|Title / Titel||Detection and identification of retroviruses using an ultrasensitive reverse transcriptase test and a highly efficient method for molecular cloning|
|Abstract (PDF, 14 KB)|
|Summary / Zusammenfassung||We have developed an ultrasensitive test for the retroviral enzyme reverse transcriptase (RT) called PERT (Product-Enhanced Reverse Transcriptase) assay (Pyra et al. PNAS 1994;91:1544-8; U.S. patent nr.5'807'669; EU patent nr.0623176). The PERT assay permits detection of all retrovirus particles exhibiting an enzymatically active RT at a diagnostic sensitivity which is equal to that of specific viral RNA detection by PCR and has the additional advantage of not being negatively affected by sequence variability. For the identification of candidate retroviruses positive by the PERT assay, but negative by PCR for known retroviral sequences we have developed another highly efficient procedure. It permits amplification of the viral RU5 RNA sequence located at the 5'-end of the retroviral RNA from as little as 1000 copies. By making use of the redundancy of R at the 3'-end of the viral RNA genome, subsequent amplification of the entire retroviral genome by "long PCR" can be achieved (Weissmahr et al., J. Virol. 1997;71:3005-12).
We apply these methods to the following goals:
- Screening of human or animal disorders (cancer, autoimmune & other chronic diseases) for a possible involvement of exogenous or endogenous retroviruses (cf. Conrad et al., Cell 1997;90:303-13).
- Safety of biomedical materials (blood products, cell cultures, vaccines etc.).
- Safety aspects of xenotransplantation
- Characterization of candidate retroviruses thus identified.
- Quantification of specific retroviruses, HIV monitoring
- Drug susceptibility testing (HIV)
|Publications / Publikationen||Pyra H, Böni J, and Schüpbach J. Ultrasensitive retrovirus detection by a reverse transcriptase assay based on product enhancement. Proc. Natl. Acad. Sci. USA 1994; 91:1544-1548.Böni J, Pyra H, Schüpbach J. Sensitive detection and quantification of particle-associated reverse transcriptase in plasma of HIV-1 infected individuals by the product-enhanced reverse transcriptase assay. J. Med. Virol. 1996; 49:23-28.Schüpbach J, Pyra H, Jendis J., Tomasik Z, Böni J. Isolation of an in vitro transmissible agent with reverse transcriptase activity from a blood donor with a borderline-positive HIV-1 serology for more than five years. Clin. Diagn. Virol. 1996; 5: 197-203.Böni J, Stalder J, Reigel F, Schüpbach J. Detection of reverse transcriptase activity in live attenuated virus vaccines. Clin. Diagn. Virol. 1996; 5:43-53.Weissmahr RN, Schüpbach J, Böni J. Reverse transcriptase activity in chicken embryo fibroblast culture supernatants is associated with particles containing endogenous avian retrovirus EAV-0 RNA. J. Virol. 1997; 71: 3005-3012.Conrad B, Weissmahr RN, Böni J, Arcari R, Schüpbach J, Mach B. A human endogenous retroviral superantigen as candidate autoimmune gene in type 1 diabetes. Cell 1997; 90:303-313.Böni J and Schüpbach J. Reverse transcriptase assay based on product enhancement for assessing the drug susceptibility of retroviruses. In: Methods in Molecular Medicine, vol. 24: Antiviral Methods and Protocol. D. Edited by: D. Kinchington and R.F. Schinazi. HUMANA PRESS, Totowa NJ. 1999, pp.301-312.Bürgisser P, Vernazza P, Flepp M, Böni J, Tomasik Z, Hummel U, Pantaleo G, Schüpbach J and the Swiss HIV Cohort Study. Performance of five different assays for the quantification of viral load in subjects infected with various subtypes of HIV-1. J. AIDS 2000; 23:138-144.Huder JB, Böni J, Hatt J-M, Lutz H, Schüpbach J. Identification and characterization of two closely related unclassifiable endogenous retroviruses in pythons (Python molurus and P. curtus). J Virol 2002 76:7607-7615.Deichmann M, Huder JB, Kleist C, Naher H, Schupbach J, Boni J. Detection of reverse transcriptase activity in human melanoma cell lines and identification of a murine leukemia virus contaminant. Arch. Dermatol. Res. 2005; 296:345-352.|
|Keywords / Suchbegriffe||retrovirus, reverse transcriptase, diseases of unknown origin, autoimmune disorders, cancer, chronic diseases, xenotransplantation, veterinary disorders|
|Project leadership and contacts /
Projektleitung und Kontakte
|Funding source(s) /
|SNF (Personen- und Projektförderung), Other Public Sources (e.g. Federal or Cantonal Agencies), Others
|In collaboration with /
In Zusammenarbeit mit
|Duration of Project / Projektdauer||Jan 1994 to Apr 2015|