Quantification of HIV-1 using multiple competitors in a single-tube assay

Biotechniques. 1996 Aug;21(2):248-52, 253-5. doi: 10.2144/96212st01.


Methods for quantification of human immunodeficiency virus type 1 (HIV-1) based on competitive PCR and fragment analysis have been developed. Samples containing HIV-1 DNA and known amounts of three cloned competitors were co-amplified by PCR with semi-nested primers. The competitor DNAs contained the same long terminal repeat primer binding sequences as the wild-type DNA, but they are different in internal sequences and length. One of the inner primers was fluorescent-labeled to allow discrimination between the wild-type DNA and the three competitors by fragment analysis using a standard automated sequencer. A calibration curve using the peak area of the three competitors enabled accurate determination of target amount with minimal variations. The method presented here can be used for quantification of HIV-1 in clinical samples and will be useful for monitoring disease progression and treatment effects.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Binding, Competitive
  • Calibration
  • DNA Primers
  • HIV Infections / blood
  • HIV Infections / virology
  • HIV Long Terminal Repeat / genetics
  • HIV-1 / genetics
  • HIV-1 / isolation & purification*
  • Humans
  • Leukocytes, Mononuclear / virology
  • Polymerase Chain Reaction / instrumentation
  • Polymerase Chain Reaction / methods*
  • Viremia / virology


  • DNA Primers