Iranian Journal of Microbiology 2011. 3(4):177-182.

Detection of human Papillomavirus 18 in cervical cancer samples using PCR-ELISA (DIAPOPS)
N Raji, M Sadeghizadeh, KN Tafreshi, E Jahanzad

Abstract


Background and Objectives: Human Papillomavirus (HPV) infection is a major risk factor for adenocarcinoma of the cervix. The high-risk types of the virus such as HPV16 and HPV18, which possess the E6 and E7 oncogenes, are responsible for approximately 50% of all cervical cancers. A rapid, sensitive and specific test has been proposed for detection of HPV to improve cervical cancer screening programs.
Objectives: The aim of this study was to develop a fast PCR-ELISA assay designated as DIAPOPS (Detection of Immobilized Amplified Products in a One Phase System)for detection of HPV16 and HPV18 types in SCC samples and Pap smears. The type specific primers and probes were designed for PCR and PCR-ELISA. The amplified products were hybridized with a specific biotin-labeled probe for HPV18 inner amplicons. The hybrids were detected with peroxidase conjugated avidin. The test was performed on the paraffin block and Pap smear samples from the cervical cancer patients, and the results of DIAPOPS were compared with conventional PCR assay.
Results: The 70 samples (SCC and Pap smear samples) were collected from Imam Khomeini and Mirzakoochak Khan Hospitals in Tehran. The PCR-based method detected six HPV16 positive, three HPV18 positive and Two HPV33 positive samples. DIAPOPS results were compared with the conventional PCR results and they showed an increase in sensitivity of the DIAPOPS test. Not only all of them were confirmed by PCR-ELISA but also three samples that conventional PCR showed negative for HPV18, were demonstrated positive by the PCR-ELISA method.
Conclusion: The results of the study show that modified PCR-ELISA assay is more sensitive to detect HPV types and can be used for diagnostic purposes.


Keywords


DIAPOPS; human papillomavirus; cervical cancer

Full Text:

PDF

References


Walboomers JMM, Jacobs MV, Manos MM, Bosch FX, Kummer JA, Shah K V, et al. Human papillomavirus is a necessary cause of invasive cervical cancer worldwide. J Pathol 1999; 189: 12-19.

Solomon D, Schiffman M, Tarone R. Comparison of three management strategies for patients with atypical squamous cells of undetermined significance: baseline results from a randomized trial. J Natl Cancer Inst 2001; 93: 293-299.

Gravitt PE, Peyton CL, Alessi TQ, Wheeler CM, Coutlee F, Hildesheim A, Schiffman MH, et al. Improved amplification of genital human papillomaviruses. J Clin Microbiol 2000; 38: 357-361.

De Roda Husman AM, Walboomers JMM, van den Brule AJC, Meijer CJLM, Snijders PJF. The use of general primers GP5 and GP6 elongated at thrir 3ends with adjacent highly conseved sequences improves human papillomavirus detection by polymerase chain reaction. J Gen Virol 1995; 76: 1057-1062.

Nelson JH, Howkins GA, Edlund K, Evander M, Kajelber L, Wadell G, Dillner J, Gerasimova T, Coker AL. Anoveland rapid PCR basedmethod for genotyping human papillomaviruses in clinical samples. J Clin Microbiol 2000; 38: 688-95.

Jacobs M V, Snijders P J, van den Brule A J, Helmerhorst T J, Meijer C J, Walboomers JM. A general primer GP5+/ GP6+ mediated PCR enzyme immunoassay method for rapid detection of 14 high-risk and 6 low-risk human papillomavirus genotypes in cervical scrapings. J Clin Microbiol 1997; 35: 791-795.

Venturoli S, Zerbini M, Laplaca M Jr, Antuono AD, Negosanti M, Gentilomi G, et al. Evaluatin of immunoassays for the detection and typing of PCR amplified human papilloma virus DNA. J Clin Pathol 1998; 51: 143-148.

Nagai Y, Maehama T, Asato T, Kanazawa K.Detection of Human Papillomavirus DNA in Primary and Metastatic Lisions of Carcinoma of the Cervix in Women from Okinawa, Japan. Am. J Clin Oncol (CCT)2001; 24: 160-166.

Nakhaie RS, Sadeghizadeh M, Saderi H, Tafreshi N, Behmanesh M, Shirzad H. Detection of types 40 and 41 adenoviruses in stool samples of diarrheal children by solid phase PCR. Iranian J Biotechnol 2007; 5:42-47.

Burd EM. Human papillomavirus and cervical cancer.Clin Microbiol Rev 2003; 16: 1-17.

Van den Brule AJ, Pol R, Fransen-Daalmeijer N, Schouls LM, Meijer CJ, Snijders PJ. GP5+/6+ PCR followed by reverse line blot analysis enables rapid and high-throughput identification of human papillomavirus genotypes. J Clin Microbiol 2002; 40: 779-787.

Zerbini M, Venturoli S, Cricca M, Gallinella G, De Simone P, Costa S, et al. Distribution and viral load of type specific HPVs in different cervical lesions as detected by PCR-ELISA. J Clin Pathol 2001; 54:377-380.

Schiffman MR, Herrero A, Hildesheim M E, Sherman M, Bratti S, Wacholder M, et al. HPV DNA testing in cervical cancer screening. Results from women in a high- risk province of Costa Rica. JAMA 2000; 283: 87-93.


Refbacks

  • There are currently no refbacks.


Creative Commons Attribution-NonCommercial 3.0

This work is licensed under a Creative Commons Attribution-NonCommercial 3.0 Unported License which allows users to read, copy, distribute and make derivative works for non-commercial purposes from the material, as long as the author of the original work is cited properly.