title: Heat-mediated immune complex dissociation and enzyme-linked immunosorbent assay signal amplification render p24 antigen detection in plasma as sensitive as HIV-1 RNA detection by polymerase chain reaction.
abstract: OBJECTIVE
  To compare heat denaturation and acidification as immune complex dissociation (ICD) methods in adult HIV-1 infection and to increase the sensitivity by a signal-amplification-boosted HIV-1 p24 antigen enzyme-linked immunosorbent assay (ELISA).
  
  
  METHODS
  Prospective and retrospective blinded study of paired serum and plasma samples from 245 patients (112 of class A, 66 of B, 67 of C of the Centers for Disease Control and Prevention 1993 classification).
  
  
  METHODS
  Plasma and sera were prospectively tested for antigen by ELISA using native, acidified, or heat-denatured material. Retrospective tests included batch analysis of heat-denatured samples for antigen by the signal-amplification-boosted ELISA and for viral RNA.
  
  
  RESULTS
  With serum, native antigen was reactive in 26.5%. Acidification increased the rate to 53.1% (P < or = 0.0001), but was inefficient at a CD4+ count > or = 500 x 10(6)/l. Heat denaturation further elevated the rate to 67.8% (P < or = 0.0007) and the use of plasma to 78.0% (P < or = 0.008). The boosted ELISA, performed with plasma samples diluted 1 :6, which eliminated the problem of heat-induced sample coagulation, was confirmed positive in 89.5% of serum and 97.8% of plasma samples. RNA was detected in 95.7%.
  
  
  CONCLUSIONS
  Heat-mediated ICD combined with a signal-amplification-boosted ELISA detects HIV-1 expression as sensitively as a polymerase chain reaction kit for viral RNA, but at only a fraction of the cost. The procedure uses a 50 microliters plasma sample and should be fully automatable.
authors.affiliation: Swiss National Centre for Retroviruses, University of Zurich, Switzerland.
authors.forename: J
authors.initials: J
authors.lastname: Schüpbach
authors.forename: M
authors.initials: M
authors.lastname: Flepp
authors.forename: D
authors.initials: D
authors.lastname: Pontelli
authors.forename: Z
authors.initials: Z
authors.lastname: Tomasik
authors.forename: R
authors.initials: R
authors.lastname: Lüthy
authors.forename: J
authors.initials: J
authors.lastname: Böni
mesh_terms: D000328:Adult; D000936:Antigen-Antibody Complex; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D016655:HIV Core Protein p24 / Q000097:blood*; D015658:HIV Infections / Q000175:diagnosis*; D015497:HIV-1; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011489:Protein Denaturation; D012367:RNA, Viral / Q000097:blood*; D011933:Reagent Kits, Diagnostic; D016037:Single-Blind Method
chemical_list: D000936:Antigen-Antibody Complex; D016655:HIV Core Protein p24; D012367:RNA, Viral; D011933:Reagent Kits, Diagnostic