Analytical characterization of the APTIMA® HPV Assay

Dockter, Janel; Schröder, Astrid; Eaton, Barbara; Wang, Ann; Sikhamsay, Nathan; Morales, Liezel; Giachetti, Cristina

doi:10.1016/s1386-6532(09)70007-1

Cited by 70 publications

(68 citation statements)

References 22 publications

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“…34,35 The multiplex type-specific E7 PCR utilizes HPV type-specific primers targeting the E7 region for the detection of 19 HR-/pHR (HPV16, 18,26,31,33,35,39,45, 51, 52, 53, 56, 58, 59, 66, 68a, 68b, 70, 73, 82) and 2 LRtypes (HPV6, 11), with detection limits ranging from 10 to 1,000 copies of the viral genome. 34 Two primers for the amplification of b-globin were also included to provide a positive control for the quality of the template DNA.…”

Section: Type-specific E7 Pcr-mpg (Ts-e7-mpg) Assaymentioning

confidence: 99%

“…These are predominantly available as formalin-fixed paraffin-embedded (FFPE) tissues, which are suitable for IHC analysis, but due to the degradation and cross-linking present a technical challenge for nucleic acid analysis. 24 Several commercially available RNA assays based on detection of full-length E6/E7 transcripts, 25,26 as well as ''in house'' RNA assays based on detection of spliced E6*I/E7 transcripts, 15,16,27 have been described. These assays aim to amplify rather long cDNA sequence from cervical smear samples (up to 620 bp for full-length and up to 280 bp for spliced transcripts, respectively) and therefore are not suited for FFPE tissue analysis.…”

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confidence: 99%

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Biological activity of probable/possible high‐risk human papillomavirus types in cervical cancer

Halec

Schmitt

Dondog

et al. 2012

Intl Journal of Cancer

116

155

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Judging the carcinogenicity of human papillomavirus (HPV) types rarely found in cervical cancer (CxCa) is hindered by lack of studies of their biological activity in cancer tissues. To asses transcriptional activity of HPV types, we have developed ultrashort amplimer, splice-site specific, E6*I mRNA RT-PCR assays for 12 high-risk (HR)-HPV (IARC Group 1) and eight probable/ possible high-risk (pHR)-HPV types (IARC Group 2A/B carcinogens). Previously unreported E6*I splice sites of the six pHR-HPV types 26, 53, 67, 70, 73 and 82 were identified by cloning and sequencing. We analyzed 97 formalin-fixed paraffin-embedded (FFPE) Mongolian CxCa biopsies for presence of HPV DNA by two sensitive genotyping assays, for E6*I transcripts of all HR-/ pHR-HPV types identified and for expression of HPV surrogate markers p16 INK4a , pRb and p53. E6*I of at least one HR-/pHR-HPV was expressed in 94 (98%) of cancer tissues including seven with pHR-HPV types 26, 66, 70 or 82 as single transcribed types. Fifty-eight of E6*I mRNA transcribing cases were analyzable by immunohistochemistry and displayed p16 INK4a overexpression in 57 (98%), pRb downregulation in 56 (97%) and p53 downregulation in 36 (62%) tissues. The newly developed E6*I mRNA RT-PCR assays appeared to be highly sensitive method to analyze HPV transcription in FFPE materials. Our finding of viral oncogene transcription of pHR-HPV types 26, 66, 70 and 82 in cervical tumors, in the absence of any other transcriptionally active HR-type and with p16 INK4a overexpression and pRb downregulation, may support a reassessment of the carcinogenicity classification of these pHR-HPV types.Human papillomaviruses (HPV) with currently more than 150 types defined are a complex group differing in tropism and carcinogenic potential. Detection of HPV DNA in 99.7% of cervical cancer (CxCa) cases has established HPV as an etiological factor for CxCa development. 1,2 Epidemiological studies have demonstrated that 20 mucosal HPV types from five phylogenetically related species of the genus alpha (a5, a6, a7, a9 and a11-termed as ''high-risk clade'') are consistently found as single HPV infections in CxCa worldwide. [3][4][5][6] Based on their frequency in CxCa and available biological data, 12 of these types, i.e., types 16,18,31,33,35,39, 45, 51, 52, 56, 58 and 59 have been defined as carcinogens (IARC Group 1A)-hereafter referred to as high-risk (HR)-HPV. For eight other types, in the high-risk clade, i.e., types 26, 53, 66, 67, 68, 70, 73 and 82, the combination of their low frequency, lack of data on their active transcription and their transforming potential in model systems, has led them to be classified as only probable/possible carcinogens (IARC Groups 2A and 2B)-hereafter referred to as possible highrisk (pHR)-HPV types. 7 In the era of HPV-based cervical cancer precursor screening and of type-specific HPV vaccination, understanding the oncogenic properties of individual

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Section: Type-specific E7 Pcr-mpg (Ts-e7-mpg) Assaymentioning

confidence: 99%

mentioning

confidence: 99%

Biological activity of probable/possible high‐risk human papillomavirus types in cervical cancer

Halec

Schmitt

Dondog

et al. 2012

Intl Journal of Cancer

116

155

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“…Yöntem HR-HPV varlığı/yokluğu için pozitif veya negatif şeklinde kalitatif sonuç verirken, örnekte bulunan özgül HPV tiplerini tanımlamaz. Yöntem tek tüp içinde HPV'ye özgül yakalayıcı oligomerler ve manyetik mikropartiküller kullanılarak hedef mRNA'ların yakalanması, TMA ile hedef mRNA'ların amplifi kasyonu ve amplifi kasyon ürünlerinin saptanması olmak üzere üç ana basamak içerir 50 . Her üç basamağın (yakalama, amplifi kasyon ve saptama) performansını değerlendirmek üzere teste bir internal kontrol transkripti eklenmiştir.…”

Section: Hpv-rna Amplifi Kasyon Testleriunclassified

Current Problems and Recent Advances in the Molecular Diagnosis of Genital Human Papillomavirus Infections

Şahiner¹

2014

Mikrobiyol Bul

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ÖZETMoleküler tanı yöntemlerindeki gelişmelerin bir sonucu olarak, 1980'li yılların başlarında servikal kanser (SK) biyopsileri ve SK hücre kültürlerinin insan papillomavirus (HPV) DNA dizilerini içerdiği göste-rilmiştir. Sonraki yıllarda, HPV E6 ve E7 genlerinin SK dokularında eksprese edildiği ve bu onkoproteinlerin pRb ve p53 başta olmak üzere çeşitli hücresel proteinlerle etkileşim içinde olduğu keşfedilmiştir. Serviks kanseri ile HPV enfeksiyonları arasındaki ilişkinin gösterilmesi bu konuya olan ilginin artmasına neden olmuş ve yapılan geniş ölçekli epidemiyolojik çalışmalarda bazı HPV tiplerinin SK için majör risk faktörü olduğu belirlenmiştir. Yüksek riskli veya onkojenik tipler olarak adlandırılan bu tiplerin diğer anogenital kanserler ve baş-boyun kanserlerinin bir alt grubu ile de ilişkili olduğu belirlenmiştir. Bu süreçte her biri farklı yaklaşım ve uygulamaları içeren çok sayıda ticari ve kullanıcı tasarımlı (in-house) tanı yöntemi geliş-tirilmiş ve HPV-DNA tespitine dayalı testler SK tarama programlarının önemli bir parçası haline gelmiştir. HPV enfeksiyonlarının tanı ve takibinde (yönetiminde) kullanılan tarama-tiplendirme yöntemlerinin ve kombine yaklaşımların her birinin kendilerine özgü güçlü ve zayıf yönleri mevcuttur. Yöntem seçimini kritik ve karmaşık hale getiren, gerek metodoloji gerekse virusun doğasından kaynaklanan çeşitli zorluklar bulunmaktadır. Çok sayıda farklı HPV tipinin genital bölgede enfeksiyon etkeni olarak saptanabilmesi, viral genomun aynı HPV tipi içinde bile çok sayıda dizi varyasyonları gösterebilmesi, mevcut yöntemlerin saptama duyarlılığının HPV tiplerine ve incelenen örnekte bulunan viral DNA kopya sayılarına bağlı olarak değişebilmesi ve son olarak çoklu enfeksiyonları saptama ve tip tayini yapabilme yeteneğinin yöntemler arasında farklılıklar göstermesi bu zorluklardan bazılarıdır. Bu derleme yazıda, HPV enfeksiyonlarının tanı ve takibi için kullanılan yöntemlerin özetlenmesi ve yöntem seçiminde yol gösterebileceği düşünülen güncel bilgilerin paylaşılması hedefl enmiştir. Bu amaçla, geçmiş yıllarda kullanılan moleküler temelli HPV testleri ve bu testlerin gelişim sürecine kısaca değinilmiş, günümüzde kabul gören ve yaygın olarak kullanılan tanı, tarama ve tiplendirme yöntemlerinin avantaj ve dezavantajları ayrıntılı olarak tartışılmıştır.

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“…All studies have shown that HPV-Proofer has a lower clinical sensitivity for the detection of CIN2+ lesions than DNA-based assays, but a significantly higher clinical specificity. It is believed that the lower sensitivity of HPV-Proofer is mainly due to the detection of only five hr-HPVs, rather than the 13-14 types detected by DNAbased screening assays [139]. In the largest cross-sectional study to have compared HPV-Proofer and a DNA-based assay (GP5+/6+ PCR) on a total of 4136 women, 2.4 and 9.3% of cytologically normal women over 30 years were mRNA and DNA HPV positive, respectively [136].…”

Section: Pretect Hpv-proofermentioning

confidence: 99%

“…The assay generates a qualitative result for the presence/absence of 14 targeted HPVs and does not allow the exact determination of HPV type(s) present in a clinical specimen [139]. APTIMA is a single tube test and is based on three main steps:…”

Section: Aptima Hpv Assaymentioning

confidence: 99%

Commercially available assays for multiplex detection of alpha human papillomaviruses

Poljak

Kocjan

2010

Expert Review of Anti-infective Therapy

115

129

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Analytical characterization of the APTIMA® HPV Assay

Cited by 70 publications

References 22 publications

Biological activity of probable/possible high‐risk human papillomavirus types in cervical cancer

Biological activity of probable/possible high‐risk human papillomavirus types in cervical cancer

Current Problems and Recent Advances in the Molecular Diagnosis of Genital Human Papillomavirus Infections

Commercially available assays for multiplex detection of alpha human papillomaviruses

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