Use of competitive polymerase chain reaction to determine HIV-1 levels in response to antiviral treatments

S. M. Bruisten, M. H. Koppelman, M. T. Roos, A. E. Loeliger, P. Reiss, C. A. Boucher, H. G. Huisman

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

OBJECTIVE: To develop a competitive polymerase chain reaction technique with which to evaluate the usefulness of HIV-1 level as a marker of response to antiviral treatment. DESIGN: HIV-1 sequences were assessed by competitive polymerase chain reaction in four subjects participating in a double-blind study of monotherapy versus combination therapy with nucleoside analogues. METHODS: We inserted a mutant construct of the HIV-1 pol sequence into a commercial vector, enabling us to generate known amounts of mutant DNA and RNA for competitive polymerase chain reaction. To measure HIV-1 DNA copies in cells, the mutant DNA fragments were allowed to compete in a 10-fold dilution series with a constant amount of nucleic acid from the subject. To measure HIV-1 RNA copies in plasma, in vitro synthesized mutant RNA was added in a 10-fold dilution series to a constant amount of subject RNA and copy DNA was synthesized. DNA and copy DNA were used as the input for nested pol polymerase chain reaction. Mutant and wild-type amplimers were discriminated by size. RESULTS: The competitive polymerase chain reaction technique has been validated in model experiments and can be used over a broad range (at least 6 logs) of levels. Three of the four subjects showed a decline of 1 log in proviral DNA levels in cells after beginning antiviral treatment. All four showed a decline of at least 1 log in viral RNA levels in plasma, but this decline was transient in one subject. CONCLUSION: The HIV-1 sequence level is a useful marker in antiviral treatment studies
Original languageEnglish
Pages (from-to)S15-S20
JournalAIDS (London, England)
Volume7
Issue numberSuppl. 2
DOIs
Publication statusPublished - 1993

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