A computer-based, image-analysis method to quantify HIV-1 infection in a single-cycle infectious center assay

被引:18
作者
Day, John R.
Martinez, Laura E.
Sasik, Roman
Hitchin, Douglas L.
Dueck, Megan E.
Richman, Douglas D.
Guatelli, John C.
机构
[1] Univ Calif San Diego, Dept Pathol, Dept 0679, La Jolla, CA 92093 USA
[2] San Diego Vet Affairs Healthcare Syst, San Diego, CA USA
[3] Univ Calif San Diego, Dept Med, La Jolla, CA 92093 USA
关键词
HIV; infectivity; beta-galactosidase; software; quantify; rapid;
D O I
10.1016/j.jviromet.2006.06.019
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Rapid and precise quantitation of the infectivity of HIV is important for molecular virologic studies as well as for measuring the activities of antiviral drugs and neutralizing antibodies. In the present study, an indicator cell line and image-analysis software were used to quantify HIV infectivity. Cells of the P4R5 line, which express the receptors for HIV infection as well as P-galactosidase under the control of the HIV-1 long terminal repeat, were infected either with CXCR4- or CCR5-using viruses, including primary isolates, then stained 2 days later with X-gal to turn infected cells blue. Digital images of monolayers of the infected cells were captured using a high resolution CCD video camera and a macro video zoom lens. A software program was developed to process the images and to count the blue-stained foci of infection. The assay was applied to assess the infectivity of site-directed viral mutants, and to measure the activity of antiviral drugs and neutralizing antibody. The results indicate that the described method allows for the rapid quantitation of infected cells over a wide range of viral inocula with reproducibility, accuracy and relatively low cost. (c) 2006 Elsevier B.V. All rights reserved.
引用
收藏
页码:125 / 133
页数:9
相关论文
共 21 条
[1]   HIV-1 INDICATOR CELL-LINES [J].
AKRIGG, A ;
WILKINSON, GWG ;
ANGLISS, S ;
GREENAWAY, PJ .
AIDS, 1991, 5 (02) :153-158
[2]   Comprehensive cross-clade neutralization analysis of a panel of anti-human immunodeficiency virus type 1 monoclonal antibodies [J].
Binley, JA ;
Wrin, T ;
Korber, B ;
Zwick, MB ;
Wang, M ;
Chappey, C ;
Stiegler, G ;
Kunert, R ;
Zolla-Pazner, S ;
Katinger, H ;
Petropoulos, CJ ;
Burton, DR .
JOURNAL OF VIROLOGY, 2004, 78 (23) :13232-13252
[3]   Human immunodeficiency virus type 1 coreceptors participate in postentry stages in the virus replication cycle and function in simian immunodeficiency virus infection [J].
Chackerian, B ;
Long, EM ;
Luciw, PA ;
Overbaugh, J .
JOURNAL OF VIROLOGY, 1997, 71 (05) :3932-3939
[4]   HIV-1 REVERSE TRANSCRIPTION - A TERMINATION STEP AT THE CENTER OF THE GENOME [J].
CHARNEAU, P ;
MIRAMBEAU, G ;
ROUX, P ;
PAULOUS, S ;
BUC, H ;
CLAVEL, F .
JOURNAL OF MOLECULAR BIOLOGY, 1994, 241 (05) :651-662
[5]   DEVELOPMENT OF A SENSITIVE QUANTITATIVE FOCAL ASSAY FOR HUMAN IMMUNODEFICIENCY VIRUS INFECTIVITY [J].
CHESEBRO, B ;
WEHRLY, K .
JOURNAL OF VIROLOGY, 1988, 62 (10) :3779-3788
[6]   OPTIMAL INFECTIVITY IN-VITRO OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 REQUIRES AN INTACT NEF GENE [J].
CHOWERS, MY ;
SPINA, CA ;
KWOH, TJ ;
FITCH, NJS ;
RICHMAN, DD ;
GUATELLI, JC .
JOURNAL OF VIROLOGY, 1994, 68 (05) :2906-2914
[7]   The membrane-proximal tyrosine-based sorting signal of human immunodeficiency virus type 1 gp41 is required for optimal viral infectivity [J].
Day, JR ;
Münk, C ;
Guatelli, JC .
JOURNAL OF VIROLOGY, 2004, 78 (03) :1069-1079
[8]   Identification of a major co-receptor for primary isolates of HIV-1 [J].
Deng, HK ;
Liu, R ;
Ellmeier, W ;
Choe, S ;
Unutmaz, D ;
Burkhart, M ;
DiMarzio, P ;
Marmon, S ;
Sutton, RE ;
Hill, CM ;
Davis, CB ;
Peiper, SC ;
Schall, TJ ;
Littman, DR ;
Landau, NR .
NATURE, 1996, 381 (6584) :661-666
[9]   INFECTION OF HTLV-III/LAV IN HTLV-I-CARRYING CELLS MT-2 AND MT-4 AND APPLICATION IN A PLAQUE-ASSAY [J].
HARADA, S ;
KOYANAGI, Y ;
YAMAMOTO, N .
SCIENCE, 1985, 229 (4713) :563-566
[10]   DETECTION OF REPLICATION-COMPETENT AND PSEUDOTYPED HUMAN-IMMUNODEFICIENCY-VIRUS WITH A SENSITIVE CELL-LINE ON THE BASIS OF ACTIVATION OF AN INTEGRATED BETA-GALACTOSIDASE GENE [J].
KIMPTON, J ;
EMERMAN, M .
JOURNAL OF VIROLOGY, 1992, 66 (04) :2232-2239