Replication competent molecular clones of HIV-1 expressing Renilla luciferase facilitate the analysis of antibody inhibition in PBMC.
Effective vaccine development for human immunodeficiency virus type 1 (HIV-1) will require assays that ascertain the capacity of vaccine immunogens to elicit neutralizing antibodies (NAb) to diverse HIV-1 strains. To facilitate NAb assessment in peripheral blood mononuclear cell (PBMC)-based assays, we developed an assay-adaptable platform based on a Renilla luciferase (LucR) expressing HIV-1 proviral backbone. LucR was inserted into pNL4-3 DNA, preserving all viral open reading frames. The proviral genome was engineered to facilitate expression of diverse HIV-1 env sequences, allowing analysis in an isogenic background. The resulting Env-IMC-LucR viruses are infectious, and LucR is stably expressed over multiple replications in PBMC. HIV-1 neutralization, targeting TZM-bl cells, was highly correlative comparing virus (LucR) and cell (firefly luciferase) readouts. In PBMC, NAb activity can be analyzed either within a single or multiple cycles of replication. These results represent advancement toward a standardizable PBMC-based neutralization assay for assessing HIV-1 vaccine immunogen efficacy.
Edmonds
Tara G
TG
Department of Molecular and Cellular Pathology, University of Alabama at Birmingham, 701 19th Street South, Birmingham, AL 35294, USA. edmondst@uab.edu
Ding
Haitao
H
Yuan
Xing
X
Wei
Qing
Q
Smith
Kendra S
KS
Conway
Joan A
JA
Wieczorek
Lindsay
L
Brown
Bruce
B
Polonis
Victoria
V
West
John T
JT
Montefiori
David C
DC
Kappes
John C
JC
Ochsenbauer
Christina
C
D016915:AIDS Vaccines / Q000276:immunology; D057134:Antibodies, Neutralizing / Q000276:immunology; D015483:HIV Antibodies / Q000276:immunology*; D015497:HIV-1 / Q000235:genetics / Q000254:growth & development* / Q000276:immunology; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000821:virology*; D049410:Luciferases, Renilla / Q000096:biosynthesis* / Q000235:genetics; D009500:Neutralization Tests / Q000379:methods; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D013194:Staining and Labeling / Q000379:methods*; D014773:Virology / Q000379:methods*; D014779:Virus Replication
D016915:AIDS Vaccines; D057134:Antibodies, Neutralizing; D015483:HIV Antibodies; D011994:Recombinant Proteins; D049410:Luciferases, Renilla