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We developed an improved, highly sensitive non-radioisotopic (non-RI) reverse transcriptase (RT) assay (RTA). While the original non-RI method previously reported made use of primer immobilization, our improved method was based on a primer-template immobilization procedure. We tested the template specificity, reproducibility and linearity of the new method in assays of human immunodeficiency virus type-1 (HIV-1) RT. The sensitivities of the method previously reported, the improved method and the sensitive radioisotopic (RI-) RTA were compared in assays of recombinant HIV-1 RT, partially purified HIV-1 particles, and the culture supernatant derived from HIV-1-infected cells. For each of these samples except the culture supernatant the improved method was the most sensitive. It appeared that the fetal bovine serum presented in the culture medium interfered with the assay reaction. The curve describing inhibition of the assay reaction by fetal bovine serum showed that the highest degree of sensitivity in the assay was obtained when the culture supernatant sample was diluted four times. With this degree of dilution, the sensitivity of the new method for assay of culture supernatant sample was still half that of the sensitive RI-RTA. Culture supernatants of five peripheral blood mononuclear cell samples obtained from HIV-1-seropositive carriers were assayed by both the improved method and the sensitive RI-RTA; and with each of the methods, however all virus-positive cultures could be detected. The improved non-RI RTA was considered especially useful for assay of culture supernatants for purposes of virus isolation because of its advantages of excellent sensitivity and lack of requirement for radioisotopes.
T Nakano, and
K Sano, and
F Odawara, and
Y Saitoh, and
T Otake, and
T Nakamura, and
K Hayashi, and
H Misaki, and
M Nakai
April 1997,
Journal of virological methods,
T Nakano, and
K Sano, and
F Odawara, and
Y Saitoh, and
T Otake, and
T Nakamura, and
K Hayashi, and
H Misaki, and
M Nakai
November 1992,
Journal of virological methods,
T Nakano, and
K Sano, and
F Odawara, and
Y Saitoh, and
T Otake, and
T Nakamura, and
K Hayashi, and
H Misaki, and
M Nakai
October 1999,
Journal of virological methods,
T Nakano, and
K Sano, and
F Odawara, and
Y Saitoh, and
T Otake, and
T Nakamura, and
K Hayashi, and
H Misaki, and
M Nakai
August 2015,
Journal of clinical microbiology,
T Nakano, and
K Sano, and
F Odawara, and
Y Saitoh, and
T Otake, and
T Nakamura, and
K Hayashi, and
H Misaki, and
M Nakai
June 1999,
Journal of clinical microbiology,
T Nakano, and
K Sano, and
F Odawara, and
Y Saitoh, and
T Otake, and
T Nakamura, and
K Hayashi, and
H Misaki, and
M Nakai
March 2008,
Analytical biochemistry,
T Nakano, and
K Sano, and
F Odawara, and
Y Saitoh, and
T Otake, and
T Nakamura, and
K Hayashi, and
H Misaki, and
M Nakai
February 2003,
International journal of STD & AIDS,
T Nakano, and
K Sano, and
F Odawara, and
Y Saitoh, and
T Otake, and
T Nakamura, and
K Hayashi, and
H Misaki, and
M Nakai
August 2012,
Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology,
T Nakano, and
K Sano, and
F Odawara, and
Y Saitoh, and
T Otake, and
T Nakamura, and
K Hayashi, and
H Misaki, and
M Nakai
March 2015,
Journal of medicinal chemistry,
T Nakano, and
K Sano, and
F Odawara, and
Y Saitoh, and
T Otake, and
T Nakamura, and
K Hayashi, and
H Misaki, and
M Nakai
January 1991,
Journal of virological methods,
