Mapping T cell epitopes by flow cytometry

Hoffmeister, Bodo; Kiecker, Felix; Tesfa, Lydia; Volk, Hans Dieter; Picker, Louis J.; Kern, Florian

doi:10.1016/s1046-2023(02)00349-3

Cited by 69 publications

(61 citation statements)

References 11 publications

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“…Pol, we performed IFN-g ELISPOT assays using matrices of overlapping peptides as described previously (50)(51)(52). Consistent with other reports, the H-2K d -restricted epitope Gag 65-73 AMQMLKETI (Gag-AI9) was the immunodominant target derived from this protein in BALB/c mice (53).…”

Section: Resultsmentioning

confidence: 56%

Epitope Specificity Delimits the Functional Capabilities of Vaccine-Induced CD8 T Cell Populations

Hill

Darrah

Ende

et al. 2014

The Journal of Immunology

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Despite progress toward understanding the correlates of protective T cell immunity in HIV infection, the optimal approach to Ag delivery by vaccination remains uncertain. We characterized two immunodominant CD8 T cell populations generated in response to immunization of BALB/c mice with a replication-deficient adenovirus serotype 5 vector expressing the HIV-derived Gag and Pol proteins at equivalent levels. The Gag-AI9/H-2Kd epitope elicited high-avidity CD8 T cell populations with architecturally diverse clonotypic repertoires that displayed potent lytic activity in vivo. In contrast, the Pol-LI9/H-2Dd epitope elicited motif-constrained CD8 T cell repertoires that displayed lower levels of physical avidity and lytic activity despite equivalent measures of overall clonality. Although low-dose vaccination enhanced the functional profiles of both epitope-specific CD8 T cell populations, greater polyfunctionality was apparent within the Pol-LI9/H-2Dd specificity. Higher proportions of central memory-like cells were present after low-dose vaccination and at later time points. However, there were no noteworthy phenotypic differences between epitope-specific CD8 T cell populations across vaccine doses or time points. Collectively, these data indicate that the functional and phenotypic properties of vaccine-induced CD8 T cell populations are sensitive to dose manipulation, yet constrained by epitope specificity in a clonotype-dependent manner.

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Section: Resultsmentioning

confidence: 56%

Epitope Specificity Delimits the Functional Capabilities of Vaccine-Induced CD8 T Cell Populations

Hill

Darrah

Ende

et al. 2014

The Journal of Immunology

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“…Intracellular cytokine staining (ICS) was performed as previously described (8,9,34,43) with some minor modifications. Briefly, whole blood was collected in sodium heparin tubes and prepared for peptide stimulation either on the day of collection or after keeping blood at room temperature overnight.…”

Section: Methodsmentioning

confidence: 99%

Reduced Hepatitis B Virus (HBV)-Specific CD4⁺T-Cell Responses in Human Immunodeficiency Virus Type 1-HBV-Coinfected Individuals Receiving HBV-Active Antiretroviral Therapy

Chang

Wightman

Bartholomeusz

et al. 2005

J Virol

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Functional hepatitis B virus (HBV)-specific T cells are significantly diminished in individuals chronicallyinfected with HBV compared to individuals with self-limiting HBV infection or those on anti-HBV therapy. In individuals infected with human immunodeficiency virus type 1 (HIV-1), coinfection with HBV is associated with an increased risk of worsening liver function following antiviral therapy and of more rapid HBV disease progression. Total HBV-specific T-cell responses in subjects with diverse genetic backgrounds were characterized by using a library of 15-mer peptides overlapping by 11 amino acids and spanning all HBV proteins. The magnitude and breadth of CD4 ؉ and CD8 ؉ T-cell responses to HBV in peripheral blood were examined by flow cytometry to detect gamma interferon production following stimulation with HBV peptide pools. Chronic HBV carriers (n ‫؍‬ 34) were studied, including individuals never treated for HBV infection (n ‫؍‬ 7), HBV-infected individuals receiving anti-HBV therapy (n ‫؍‬ 13), and HIV-1-HBV-coinfected individuals receiving anti-HBV therapy (n ‫؍‬ 14). CD4؉ and CD8 ؉ HBV-specific T-cell responses were more frequently detected and the CD8 ؉ T-cell responses were of greater magnitude and breadth in subjects on anti-HBV treatment than in untreated chronic HBV carriers. There was a significant inverse correlation between detection of a HBVspecific T-cell response and HBV viral load. HBV-specific CD4؉ and CD8 ؉ T-cell responses were significantly (fivefold) reduced compared with HIV-specific responses. Although, the frequency and breadth of HBV-specific CD8 ؉ T-cell responses were comparable in the monoinfected and HIV-1-HBV-coinfected groups, HBV-specific CD4 ؉ T-cell responses were significantly reduced in HIV-1-HBV-coinfected individuals. Therefore, HIV-1 infection has a significant and specific effect on HBV-specific T-cell immunity.There are 350 million individuals chronically infected with hepatitis B virus (HBV) worldwide (36,42,59). Each year 1 million to 1.5 million carriers die from HBV-related liver disease and liver cancer such as hepatocellular carcinoma (36,48). In the United States, Europe, and Australia, approximately 6 to 7% of individuals infected with human immunodeficiency virus type 1 (HIV-1) are also coinfected with HBV (23, 41). Liver disease is now a major comorbidity in HIV-1-infected individuals (11,66). Although hepatitis C virus (HCV)-HIV-1 coinfection is more common than HBV infection, liver failure occurs more frequently with persistent HBV infection (60, 63). Coinfection of HBV with HIV-1 leads to elevated HBV DNA levels, a lower rate of seroconversion to HBeAg, and lower alanine aminotransferase (ALT) levels than those in HBV monoinfection (21, 31). Spontaneous flares and HBeAg seroconversion occur in 5% of HBV-monoinfected individuals; however, in the setting of HIV-1-HBV coinfection, spontaneous flares or seroconversion to HBeAg is rare (18). In individuals infected with HIV-1 who subsequently acquire HBV infection, there is an increased risk of persisten...

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“…In each tube, we used 10 l of complete medium containing a 1 g/ml final concentration of each peptide spanning the pp65 or IE-1 CMV protein. Intracellular cytokine detection was performed in accordance with the protocol recommended by JPT Peptide Technologies and was well-described (19), together with the measurement of CD107a Ag exposure (20). Briefly, 10 6 PBMC were placed in 15-ml conical polypropylene tubes (Corning) in a final volume of 500 l of complete medium and incubated with: 10 l of complete medium containing one test volume of each PepMix, 1 g each of the costimulatory mAbs CD28 and CD49d (BD Immunocytometry Systems) and a saturating amount of PE-Cy5-conjugated anti-CD107a mAb (BD Biosciences/BD Pharmingen).…”

Section: Subjectsmentioning

confidence: 99%

Massive Load of Functional Effector CD4+ and CD8+ T Cells against Cytomegalovirus in Very Old Subjects

Vescovini

Biasini

Fagnoni³

et al. 2007

The Journal of Immunology

153

111

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A progressive, systemic, and low-grade proinflammatory status is one of the major characteristics of immunosenescence. Emerging data suggest a possible contribution of CMV, known to chronically infect a large proportion of humans, lifelong from newborns to centenarians. To test this hypothesis, we evaluated functional T cell responses to two CMV immunogenic proteins, pp65 and IE-1, in 65 chronically infected subjects aged 25–100 years. PBMC were stimulated with mixtures of peptides spanning the entire sequence of both proteins, and Ag specificity and magnitude of intracellular IFN-γ- and TNF-α-positive cells were then analyzed within both CD4+ and CD8+ T cells. Results indicate that pp65 and, to a lesser extent, IE-1 constitute major Ags against which aged people target functionally efficient T cell effector responses with massive production of Th1 cytokines and exhibition of CD107a degranulation marker. As a result, the production of IFN-γ induced in T cells by both Ags was seven to eight times greater in very old than in young subjects. The comparative analysis of pp65-specific responses in these very long-term carriers revealed a reciprocal relationship between CD4+ and CD8+ producing IFN-γ in the same individuals. These results indicate that CMV represents an important pathogen responsible for a strong immune activation in human aging. Such a remarkable burden of effector CD4+ and CD8+ T cells may be necessary to protect the elderly from CMV endogenous reactivation, but can turn detrimental by giving a substantial contribution to the proinflammatory status that accompanies the main age-related diseases.

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Mapping T cell epitopes by flow cytometry

Cited by 69 publications

References 11 publications

Epitope Specificity Delimits the Functional Capabilities of Vaccine-Induced CD8 T Cell Populations

Epitope Specificity Delimits the Functional Capabilities of Vaccine-Induced CD8 T Cell Populations

Reduced Hepatitis B Virus (HBV)-Specific CD4⁺T-Cell Responses in Human Immunodeficiency Virus Type 1-HBV-Coinfected Individuals Receiving HBV-Active Antiretroviral Therapy

Massive Load of Functional Effector CD4+ and CD8+ T Cells against Cytomegalovirus in Very Old Subjects

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Mapping T cell epitopes by flow cytometry

Cited by 69 publications

References 11 publications

Epitope Specificity Delimits the Functional Capabilities of Vaccine-Induced CD8 T Cell Populations

Epitope Specificity Delimits the Functional Capabilities of Vaccine-Induced CD8 T Cell Populations

Reduced Hepatitis B Virus (HBV)-Specific CD4+T-Cell Responses in Human Immunodeficiency Virus Type 1-HBV-Coinfected Individuals Receiving HBV-Active Antiretroviral Therapy

Massive Load of Functional Effector CD4+ and CD8+ T Cells against Cytomegalovirus in Very Old Subjects

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Reduced Hepatitis B Virus (HBV)-Specific CD4⁺T-Cell Responses in Human Immunodeficiency Virus Type 1-HBV-Coinfected Individuals Receiving HBV-Active Antiretroviral Therapy