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Miniaturized and High-Throughput Assays for Analysis of T-Cell Immunity Specific for Opportunistic Pathogens and HIV

Miniaturized and High-Throughput Assays for Analysis of T-Cell Immunity Specific for... Miniaturized and High-Throughput Assays for Analysis of T-Cell Immunity Specific for Opportunistic Pathogens and HIV Giuseppina Li Pira a , Federico Ivaldi b , Nadia Starc a , Fabiola Landi a , Franco Locatelli a , Sergio Rutella a , Gino Tripodi c and Fabrizio Manca d a Bambino Gesù Children's Hospital, Rome, Italy b Advanced Biotechnology Center, Genoa, Italy c G. Gaslini Institute, Genoa, Italy d MASIR Foundation, Rotterdam, The Netherlands T. S. Alexander , Editor ABSTRACT Monitoring of antigen-specific T-cell responses is valuable in numerous conditions that include infectious diseases, vaccinations, and opportunistic infections associated with acquired or congenital immune defects. A variety of assays that make use of peripheral lymphocytes to test activation markers, T-cell receptor expression, or functional responses are currently available. The last group of assays calls for large numbers of functional lymphocytes. The number of cells increases with the number of antigens to be tested. Consequently, cells may be the limiting factor, particularly in lymphopenic subjects and in children, the groups that more often require immune monitoring. We have developed immunochemical assays that measure secreted cytokines in the same wells in which peripheral blood mononuclear cells (PBMC) are cultured. This procedure lent itself to miniaturization and automation. Lymphoproliferation and the enzyme-linked immunosorbent spot (ELISPOT) assay have been adapted to a miniaturized format. Here we provide examples of immune profiles and describe a comparison between miniaturized assays based on cytokine secretion or proliferation. We also demonstrate that these assays are convenient for use in testing antigen specificity in established T-cell lines, in addition to analysis of PBMC. In summary, the applicabilities of miniaturization to save cells and reagents and of automation to save time and increase accuracy were demonstrated in this study using different methodological approaches valuable in the clinical immunology laboratory. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Clinical and Vaccine Immunology American Society For Microbiology

Miniaturized and High-Throughput Assays for Analysis of T-Cell Immunity Specific for Opportunistic Pathogens and HIV

Miniaturized and High-Throughput Assays for Analysis of T-Cell Immunity Specific for Opportunistic Pathogens and HIV

Clinical and Vaccine Immunology , Volume 21 (4): 488 – Apr 1, 2014

Abstract

Miniaturized and High-Throughput Assays for Analysis of T-Cell Immunity Specific for Opportunistic Pathogens and HIV Giuseppina Li Pira a , Federico Ivaldi b , Nadia Starc a , Fabiola Landi a , Franco Locatelli a , Sergio Rutella a , Gino Tripodi c and Fabrizio Manca d a Bambino Gesù Children's Hospital, Rome, Italy b Advanced Biotechnology Center, Genoa, Italy c G. Gaslini Institute, Genoa, Italy d MASIR Foundation, Rotterdam, The Netherlands T. S. Alexander , Editor ABSTRACT Monitoring of antigen-specific T-cell responses is valuable in numerous conditions that include infectious diseases, vaccinations, and opportunistic infections associated with acquired or congenital immune defects. A variety of assays that make use of peripheral lymphocytes to test activation markers, T-cell receptor expression, or functional responses are currently available. The last group of assays calls for large numbers of functional lymphocytes. The number of cells increases with the number of antigens to be tested. Consequently, cells may be the limiting factor, particularly in lymphopenic subjects and in children, the groups that more often require immune monitoring. We have developed immunochemical assays that measure secreted cytokines in the same wells in which peripheral blood mononuclear cells (PBMC) are cultured. This procedure lent itself to miniaturization and automation. Lymphoproliferation and the enzyme-linked immunosorbent spot (ELISPOT) assay have been adapted to a miniaturized format. Here we provide examples of immune profiles and describe a comparison between miniaturized assays based on cytokine secretion or proliferation. We also demonstrate that these assays are convenient for use in testing antigen specificity in established T-cell lines, in addition to analysis of PBMC. In summary, the applicabilities of miniaturization to save cells and reagents and of automation to save time and increase accuracy were demonstrated in this study using different methodological approaches valuable in the clinical immunology laboratory.

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References (21)

Publisher
American Society For Microbiology
Copyright
Copyright © 2014 by the American society for Microbiology.
ISSN
1556-6811
eISSN
1556-679X
DOI
10.1128/CVI.00660-13
pmid
24477854
Publisher site
See Article on Publisher Site

Abstract

Miniaturized and High-Throughput Assays for Analysis of T-Cell Immunity Specific for Opportunistic Pathogens and HIV Giuseppina Li Pira a , Federico Ivaldi b , Nadia Starc a , Fabiola Landi a , Franco Locatelli a , Sergio Rutella a , Gino Tripodi c and Fabrizio Manca d a Bambino Gesù Children's Hospital, Rome, Italy b Advanced Biotechnology Center, Genoa, Italy c G. Gaslini Institute, Genoa, Italy d MASIR Foundation, Rotterdam, The Netherlands T. S. Alexander , Editor ABSTRACT Monitoring of antigen-specific T-cell responses is valuable in numerous conditions that include infectious diseases, vaccinations, and opportunistic infections associated with acquired or congenital immune defects. A variety of assays that make use of peripheral lymphocytes to test activation markers, T-cell receptor expression, or functional responses are currently available. The last group of assays calls for large numbers of functional lymphocytes. The number of cells increases with the number of antigens to be tested. Consequently, cells may be the limiting factor, particularly in lymphopenic subjects and in children, the groups that more often require immune monitoring. We have developed immunochemical assays that measure secreted cytokines in the same wells in which peripheral blood mononuclear cells (PBMC) are cultured. This procedure lent itself to miniaturization and automation. Lymphoproliferation and the enzyme-linked immunosorbent spot (ELISPOT) assay have been adapted to a miniaturized format. Here we provide examples of immune profiles and describe a comparison between miniaturized assays based on cytokine secretion or proliferation. We also demonstrate that these assays are convenient for use in testing antigen specificity in established T-cell lines, in addition to analysis of PBMC. In summary, the applicabilities of miniaturization to save cells and reagents and of automation to save time and increase accuracy were demonstrated in this study using different methodological approaches valuable in the clinical immunology laboratory.

Journal

Clinical and Vaccine ImmunologyAmerican Society For Microbiology

Published: Apr 1, 2014

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