Performance of a fourth-generation HIV screening assay and an alternative HIV diagnostic testing algorithm

AIDS. 2013 Mar 13;27(5):731-7. doi: 10.1097/QAD.0b013e32835bc535.


Objective: We evaluated the performance of the GS fourth-generation antigen/antibody assay and compared Centers for Disease Control and Prevention's (CDC's) proposed alternative algorithm [repeatedly reactive fourth-generation immunoassay followed by an HIV-1/HIV-2 differentiation immunoassay and, if needed, nucleic acid test (NAT)] with the current algorithm (repeatedly reactive third-generation immunoassay followed by HIV-1 western blot).

Design: A convenience sample of the following four specimen sets was acquired: 10 014 from insurance applicants, 493 known western blot-positive, 20 known western blot-indeterminate specimens, and 230 specimens from 26 HIV-1 seroconverters.

Methods: Specimens were tested with the GS third-generation and fourth-generation immunoassays, the Multispot HIV-1/HIV-2 differentiation immunoassay, NAT, and western blot. We applied the two algorithms using these results.

Results: Among insurance specimens, 13 (0.13%) specimens were immunoassay repeatedly reactive: two were HIV-positive (repeatedly reactive by third-generation and fourth-generation immunoassays, and western blot and Multispot positive); two third-generation repeatedly reactive and nine fourth-generation repeatedly reactive specimens were false-positive. Third-generation and fourth-generation specificities were 99.98% [95% confidence interval (CI) 99.93-100%] and 99.91% (95% CI 99.84-99.96%), respectively.All HIV-1 western blot-positive specimens were repeatedly reactive by third-generation and fourth-generation immunoassays. By Multispot, 491 (99.6%) were HIV-1-positive and two (0.4%) were HIV-2-positive.Only eight (40%) western blot-indeterminate specimens were fourth-generation repeatedly reactive: six were Multispot and NAT-negative and two were Multispot HIV-1-positive but NAT-negative.The alternative algorithm correctly classified as positive 102 seroconverter specimens with the third-generation immunoassay and 130 with the fourth-generation immunoassay compared with 56 using the western blot with either immunoassay.

Conclusion: The alternative testing algorithm improved early infection sensitivity and identified HIV-2 infections. Two potential false-positive algorithm results occurred with western blot-indeterminate specimens.

Publication types

  • Evaluation Study

MeSH terms

  • Algorithms
  • Blotting, Western / methods
  • Diagnostic Tests, Routine / methods
  • HIV Antibodies* / immunology
  • HIV Infections / diagnosis*
  • HIV Infections / immunology
  • HIV-1 / immunology*
  • HIV-2 / immunology*
  • Humans
  • Immunoassay / methods*
  • Mass Screening / methods
  • Nucleic Acid Amplification Techniques / methods
  • Sensitivity and Specificity


  • HIV Antibodies