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Sections of Infectious Diseases and Immunobiology, Yale University School of Medicine, New Haven, CT 06520
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The role of the TCR repertoire in determining the magnitude of a T cell
response is unknown. Approaching this issue has been difficult because
of the complexity of Ag-specific T cell populations and the difficulty
of identifying, isolating, and characterizing Ag-specific T cells ex
vivo (11). TCR analyses of T cell clones specific for the same epitope
demonstrated highly diverse repertoires in some cases (12, 13, 14) and
restriction to specific TCR 
- and ß-chains in others (15, 16, 17, 18, 19).
While it is clear that positive and negative selection determine the
ultimate number of naive peripheral T cells specific for a particular
Ag, the relationship between naive precursor frequencies and
immunodominance has not been established. The role of T cell avidities
for different MHC/peptide complexes is also unknown.
CD8+ cytolytic T cells (CTL) play an important role in the host immune defense against Listeria monocytogenes, a Gram-positive intracellular bacterium that enters the cytoplasm of infected cells (20, 21). Intravenous infection of mice with a subethal dose of L. monocytogenes primes specific CTL, which participate in clearing the infection and mediate long-lasting, protective immunity (22). Four different L. monocytogenes epitopes are presented by MHC class I H2-Kd molecules to CTL (23, 24, 25, 26). These epitopes derive from bacterial virulence factors that are expressed and degraded in the cytosol of infected cells. Remarkably, there is no correlation between the amount of epitope that is presented by infected cells and the magnitude of the CTL response. Thus, LLO 91–99, a relatively sparsely presented epitope, elicits a quantitatively large T cell response, while p60 449–457, which is presented in large quantities, elicits a small response (27).
In this report, we investigate differences between two dominant and two subdominant T cell responses following L. monocytogenes infection. We find that short term, in vitro T cell expansion with peptide-coated stimulator cells provides misleading results when comparing dominant with subdominant T cell responses. Immunodominant peptides inhibit T cell expansion at conventional peptide concentrations and stimulate maximal T cell proliferation at significantly lower concentrations than subdominant peptides. The optimal peptide concentration for stimulating T cell expansion correlates inversely with peptide-induced stabilization of surface H2-Kd molecules, suggesting that MHC/peptide stability is an important determinant for immunodominance. Using optimal in vitro restimulation conditions, we find that CTL lines specific for dominant epitopes express diverse TCR-Vß chains, while subdominant peptide-specific CTL lines use a restricted range of TCR-Vß chains. Thus, our findings suggest that peptides that form stable complexes with MHC molecules elicit dominant T cell responses characterized by a diverse TCR repertoire.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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CB6 ((C57BL/6 x BALB/c)F1; H2bxd) mice were obtained from The Jackson Laboratory (Bar Harbor, ME). L. monocytogenes strain 10403S was purchased from the American Type Culture Collection (ATCC, Rockville, MD) and grown in brain-heart infusion broth.
Mastocytoma cell line P815 (H2d) was obtained from ATCC, and H2-Kd-transfected RMA-S cells were kindly provided by M. Bevan (University of Washington, Seattle, WA). Cells were cultured in RP10+, consisting of RPMI 1640 medium (Life Technologies, Gaithersburg, MD) supplemented with 10% FCS, L-glutamine, HEPES (pH 7.5), ß-mercaptoethanol, penicillin (100 U/ml), streptomycin (100 µg/ml), and gentamicin (50 µg/ml).
Immunization with Listeria and harvesting spleen cells
Mice were immunized by injecting 2 x 103 L. monocytogenes 10403S into the tail vein. Spleens were removed 7 days after immunization, and splenocytes were harvested by dissociation through a wire mesh and lysis of erythrocytes with ammonium chloride, and subsequently resuspended in RP10+.
Peptides
Synthetic peptides LLO 91–99 (GYKDGNEYI), p60 217–225 (KYGVSVQDI), p60 449–457 (IYVGNGQMI), and mpl 84–92 (GYLTDNDEI) were obtained from Research Genetics (Huntsville, AL). Peptide stock solutions (1 mg/ml dissolved in PBS) were stored at -20 C until use.
IFN-
ELISPOT assays
ELISPOT assays were performed as described previously (27).
Briefly, 96-well nitrocellulose plates (Millititer HA, Millipore,
Bedford, MA) were coated with anti-mouse IFN- mAb (PharMingen,
San Diego, CA) and then blocked with medium containing 10% FCS. To
determine in vivo frequencies for epitope-specific
CD8+ T-cells, 1 x 105 spleen cells
from an immunized mouse and 1 x 105 peptide-coated
irradiated P815 cells were added per well in a total volume of 200 µl
RP10+ containing 30 U/ml IL-2. For each peptide,
measurements were performed at least in triplicates, and for no-peptide
controls uncoated P815 were used. Cells were incubated for 27 h at
37°C; thereafter, nitrocellulose membranes were washed with
PBS/0.05% Tween-20, incubated with biotinylated anti-mouse IFN-
mAb (PharMingen), and developed using peroxidase-labeled streptavidin
(Kirkegaard and Perry Laboratories, Gaithersburg, MD) and DAB substrate
(Bio-Rad, Melville, NY). The areas where a single cell was stimulated
to secrete IFN- can be easily detected as a spot on the
nitrocellulose membrane using a dissecting microscope. Frequencies of
peptide-specific T cells were expressed as the number of
IFN--secreting cells per 105 splenocytes.
For ELISPOT assays of T cell lines, T cell cultures were used directly or following serial 1:5 dilutions, and added in a volume of 100 µl to wells.
In vitro peptide stimulation and generation of T cell lines
Spleen cells (3–4 x 107) from immunized mice were resuspended in 5 ml RP10+ and added to a T 25 cell culture flask. Syngeneic splenocyte stimulators were prepared by irradiation (3000 rad) and pulsing for 1 h at 37°C with the appropriate peptide. Thereafter, cells were washed to remove free unbound peptide, and 3 x 107 cells in 5 ml RP10+ were added to the responder cells. For peptide titration experiments, cells were stimulated in 24-well plates incubating 1 x 107 responder cells and 1 x 107 peptide-pulsed stimulator cells in 2 ml RP10+.
To generate T cell lines, spleen cells from immunized mice were stimulated with the appropriate peptide in flasks as described and then restimulated weekly by adding 3 x 107 fresh peptide-coated stimulator cells. Following the second restimulation, the medium was supplemented with 10 U/ml IL-2.
CTL assay
Standard chromium release assays were performed as described (28). Briefly, 1 x 104 51Cr sodium chromate-labeled P815 cells were added as target cells per well of a 96-well tissue culture plate and tested for lysis by effector cells in the presence of peptide (10-6 M). For experiments analyzing short term, peptide-stimulated T cell lines, 2 ml were taken from a 10-ml total volume of cell cultures, pelleted, and resuspended in 300 µl RP10+. Then, 100 µl of this cell suspension, directly or from serial 1:3 dilutions, was added to target cells. The percentage of specific lysis was calculated as previously described (28). For peptide titrations, assays were performed in triplicates at a constant E:T ratio of approximately 10:1.
RMA-S-Kd assay
The assay was performed as described recently (29). RMA-S Kd cells were grown in serum-free HL-1 medium (BioWhittaker, Walkersville, MD) containing 400 µg/ml G418 (Life Technologies, Grand Island, NY). 1 x 106 cells were incubated overnight at 37°C in the presence of 30 µM synthetic LLO 91–99, p60 217–225, p60 449–457, mpl 84–92 or without peptide. Cells were washed three times with PBS to remove free peptide and resuspended in fresh serum-free medium. After 0, 2, 4, and 6 h, samples were taken and immediately stained for H2-Kd-expression, using the conformation dependent mAb SF1-1.1.1 (obtained from ATCC) and FITC-conjugated goat anti-mouse IgG (PharMingen) as the secondary Ab. Staining was performed in ice-cold PBS containing 0.5% BSA and 0.02% sodium azide, and cells were immediately fixed in 1% paraformaldehyde after staining. Expression of H2-Kd was determined by FACS analysis (see below).
Flow cytometry and TCR-Vß chain analysis
Five days after the third in vitro peptide stimulation, T cell
lines were examined for expression of different TCR-Vß segments.
Cells (2 x 105) were added per well of a 96-well
tissue culture plate and stained with mAbs specific for 13 different
Vß segments (Vß 2, 3, 4, 5.1/2, 6, 7, 8.1/2/3, 9, 10, 11, 12, 13,
14; all obtained from PharMingen). Simultaneously, cells were stained
with phycoerythrin-conjugated anti-CD8 and biotinylated
anti-CD62L mAbs (PharMingen). After 30 min of incubation, cells
were washed twice in staining buffer, incubated for 30 min with
Cy-Chrome-conjugated streptavidin (PharMingen), and then washed three
times. Three-color flow cytometry was performed using a FACScan flow
cytometer and further analyzed with CELLQuest software (Becton
Dickinson, Mountain View, CA).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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A conventional method for characterizing CTL following infection
is to restimulate primed splenocytes in vitro with peptide-coated,
irradiated, naive splenocytes and to assay for peptide-specific
cytotoxic activity 5 days later in a standard chromium release assay.
We restimulated L. monocytogenes-immune splenocytes with
irradiated, naive spleen cells coated with 10-6 M LLO
91–99, p60 217–225, p60 449–457, and mpl 84–92 and tested
responders for specific lysis using peptide-coated P815
(H2d) cells. When equivalent proportions of different
responders from the original cell cultures were tested, we obtained
different degrees of % specific lysis for the various epitopes.
Responders restimulated with mpl 84–92 and p60 217–225 showed a high
percentage of specific lysis, while LLO 91–99- and p60
449–457-restimulated responders gave a lower specific lysis (Fig. 1
, a and c).
Although it is well known that the amount of specific lysis correlates
with the ratio of effector (CTL) to target cells, we wanted to be
certain that the different degrees of lysis obtained with CTL
stimulated with the four H2-Kd-restricted epitopes could be
attributed to different numbers of responding CTL and not to
differences in the lytic capacity of the different responder
populations. Because Ag-specific T cells constitute only a small
fraction of the total cellular content of short term, in vitro
restimulations, we quantified Ag-specific T cells in responder flasks
by ELISPOT. The number of IFN--secreting, peptide-specific T cells
in flasks restimulated with each of the four
H2-Kd-restricted epitopes correlates precisely with the
specific lysis obtained in CTL assays (Fig. 1b).
Thus, the differences in specific lysis that we observe can be
attributed to the number of CTL following in vitro peptide
restimulation.
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The frequency of T cells specific for LLO 91–99, p60
217–225, and p60 449–457 1 wk following L. monocytogenes
infection was previously found by ELISPOT to be roughly 80, 40, and 4
per 100,000 immune splenocytes (27). Because the frequency of T
lymphocytes specific for the recently described mpl 84–92 epitope was
unknown, we performed ELISPOT frequency analysis on immune splenocytes
for this epitope and the previously characterized
H2-Kd-restricted epitopes. Surprisingly, the frequency of
mpl 84–92-specific T cells is very low and is comparable to p60
449–457-specific responses (Fig. 2). As
shown previously, the level of responses to LLO 91–99 and p60 217–225
is high. Thus, as determined by direct frequency analysis, the relative
magnitude of the T lymphocyte responses can be ranked as follows: LLO
91–99 > p60 217–225 >> p60 449–457 = mpl 84–92. These
frequencies contrast dramatically with the results obtained by in vitro
peptide restimulation (Fig. 1
, a and b),
indicating that LLO 91–99-specific T cells proliferate poorly in
vitro, while at the other extreme, mpl 84–92-specific T cells undergo
far greater in vitro expansion.
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Different in vitro expansion rates may result from intrinsic
differences between responding T cells with differing peptide
specificities. Perhaps vigorous in vivo activation and expansion of LLO
91–99-specific CTL interferes with further in vitro expansion
following peptide restimulation. It is possible that the majority of
these cells are programmed to undergo cell death, since it is well
known that most of the primary effector cells rapidly disappear as a
result of apoptosis or other mechanisms (30). To determine whether this
is the case, we stimulated immune splenocytes nonspecifically with the
mitogen Con A (37) and tested responders 5 days later with CTL and
ELISPOT assays for their peptide specificity. As shown in Figure 3, a and b,
mitogenic stimulation of immune splenocytes expands epitope-specific T
cell populations in proportions that reflect the immunodominance
hierarchy observed in vivo. Even small differences, such as the lower
number of p60 217–225-specific CTL as compared with LLO 91–99, are
conserved. Thus, CTL specific for immunodominant epitopes do not have a
diminished ability to expand in vitro. This suggests that some aspect
of in vitro peptide restimulation causes disparate expansion of CTL
specific for different epitopes.
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Recent studies have demonstrated that in vitro expansion of some T
cells is inhibited when they are restimulated with high peptide
concentrations (31, 32). To determine whether the peptide concentration
that we used to stimulate T cells inhibited their in vitro expansion,
we coated naive splenocytes with a range of peptide concentrations
before in vitro restimulation of immune splenocytes. We found
significant differences in the optimal peptide concentrations required
for maximal in vitro CTL expansion (Fig. 4). T cells specific for immunodominant
epitopes expanded most vigorously with lower peptide coating
concentrations (10-9 M for LLO 91–99 and
10-8 M for p60 217–225), while CTL specific for
subdominant epitopes responded best when stimulators were coated with a
100- to 1,000-fold higher peptide concentration (10-6 M
for p60 449–457 and mpl 84–92). Notably, T cells specific for
subdominant epitopes did not undergo in vitro expansion in response to
peptide concentrations of 10-8 or 10-9 M.
Thus, when immune splenocytes are stimulated with high concentrations
of dominant and subdominant peptides, T cells responding to subdominant
peptides have a selective advantage and, following short term
restimulation, can appear to be more prevalent than dominant T cell
populations.
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The observation that T cells specific for immunodominant
epitopes expand most vigorously when lower peptide concentrations are
used for stimulation prompted us to determine the stability of
MHC/peptide complexes with each of the four
H2-Kd-restricted epitopes. RMA-S-Kd cells,
which lack functional TAP2 molecules, express negligible levels of
surface H2-Kd in the absence of exogenous peptide. We
incubated RMA-S-Kd cells in the presence of each of the
L. monocytogenes-derived epitopes and monitored the
stability of surface H2-Kd molecules using the
conformation-dependent, H2-Kd-specific Ab SF1-1.1.1 and
flow cytometry. All four peptides stabilized surface expression of
H2-Kd nearly equivalently (Fig. 5), and during the first 2 h after
peptide removal there was rapid loss of SF1-1.1.1-reactive
H2-Kd, perhaps resulting from rapid dissociation of
incompletely bound epitopes. As we have shown in previous studies (29),
LLO 91–99 and p60 217–225 stabilize H2-Kd more
effectively than does p60 449–457 between 2 and 6 h after peptide
removal (Fig. 5). Remarkably, for mpl 84–92 we observe a rapid loss of
SF1-1.1.1-reactive H2-Kd (Fig. 5), indicating that mpl
84–92, like p60 449–457, dissociates from H2-Kd rapidly,
while LLO 91–99 and p60 217–225 dissociate slowly. In previous
studies, we used biochemical approaches to determine that the
t1/2 of H2-Kd complexes with LLO
91–99 or p60 217–225 is 
6 h (29). In contrast, the
t1/2 of H2-Kd complexes with p60
449–457 is 1 h. Since mpl 84–92 stabilizes surface
H2-Kd slightly more effectively than p60 449–457, but much
less effectively than LLO 91–99 or p60 217–225, we estimate the
t1/2 of mpl 84–92/H2-Kd complexes
to be in the range of 1 to 2 h. These findings point to a
correlation between MHC/peptide stability and the peptide concentration
required for optimal in vitro T cell expansion.
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It is not known whether T cell populations specific for
dominant and subdominant epitopes have different avidities for their
respective epitope/MHC complexes. To determine whether we select for
high avidity CTL following in vitro expansion with very low peptide
concentrations, we tested different epitope-specific CTL lines for
their relative peptide sensitivities. T cell lines were generated by in
vitro restimulation of immune splenocytes with naive splenocytes coated
with each of the epitopes at the concentration that supported optimal
proliferation (10-6 M for mpl 84–92 and p60
449–457, 10-8 for p60 217–225, and 10-9 for
LLO 91–99). The relative ability of CTL lines to lyse target cells in
the presence of decreasing concentrations of each of the peptides was
determined (Fig. 6). CTL lines specific
for all four epitopes lysed target cells maximally in the presence of
their respective peptide at concentrations as low as 10-10
M. The similar peptide sensitivities of the generated CTL lines suggest
that the avidity of in vitro-challenged CTL lines specific for each of
the L. monocytogenes-derived epitopes is similar. This
finding is noteworthy because markedly different peptide concentrations
were used for in vitro restimulation. We attempted to obtain CTL lines
specific for LLO 91–99 and p60 217–225 by restimulation with higher
peptide concentrations; however, these lines uniformly could not be
maintained beyond two or three restimulations. This indicates that the
optimal peptide concentrations determined for short term restimulation
are neccessary for long term, in vitro expansion.
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Although the number and range of naive, Ag-specific T cells may be
important determinants of the ultimate response magnitude to a
particular epitope, few studies have compared the TCR diversity of CTL
specific for dominant and subdominant epitopes. We decided, therefore,
to stain the four epitope-specific CTL lines with mAbs specific for 13
different Vß TCR chains. Short term, in vitro peptide-restimulated
CTL lines were generated and consisted of >90% specific CTL, as
determined by H2-Kd tetramer staining (D.H.B. and
E.G.P, unpublished results). As shown in Figure 7, we found remarkable differences
between the TCR-Vß usage of T cell lines specific for immunodominant
vs subdominant epitopes: T cell lines specific for both immunodominant
epitopes show an extremely diverse TCR-Vß repertoire, whereas T cell
lines specific for the subdominant epitopes express far more restricted
TCR-Vß segments. In the case of mpl 84–92-specific T cells,
essentially all TCRs contain the Vß14 chain. Although the restriction
of TCRs specific for p60 449–457 is less stringent, a predominance of
TCR-Vß8 segments is readily and reproducibly detected.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Because of the low frequency of Ag-specific CTL following most
infections, studies of CTL generally require in vitro expansion to
obtain sufficient numbers of cells. One common method for expanding T
lymphocytes involves in vitro peptide restimulation. This method has
been used to generate T cell lines and clones and has also been used to
determine the frequencies of epitope-specific T cells by limiting
dilution (6, 7, 25). Our studies demonstrate very clearly that the
hierarchy of T cell responses following infection cannot be readily
determined with assays that depend on in vitro T cell proliferation
following peptide restimulation. Our finding that epitopes that
stimulate in vitro T cell proliferation at very low concentrations form
stable complexes with H2-Kd is unlikely to be coincidental.
Stimulator cells coated with peptides that form stable complexes may
form more surface complexes, and perhaps more significantly, these
complexes will persist on the cell surface for much longer periods of
time. In the case of LLO 91–99 and p60 217–225, which dissociate from
H2-Kd with a t1/2 of 6 h, it is
likely that immunologically relevant numbers of
H2-Kd/peptide complexes will be present on peptide-coated
stimulator cells much longer than p60 449–457- and mpl
84–92-containing complexes, which decay much more rapidly. Recent
studies have shown that T cell lines specific for an HIV-derived
epitope are inhibited by and undergo activation-induced cell death in
response to in vitro restimulation with high peptide concentrations
(31). We see a similar inhibition of in vitro expansion of LLO
91–99-specific CTL upon stimulation with high peptide concentrations.
Interestingly, the response to p60 217–225, which is also dominant, is
not detectably inhibited by restimulation with high peptide
concentrations. The basis for this difference between LLO 91–99 and
p60 217–225 is unknown. Our findings extend previous work in this area
by demonstrating: 1) CTL with different peptide specificities have
distinct responses to changes in peptide concentrations used for in
vitro restimulation; 2) CTL specific for subdominant epitopes require
high peptide concentrations for optimal in vitro expansion; and 3) CTL
specific for dominant epitopes require lower concentrations of peptide
for optimal in vitro expansion and can be inhibited by high peptide
concentrations.
Analysis of in vitro-expanded T cell lines revealed a highly
diverse TCR-Vß repertoire for immunodominant-specific lines, whereas
subdominant epitopes appeared to elicit far more restricted TCR-Vß
repertoire. These findings are similar to a recent analysis of
EBV-specific T cell clones isolated from patients with persistent EBV
infection (34). Comparison of multiple TCR sequences demonstrated a
highly diverse TCR repertoire for T cells specific for an
immunodominant EBV epitope, whereas a more restricted TCR repertoire
was detected when T cell clones were specific for a subdominant EBV
epitope. Interestingly, these differences were also maintained on the
level of the TCR-Vß segments. In other experimental systems, a rather
limited TCR repertoire diversity had been described for immunodominant
T cell responses (15, 16, 17). However, most of the Ags investigated in
these studies share homology with self proteins, and a possible role
for self homologues in determining the size and quality of the selected
TCR repertoire for these particular Ags has been suggested (35).
Although our approach only determined how many different Vß-chains
are expressed on CTL lines specific for the different L.
monocytogenes epitopes, our study has the advantage of
characterizing the majority of T cells (the 13 Vß chain-specific Abs
should detect 90% of TCR-ß-expressing cells) that respond to
each of the epitopes, without the need for cloning. Analyzing the
TCR-Vß usage alone does not allow a definitive statement about the
TCR repertoire, since there are clearly additional layers of diversity
at the level of CDR3 (complementarity-determining region 3) length and
pairing with different TCR -chains (36). Nevertheless, the TCR-Vß
differences between dominant and subdominant epitopes are striking and
are consistent with work in other infectious systems (13). Given these
results, it is tempting to attribute T cell response sizes to a
relatively greater repertoire of T cells in immunodominant responses
and a more restricted repertoire in subdominant responses. However, the
correlation between MHC/peptide stability and immunodominance is not
readily explained by this model.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Eric G. Pamer, Sections of Infectious Diseases and Immunobiology, Yale University, New Haven, CT 06520. E-mail address:
3 Abbreviations used in this paper: TAP, transporter associated with Ag processing; LLO, listeriolysin O; mpl, metalloprotease; ELISPOT, enzyme-linked immunospot.
Received for publication October 3, 1997. Accepted for publication December 22, 1997.
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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 D. Meyer-Olson, N. H. Shoukry, K. W. Brady, H. Kim, D. P. Olson, K. Hartman, A. K. Shintani, C. M. Walker, and S. A. Kalams Limited T Cell Receptor Diversity of HCV-specific T Cell Responses Is Associated with CTL Escape J. Exp. Med., August 2, 2004; 200(3): 307 - 319. [Abstract] [Full Text] [PDF]
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 W. Zhong, P. A. Reche, C.-C. Lai, B. Reinhold, and E. L. Reinherz Genome-wide Characterization of a Viral Cytotoxic T Lymphocyte Epitope Repertoire J. Biol. Chem., November 14, 2003; 278(46): 45135 - 45144. [Abstract] [Full Text] [PDF]
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 Z. Li, C. M. Diaz-Montero, G. Valbuena, X.-J. Yu, J. P. Olano, H.-M. Feng, and D. H. Walker Identification of CD8 T-Lymphocyte Epitopes in OmpB of Rickettsia conorii Infect. Immun., July 1, 2003; 71(7): 3920 - 3926. [Abstract] [Full Text] [PDF]
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Y. Nakamura, T. Suda, T. Nagata, T. Aoshi, M. Uchijima, A. Yoshida, K. Chida, Y. Koide, and H. Nakamura Induction of Protective Immunity to Listeria monocytogenes with Dendritic Cells Retrovirally Transduced with a Cytotoxic T Lymphocyte Epitope Minigene Infect. Immun., April 1, 2003; 71(4): 1748 - 1754. [Abstract] [Full Text]
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S. Oh, J. W. Hodge, J. D. Ahlers, D. S. Burke, J. Schlom, and J. A. Berzofsky Selective Induction of High Avidity CTL by Altering the Balance of Signals from APC J. Immunol., March 1, 2003; 170(5): 2523 - 2530. [Abstract] [Full Text] [PDF]
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K. Ogata, A. Jaramillo, W. Cohen, J.-P. Briand, F. Connan, J. Choppin, S. Muller, and S. J. Wodak Automatic Sequence Design of Major Histocompatibility Complex Class I Binding Peptides Impairing CD8+ T Cell Recognition J. Biol. Chem., January 3, 2003; 278(2): 1281 - 1290. [Abstract] [Full Text] [PDF]
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P. M. Gray, G. D. Parks, and M. A. Alexander-Miller High Avidity CD8+ T Cells Are the Initial Population Elicited Following Viral Infection of the Respiratory Tract J. Immunol., January 1, 2003; 170(1): 174 - 181. [Abstract] [Full Text] [PDF]
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S. E. Hamilton and J. T. Harty Quantitation of CD8+ T Cell Expansion, Memory, and Protective Immunity After Immunization with Peptide-Coated Dendritic Cells J. Immunol., November 1, 2002; 169(9): 4936 - 4944. [Abstract] [Full Text] [PDF]
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R. J. DiPaolo and E. R. Unanue Cutting Edge: Chemical Dominance Does Not Relate to Immunodominance: Studies of the CD4+ T Cell Response to a Model Antigen J. Immunol., July 1, 2002; 169(1): 1 - 4. [Abstract] [Full Text] [PDF]
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F. Rodriguez, S. Harkins, M. K. Slifka, and J. L. Whitton Immunodominance in Virus-Induced CD8+ T-Cell Responses Is Dramatically Modified by DNA Immunization and Is Regulated by Gamma Interferon J. Virol., March 27, 2002; 76(9): 4251 - 4259. [Abstract] [Full Text] [PDF]
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M. Skoberne and G. Geginat Efficient In Vivo Presentation of Listeria monocytogenes- Derived CD4 and CD8 T Cell Epitopes in the Absence of IFN-{gamma} J. Immunol., February 15, 2002; 168(4): 1854 - 1860. [Abstract] [Full Text] [PDF]
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L. Grode, M. Kursar, J. Fensterle, S. H. E. Kaufmann, and J. Hess Cell-Mediated Immunity Induced by Recombinant Mycobacterium bovis Bacille Calmette-Guerin Strains Against an Intracellular Bacterial Pathogen: Importance of Antigen Secretion or Membrane-Targeted Antigen Display as Lipoprotein for Vaccine Efficacy J. Immunol., February 15, 2002; 168(4): 1869 - 1876. [Abstract] [Full Text] [PDF]
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M. Skoberne, R. Holtappels, H. Hof, and G. Geginat Dynamic Antigen Presentation Patterns of Listeria monocytogenes-Derived CD8 T Cell Epitopes In Vivo J. Immunol., August 15, 2001; 167(4): 2209 - 2218. [Abstract] [Full Text] [PDF]
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J. Choppin, W. Cohen, A. Bianco, J.-P. Briand, F. Connan, M. Dalod, and J.-G. Guillet Characteristics of HIV-1 Nef Regions Containing Multiple CD8+ T Cell Epitopes: Wealth of HLA-Binding Motifs and Sensitivity to Proteasome Degradation J. Immunol., May 15, 2001; 166(10): 6164 - 6169. [Abstract] [Full Text] [PDF]
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T. Yamada, H. Uchiyama, T. Nagata, M. Uchijima, T. Suda, K. Chida, H. Nakamura, and Y. Koide Protective Cytotoxic T Lymphocyte Responses Induced by DNA Immunization against Immunodominant and Subdominant Epitopes of Listeria monocytogenes Are Noncompetitive Infect. Immun., May 1, 2001; 69(5): 3427 - 3430. [Abstract] [Full Text] [PDF]
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G. Geginat, S. Schenk, M. Skoberne, W. Goebel, and H. Hof A Novel Approach of Direct Ex Vivo Epitope Mapping Identifies Dominant and Subdominant CD4 and CD8 T Cell Epitopes from Listeria monocytogenes J. Immunol., February 1, 2001; 166(3): 1877 - 1884. [Abstract] [Full Text] [PDF]
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K. M. Kerksiek, D. H. Busch, and E. G. Pamer Variable Immunodominance Hierarchies for H2-M3-Restricted N-Formyl Peptides Following Bacterial Infection J. Immunol., January 15, 2001; 166(2): 1132 - 1140. [Abstract] [Full Text] [PDF]
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 V. P. Badovinac, A. R. Tvinnereim, and J. T. Harty Regulation of Antigen-Specific CD8+ T Cell Homeostasis by Perforin and Interferon-gamma Science, November 17, 2000; 290(5495): 1354 - 1357. [Abstract] [Full Text]
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L. M. Mylin, T. D. Schell, D. Roberts, M. Epler, A. Boesteanu, E. J. Collins, J. A. Frelinger, S. Joyce, and S. S. Tevethia Quantitation of CD8+ T-Lymphocyte Responses to Multiple Epitopes from Simian Virus 40 (SV40) Large T Antigen in C57BL/6 Mice Immunized with SV40, SV40 T-Antigen-Transformed Cells, or Vaccinia Virus Recombinants Expressing Full-Length T Antigen or Epitope Minigenes J. Virol., August 1, 2000; 74(15): 6922 - 6934. [Abstract] [Full Text]
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J. V. Spencer and T. J. Braciale Incomplete Cd8+ T Lymphocyte Differentiation as a Mechanism for Subdominant Cytotoxic T Lymphocyte Responses to a Viral Antigen J. Exp. Med., May 15, 2000; 191(10): 1687 - 1698. [Abstract] [Full Text] [PDF]
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Q.-J. Zhang, S. S. Chen, C.-A. Saari, M. G. Massuci, F. Tufaro, and W. A. Jefferies Evidence of Selective Processing of Immunodominant Epitopes in Virally Infected Cells J. Immunol., May 1, 2000; 164(9): 4513 - 4521. [Abstract] [Full Text] [PDF]
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S. Yang, D. Kittlesen, C. L. Slingluff Jr., C. E. Vervaert, H. F. Seigler, and T. L. Darrow Dendritic Cells Infected with a Vaccinia Vector Carrying the Human gp100 Gene Simultaneously Present Multiple Specificities and Elicit High-Affinity T Cells Reactive to Multiple Epitopes and Restricted by HLA-A2 and -A3 J. Immunol., April 15, 2000; 164(8): 4204 - 4211. [Abstract] [Full Text] [PDF]
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R. Holtappels, D. Thomas, J. Podlech, G. Geginat, H.-P. Steffens, and M. J. Reddehase The Putative Natural Killer Decoy Early Gene m04 (gp34) of Murine Cytomegalovirus Encodes an Antigenic Peptide Recognized by Protective Antiviral CD8 T Cells J. Virol., February 15, 2000; 74(4): 1871 - 1884. [Abstract] [Full Text]
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D. E. Byers and K. F. Lindahl Peptide Affinity and Concentration Affect the Sensitivity of M3-Restricted CTLs Induced In Vitro J. Immunol., September 15, 1999; 163(6): 3022 - 3028. [Abstract] [Full Text] [PDF]
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M. E. Wallace, R. Keating, W. R. Heath, and F. R. Carbone The Cytotoxic T-Cell Response to Herpes Simplex Virus Type 1 Infection of C57BL/6 Mice Is Almost Entirely Directed against a Single Immunodominant Determinant J. Virol., September 1, 1999; 73(9): 7619 - 7626. [Abstract] [Full Text]
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L. J. McHeyzer-Williams, J. F. Panus, J. A. Mikszta, and M. G. McHeyzer-Williams Evolution of Antigen-specific T Cell Receptors In Vivo: Preimmune and Antigen-driven Selection of Preferred Complementarity-determining Region 3 (CDR3) Motifs J. Exp. Med., June 7, 1999; 189(11): 1823 - 1838. [Abstract] [Full Text] [PDF]
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C. Couedel, M. Bodinier, M.-A. Peyrat, M. Bonneville, F. Davodeau, and F. Lang Selection and Long-Term Persistence of Reactive CTL Clones During an EBV Chronic Response Are Determined by Avidity, CD8 Variable Contribution Compensating for Differences in TCR Affinities J. Immunol., June 1, 1999; 162(11): 6351 - 6358. [Abstract] [Full Text] [PDF]
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S. Vijh, I. M. Pilip, and E. G. Pamer Noncompetitive Expansion of Cytotoxic T Lymphocytes Specific for Different Antigens during Bacterial Infection Infect. Immun., March 1, 1999; 67(3): 1303 - 1309. [Abstract] [Full Text] [PDF]
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D. H. Busch and E. G. Pamer T Cell Affinity Maturation by Selective Expansion during Infection J. Exp. Med., February 15, 1999; 189(4): 701 - 710. [Abstract] [Full Text] [PDF]
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H. G. A. Bouwer, H. Shen, X. Fan, J. F. Miller, R. A. Barry, and D. J. Hinrichs Existing Antilisterial Immunity Does Not Inhibit the Development of a Listeria monocytogenes-Specific Primary Cytotoxic T-Lymphocyte Response Infect. Immun., January 1, 1999; 67(1): 253 - 258. [Abstract] [Full Text] [PDF]
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M. Tsuji, C. C. Bergmann, Y. Takita-Sonoda, K.-I. Murata, E. G. Rodrigues, R. S. Nussenzweig, and F. Zavala Recombinant Sindbis Viruses Expressing a Cytotoxic T-Lymphocyte Epitope of a Malaria Parasite or of Influenza Virus Elicit Protection against the Corresponding Pathogen in Mice J. Virol., August 1, 1998; 72(8): 6907 - 6910. [Abstract] [Full Text] [PDF]
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D. H. Busch, I. Pilip, and E. G. Pamer Evolution of a Complex T Cell Receptor Repertoire during Primary and Recall Bacterial Infection J. Exp. Med., July 1, 1998; 188(1): 61 - 70. [Abstract] [Full Text] [PDF]
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