Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Experiment Videos

An improved non-radioisotopic reverse transcriptase assay and its evaluation

T Nakano1, K Sano, F Odawara

  • 1Department of Microbiology, Osaka Medical College.

Kansenshogaku Zasshi. the Journal of the Japanese Association for Infectious Diseases
|July 1, 1994
PubMed
Summary

We developed a highly sensitive, non-radioisotopic reverse transcriptase assay (RTA) using primer-template immobilization. This improved RTA detects human immunodeficiency virus type-1 (HIV-1) effectively, offering advantages over radioisotopic methods for virus isolation.

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Differential patterns of expression of glycosylphosphatidylinositol-anchored carcinoembryonic antigen and alkaline phosphatase in various cancer cell lines.

Cytotechnology·2008
Same author

Relationship between distance of schools from the nearest municipal waste incineration plant and child health in Japan.

European journal of epidemiology·2005
Same author

Increasing the thermostability of Flavobacterium meningosepticum glycerol kinase by changing Ser329 to Asp in the subunit interface region.

Protein engineering·2001
Same author

[Anhidrosis during long-term hydroxyurea therapy in a patient with chronic myelogenous leukemia].

[Rinsho ketsueki] The Japanese journal of clinical hematology·2001
Same author

Interaction of two arginine residues in lactate oxidase with the enzyme flavin: conversion of FMN to 8-formyl-FMN.

Proceedings of the National Academy of Sciences of the United States of America·2000
Same author

Surveillance of the serum Candida antigen titer for initiation of antifungal therapy after post-remission chemotherapy in patients with acute leukemia.

International journal of hematology·2000

Area of Science:

  • Biochemistry
  • Virology
  • Assay Development

Background:

  • Radioisotopic reverse transcriptase assays (RI-RTA) are standard but involve radioactivity.
  • Previous non-radioisotopic (non-RI) methods utilized primer immobilization.
  • An improved non-RI RTA is needed for sensitive and safe detection of viral reverse transcriptase activity.

Purpose of the Study:

  • To develop and validate an improved, highly sensitive non-radioisotopic reverse transcriptase assay (non-RI RTA).
  • To compare the sensitivity and performance of the new non-RI RTA against a radioisotopic RTA (RI-RTA).
  • To assess the utility of the improved non-RI RTA for human immunodeficiency virus type-1 (HIV-1) detection in clinical samples.

Main Methods:

  • Developed an improved non-RI RTA based on primer-template immobilization.

Related Experiment Videos

  • Tested template specificity, reproducibility, and linearity of the assay using human immunodeficiency virus type-1 (HIV-1) RT.
  • Compared sensitivities of the improved non-RI RTA, a previous non-RI RTA, and a sensitive RI-RTA with various HIV-1 samples.

    • Main Results:

    • The improved non-RI RTA demonstrated high sensitivity, outperforming the previous non-RI method and matching RI-RTA for most samples.
    • Fetal bovine serum in culture medium interfered with the assay, requiring a four-fold dilution for optimal sensitivity.
    • Despite a slight decrease in sensitivity for culture supernatants compared to RI-RTA, the improved non-RI RTA successfully detected all virus-positive cultures.

    Conclusions:

    • The improved non-RI RTA offers excellent sensitivity and avoids the need for radioisotopes.
    • This assay is particularly useful for virus isolation from culture supernatants, especially for HIV-1 detection.
    • The primer-template immobilization strategy enhances the performance of non-RI RTAs.