Quantification of infectious HIV-1 plasma viral load using a boosted in vitro infection protocol.
Methods currently used for HIV-1 viral load measurements are very sensitive, but cannot distinguish between infectious and noninfectious particles. Here we describe the development of a novel, sensitive, and highly reproducible method that allows rapid isolation and quantification of infectious particles from patient plasma. By immobilizing HIV-1 particles in human plasma to platelets using polybrene, we observed a 10- to 1000-fold increase in infectivity over infection protocols using free virus particles. Using this method, we evaluated infectivity in plasma from 52 patients at various disease stages. At plasma viral loads of 1000-10000 HIV-1 RNA copies/ml 18%, at 10,000-50,000 copies/ml 73%, at 50,000-100,000 copies/ml 90%, and above 100,000 copies 96% of cultures were positive. We found that infectious titers among patients vary distinctively but are characteristic for a patient over extended time periods. Furthermore, we demonstrate that by evaluating infectious titers in conjunction with total HIV RNA loads, subtle effects of treatment intervention on viremia levels can be detected. The immobilization procedure does not interfere with viral entry and does not restore the infectivity of neutralized virus. Therefore, this assay system can be utilized to investigate the influence of substances that specifically affect virion infectivity such as neutralizing antibodies, soluble CD4, or protease inhibitors. Measuring viral infectivity may thereby function as an additional, useful marker in monitoring disease progression and evaluating efficacy of antivirals in vivo.
Rusert
Peter
P
Division of Infectious Diseases, University Hospital Zurich, Switzerland.
Fischer
Marek
M
Joos
Beda
B
Leemann
Christine
C
Kuster
Herbert
H
Flepp
Markus
M
Bonhoeffer
Sebastian
S
Günthard
Huldrych F
HF
Trkola
Alexandra
A
D000328:Adult; D000368:Aged; D044966:Anti-Retroviral Agents / Q000627:therapeutic use; D001792:Blood Platelets / Q000821:virology; D002478:Cells, Cultured; D005260:Female; D015658:HIV Infections / Q000097:blood / Q000188:drug therapy / Q000821:virology*; D015497:HIV-1 / Q000235:genetics / Q000302:isolation & purification* / Q000472:pathogenicity; D006583:Hexadimethrine Bromide; D006801:Humans; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D010949:Plasma / Q000821:virology; D016133:Polymerase Chain Reaction; D012367:RNA, Viral / Q000032:analysis*; D012680:Sensitivity and Specificity; D019562:Viral Load / Q000379:methods*; D014771:Virion / Q000235:genetics
D044966:Anti-Retroviral Agents; D012367:RNA, Viral; D006583:Hexadimethrine Bromide