A critical aspect of this program project includes evaluating antibodies for their potency against HIV and affinities to gpl 20s and Fc receptors. The Cell/Biochemical Assay Automation Core will design and carry out cell-based and biochemical assays to characterize the specificity, efficacy, and potency of HIV-neutralizing antibodies. We have adapted the standard in vitro pseudovirus neutralization assay, which measures the reduction in luciferase reporter gene expression in the presence of a potential inhibitor following pseudovirus infection in TZM-bl cells, for execution by a liquid handing robot. Core A personnel will validate, refine, and trouble-shoot protocols for the automated in vitro neutralization assays and then perform automated in vitro neutralization assays for evaluating antibodies for Drs. Nussenzweig, Ravetch, and Bjorkman. In addition. Core A will perform rapid and high-throughput binding assays to assess the affinities of antibodies for gpl20s and/or gp140s, as required for Drs. Nussenzweig and Bjorkman, and the affinities of modified Fc regions for Fc receptors, as required by Dr. Ravetch. The core will train and assist students, postdoctoral fellows, research assistants, and investigators in the analysis and interpretation of data from automated assays. In addition, the core will maintain two custom-equipped Evo Freedom Liquid handling stations, including daily calibration and care, and annual preventative maintenance.

Public Health Relevance

HIV/AIDS is a global public health problem. Efforts to develop a vaccine or improve antibodies for passive immunization require screening large numbers of potential reagents: using in vitro neutralization assays to evaluate antibody potencies and protein-protein binding assays to identify interactions. We have automated these techniques in order to efficiently evaluate large numbers of reagents for the investigators in this collaborative project.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Program Projects (P01)
Project #
5P01AI100148-05
Application #
9208099
Study Section
Special Emphasis Panel (ZAI1-RRS-A)
Project Start
Project End
Budget Start
2017-02-01
Budget End
2018-01-31
Support Year
5
Fiscal Year
2017
Total Cost
$213,643
Indirect Cost
$85,329
Name
California Institute of Technology
Department
Type
Domestic Higher Education
DUNS #
009584210
City
Pasadena
State
CA
Country
United States
Zip Code
91125
Wang, Haoqing; Barnes, Christopher O; Yang, Zhi et al. (2018) Partially Open HIV-1 Envelope Structures Exhibit Conformational Changes Relevant for Coreceptor Binding and Fusion. Cell Host Microbe 24:579-592.e4
Stadtmueller, Beth M; Bridges, Michael D; Dam, Kim-Marie et al. (2018) DEER Spectroscopy Measurements Reveal Multiple Conformations of HIV-1 SOSIP Envelopes that Show Similarities with Envelopes on Native Virions. Immunity 49:235-246.e4
Gautam, Rajeev; Nishimura, Yoshiaki; Gaughan, Natalie et al. (2018) A single injection of crystallizable fragment domain-modified antibodies elicits durable protection from SHIV infection. Nat Med 24:610-616
Cohn, Lillian B; da Silva, Israel T; Valieris, Renan et al. (2018) Clonal CD4+ T cells in the HIV-1 latent reservoir display a distinct gene profile upon reactivation. Nat Med 24:604-609
Bournazos, Stylianos; Ravetch, Jeffrey V (2017) Fc? Receptor Function and the Design of Vaccination Strategies. Immunity 47:224-233
Freund, Natalia T; Wang, Haoqing; Scharf, Louise et al. (2017) Coexistence of potent HIV-1 broadly neutralizing antibodies and antibody-sensitive viruses in a viremic controller. Sci Transl Med 9:
Mayer, Christian T; Gazumyan, Anna; Kara, Ervin E et al. (2017) The microanatomic segregation of selection by apoptosis in the germinal center. Science 358:
Wang, Haoqing; Gristick, Harry B; Scharf, Louise et al. (2017) Asymmetric recognition of HIV-1 Envelope trimer by V1V2 loop-targeting antibodies. Elife 6:
Horwitz, Joshua A; Bar-On, Yotam; Lu, Ching-Lan et al. (2017) Non-neutralizing Antibodies Alter the Course of HIV-1 Infection In Vivo. Cell 170:637-648.e10
Nishimura, Yoshiaki; Gautam, Rajeev; Chun, Tae-Wook et al. (2017) Early antibody therapy can induce long-lasting immunity to SHIV. Nature 543:559-563

Showing the most recent 10 out of 52 publications