMan / sequencing (1372) manuf. / manuf. 1 1 100.0 test (99) 5 0 5 0 0 100.0 PCR / test (1199) in house / in house 4 4 100.0 nested PCR / test (1299) in house / in house 1 1 100.0 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 9
Results for sample 373013 25. Typing of "High Risk" HPV and detection of "Low Risk" HPV (without typing) Correct result: Low Risk test in 7 7 0 0 100.0 hybridization (70) 5 5 0 0 100.0 Gen ID (AID) / Gen ID (AID) 5 5 100.0 test (99) 2 2 0 0 100.0 PCR / test (1199) in house / in house 1 1 100.0 LightCycler / test (4099) in house / in house 1 1 100.0 30. Typing of HPV Correct result: HPV 6 test in Results for sample 373013 HPV 6 78 78 0 0 100.0 HPV 6 hybridization (70) 60 60 0 0 100.0 Chipron / Chipron 7 7 a 100.0 Gen ID (AID) / Gen ID (AID) 8 8 100.0 Genomica / Genomica 1 1 100.0 Greiner / Greiner 9 9 a 100.0 Innogenetics / Innogenetics 15 15 a 100.0 Roche / Roche 13 13 100.0 in house / Roche 1 1 100.0 in house / in house 1 1 100.0 in house / manufacturer 1 1 100.0 manufacturer / manufact. 1 1 100.0 nested PCR / hybridization (1270) in house / in house 1 1 100.0 andere PCR / hybridization (9870) astra Diag. / astra Diag. 2 2 a 100.0 a In 11 participants detected additionally HPV 11. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 10
30. Typing of HPV (continued) Results for sample 373013 HPV 6 sequencing (72) 13 13 0 0 100.0 PCR / sequencing (1172) in house / in house 8 8 100.0 nested PCR / sequencing (1272) in house / in house 4 4 100.0 TaqMan / sequencing (1372) manufacturer / manufact. 1 1 100.0 test (99) 5 5 0 0 100.0 PCR / test (1199) AmpliSens / AmpliSens 1 1 100.0 in house / in house 4 4 100.0 Results for sample 373013 35. Typing of "High Risk" HPV (without determination of "Low Risk" HPV) (Tests for exclusive detection and typing of "High Risk"-HPV )* Correct result: for "High Risk" HPV test in HPV 6 * 7 0 0 7 100.0 HPV 6 * hybridization (70) 6 0 0 6 100.0 Abbott / Abbott 6 6 100.0 test (99) 1 0 0 1 100.0 TaqMan / test (1399) Roche / Roche 1 1 100.0 * These tests exclusively detect and "High Risk"-HPV and cannot detect "Low Risk"-HPV. This means that a test result obtained by these tests can only be interpreted as " for High Risk-HPV". Results representing " for High Risk-HPV" will be stated as "" in your statement of individual. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 11
Results for sample 373013 60. Determination of "High Risk" HPV (without determination of "Low Risk" HPV) (Tests which exclusively detect "High Risk"-HPV)* Correct result: for "High Risk" HPV test in High Risk Low Risk indeterminate below level of * 18 1 0 0 17 94.4 High Risk Low Risk indeterminate below level of * hybridization (70) 16 1 0 0 15 93.8 Roche / Roche 2 1 1 50.0 Abbott / Abbott 7 7 100.0 hybridization / hybridization (3070) Qiagen / Qiagen 7 7 100.0 test (99) 2 0 0 0 2 100.0 PCR / test (1199) manuf. / manuf. 1 1 100.0 TMA / test (6099) Gen-Probe / Gen-Probe 1 1 100.0 * These tests exclusively detect "High Risk"-HPV (without specification of the HPV ) and cannot detect "Low Risk"-HPV. This means that a test result obtained by these tests can only be interpreted as " for High Risk-HPV". Results representing " for High Risk-HPV" will be stated as "" in your statement of individual. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 12
Results for sample 373014 20. Differentiation between "High Risk" and "Low Risk" HPV Correct result: High Risk test in High Risk Low Risk indeterminate 92 87 1 1 3 94.6 High Risk Low Risk indeterminate hybridization (70) 76 71 1 1 3 93.4 Chipron / Chipron 3 3 100.0 Gen ID (AID) / Gen ID (AID) 12 11 1 91.7 Genomica / Genomica 1 1 100.0 Greiner / Greiner 4 4 100.0 Innogenetics / Innogenetics 10 10 100.0 Roche / Roche 6 5 1 83.3 in house / Qiagen 1 1 100.0 in house / Roche 1 1 100.0 in house / manuf. 1 1 100.0 manuf. / manuf. 1 1 100.0 nested PCR / hybridization (1270) in house / in house 1 1 100.0 Abbott / Abbott 4 4 100.0 hybridization / hybridization (3070) Qiagen / Qiagen 30 28 2 93.3 PCR / hybridization (9870) astra Diag. / astra Diag. 1 1 0.0 sequencing (72) 11 11 0 0 0 100.0 PCR / sequencing (1172) in house / in house 6 6 100.0 nested PCR / sequencing (1272) in house / in house 4 4 100.0 TaqMan / sequencing (1372) manuf. / manuf. 1 1 100.0 test (99) 5 5 0 0 0 100.0 PCR / test (1199) in house / in house 4 4 100.0 nested PCR / test (1299) in house / in house 1 1 100.0 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 13
Results for sample 373014 25. Typing of "High Risk" HPV and detection of "Low Risk" HPV (without typing) Correct result: test in 7 7 0 0 100.0 hybridization (70) 5 5 0 0 100.0 Gen ID (AID) / Gen ID (AID) 5 5 100.0 test (99) 2 2 0 0 100.0 PCR / test (1199) in house / in house 1 1 100.0 LightCycler / test (4099) in house / in house 1 1 100.0 30. Typing of HPV Correct result: Results for sample 373014 test in 77 75 1 1 97.4 hybridization (70) 58 56 1 1 96.6 Chipron / Chipron 7 7 100.0 Gen ID (AID) / Gen ID (AID) 8 8 100.0 Genomica / Genomica 1 1 100.0 Greiner / Greiner 9 9 100.0 Innogenetics / Innogenetics 15 15 a, b 100.0 Roche / Roche 12 11 1 91.7 in house / Roche 1 1 100.0 in house / in house 1 1 100.0 in house / manufacturer 1 1 100.0 manufacturer / manufact. 1 1 100.0 nested PCR / hybridization (1270) in house / in house 1 1 100.0 andere PCR / hybridization (9870) astra Diag. / astra Diag. 1 1 c 0.0 a One participant detected additionally HPV 39, HPV 68 and HPV 73. b One participant detected additionally HPV 39. c Sample 373014 comprises exclusively "High Risk" HPV 18. Therefore the reporting of "Low Risk" HPV 42, HPV 43 and HPV 44 was assessed as "in" result. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 14
30. Typing of HPV (continued) Results for sample 373014 sequencing (72) 13 13 0 0 100.0 PCR / sequencing (1172) 100.0 in house / in house 8 8 100.0 nested PCR / sequencing (1272) in house / in house 4 4 100.0 TaqMan / sequencing (1372) manufacturer / manufact. 1 1 100.0 test (99) 6 6 0 0 100.0 PCR / test (1199) AmpliSens / AmpliSens 1 1 100.0 in house / in house 4 4 100.0 TaqMan / test (1399) Roche / Roche 1 1 100.0 Results for sample 373014 35. Typing of "High Risk" HPV (without determination of "Low Risk" HPV) (Tests for exclusive detection and typing of "High Risk"-HPV )* Correct result: test in * 8 8 0 0 100.0 * hybridization (70) 7 7 0 0 100.0 Abbott / Abbott 7 7 100.0 test (99) 1 1 0 0 100.0 TaqMan / test (1399) Roche / Roche 1 1 100.0 * These tests exclusively detect and "High Risk"-HPV and cannot detect "Low Risk"-HPV. This means that a test result obtained by these tests can only be interpreted as " for High Risk-HPV". Results representing " for High Risk-HPV" will be stated as "" in your statement of individual. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 15
Results for sample 373014 60. Determination of "High Risk" HPV (without determination of "Low Risk" HPV) (Tests which exclusively detect "High Risk"-HPV)* Correct result: High Risk test in High Risk Low Risk indeterminate below level of * 18 18 0 0 0 100.0 High Risk Low Risk indeterminate below level of * hybridization (70) 16 16 0 0 0 100.0 Roche / Roche 2 2 100.0 Abbott / Abbott 7 7 100.0 hybridization / hybridization (3070) Qiagen / Qiagen 7 7 100.0 test (99) 2 2 0 0 0 100.0 PCR / test (1199) manuf. / manuf. 1 1 100.0 TMA / test (6099) Gen-Probe / Gen-Probe 1 1 100.0 * These tests exclusively detect "High Risk"-HPV (without specification of the HPV ) and cannot detect "Low Risk"-HPV. This means that a test result obtained by these tests can only be interpreted as " for High Risk-HPV". Results representing " for High Risk-HPV" will be stated as "" in your statement of individual. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 16
Results for sample 373015 20. Differentiation between "High Risk" and "Low Risk" HPV Correct result: test in High Risk Low Risk indeterminate 92 0 0 1 91 98.9 High Risk Low Risk indeterminate hybridization (70) 76 0 0 1 75 98.7 Chipron / Chipron 3 3 100.0 Gen ID (AID) / Gen ID (AID) 12 1 11 91.7 Genomica / Genomica 1 1 100.0 Greiner / Greiner 4 4 100.0 Innogenetics / Innogenetics 10 10 100.0 Roche / Roche 6 6 100.0 in house / Qiagen 1 1 100.0 in house / Roche 1 1 100.0 in house / manuf. 1 1 100.0 manuf. / manuf. 1 1 100.0 nested PCR / hybridization (1270) in house / in house 1 1 100.0 Abbott / Abbott 4 4 100.0 hybridization / hybridization (3070) Qiagen / Qiagen 30 30 100.0 PCR / hybridization (9870) astra Diag. / astra Diag. 1 1 100.0 sequencing (72) 11 0 0 0 11 100.0 PCR / sequencing (1172) in house / in house 6 6 100.0 nested PCR / sequencing (1272) in house / in house 4 4 100.0 TaqMan / sequencing (1372) manuf. / manuf. 1 1 100.0 test (99) 5 0 0 0 5 100.0 PCR / test (1199) in house / in house 4 4 100.0 nested PCR / test (1299) in house / in house 1 1 100.0 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 17
Results for sample 373015 25. Typing of "High Risk" HPV and detection of "Low Risk" HPV (without typing) Correct result:, not done test in / not done 7 0 0 7 100.0 not done hybridization (70) 5 0 0 5 100.0 Gen ID (AID) / Gen ID (AID) 5 5 100.0 test (99) 2 0 0 2 100.0 PCR / test (1199) in house / in house 1 1 100.0 LightCycler / test (4099) in house / in house 1 1 100.0 30. Typing of HPV Correct result:, not done Results for sample 373015 test in not done 77 0 0 76 98.7 not done hybridization (70) 58 0 0 57 98.3 Chipron / Chipron 7 7 100.0 Gen ID (AID) / Gen ID (AID) 8 7 a 87.5 Genomica / Genomica 1 1 100.0 Greiner / Greiner 9 9 100.0 Innogenetics / Innogenetics 15 15 100.0 Roche / Roche 12 12 100.0 in house / Roche 1 1 100.0 in house / in house 1 1 100.0 in house / manufacturer 1 1 100.0 manufacturer / manufact. 1 1 100.0 nested PCR / hybridization (1270) in house / in house 1 1 100.0 andere PCR / hybridization (9870) astra Diag. / astra Diag. 1 1 100.0 a One participant did not report any result for sample 373015. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 18
30. Typing of HPV (continued) Results for sample 373015 sequencing (72) 13 0 0 13 100.0 PCR / sequencing (1172) in house / in house 8 8 100.0 nested PCR / sequencing (1272) in house / in house 4 4 100.0 TaqMan / sequencing (1372) manufacturer / manufact. 1 1 100.0 test (99) 6 0 0 6 100.0 PCR / test (1199) AmpliSens / AmpliSens 1 1 100.0 in house / in house 4 4 100.0 TaqMan / test (1399) Roche / Roche 1 1 100.0 Results for sample 373015 35. Typing of "High Risk" HPV (without determination of "Low Risk" HPV) (Tests for exclusive detection and typing of "High Risk"-HPV )* Correct result: for "High Risk" HPV test in * 8 0 0 8 100.0 * hybridization (70) 7 0 0 7 100.0 Abbott / Abbott 7 7 100.0 test (99) 1 0 0 1 100.0 TaqMan / test (1399) Roche / Roche 1 1 100.0 * These tests exclusively detect and "High Risk"-HPV and cannot detect "Low Risk"-HPV. This means that a test result obtained by these tests can only be interpreted as " for High Risk-HPV". Results representing " for High Risk-HPV" will be stated as "" in your statement of individual. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 19
Results for sample 373015 60. Determination of "High Risk" HPV (without determination of "Low Risk" HPV) (Tests which exclusively detect "High Risk"-HPV)* Correct result: for "High Risk" HPV test in High Risk Low Risk indeterminate below level of * 18 1 0 0 17 94.4 High Risk Low Risk indeterminate below level of * hybridization (70) 16 1 0 0 15 93.8 Roche / Roche 2 1 1 50.0 Abbott / Abbott 7 7 100.0 hybridization / hybridization (3070) Qiagen / Qiagen 7 7 100.0 test (99) 2 0 0 0 2 100.0 PCR / test (1199) manuf. / manuf. 1 1 100.0 TMA / test (6099) Gen-Probe / Gen-Probe 1 1 100.0 * These tests exclusively detect "High Risk"-HPV (without specification of the HPV ) and cannot detect "Low Risk"-HPV. This means that a test result obtained by these tests can only be interpreted as " for High Risk-HPV". Results representing " for High Risk-HPV" will be stated as "" in your statement of individual. 373 Human Papilloma Viruses Genome June July 2011 Table 20110830a.doc 20