Voie d'immunisation et séquence d'administration de l ... - TEL

tel-00827710, version 1 - 29 May 2013
sufficient (Seder et al., 2008). It has become clear that, more than the quantity of antigen-
specific T cells that are generated upon immunization, it is their functional qualities that are
reflective of their responsiveness and need to be analyzed.
CD8 + T cell responses directed against a pathogen or elicited by efficient vaccines are usually
robust enough to allow for the detection and characterization of antigen-specific T cells ex
vivo without performing an enrichment. However, in some cases, such as for the study of
naïve T cells or low frequency populations, an enrichment step may be necessary to
sufficiently dissect a functionally effective immune response. It is important to note that
tetramer-based enrichment can be also applied to the study of T cell populations in humans.
For this purpose, the tetramer-based enrichment strategy has been developed and optimized
for the study of human samples in our lab (Alanio et al., 2010). This method allowed for the
detection and characterization of antigen-specific, naive T cells in the blood of healthy
donors. Interestingly, it was shown that specific precursor frequencies are conserved between
human donors, as it has been observed in mice. Using similar techniques, one group has
demonstrated that in chronically HCV-infected patients, immunodominance is correlated to
the naïve, HCV-specific precursor T cell frequency (Schmidt et al., 2011).
3) Limitations
Despite the clear advantages of the tetramer-based enrichment system, there are also
limitations, mostly technical in nature. The protocol for enrichment and staining of T cells is
extensive and time consuming, thus fewer samples can be processed concurrently. Although
this approach allows for the characterization of smaller populations, there is a limit to the
number of conditions that can feasibly be compared in the same experiment. Moreover, in the
case that an experiment requires the use of transgenic cells or to follow CFSE-labeled specific
cells in vivo, it remains extremely difficult to combine the techniques and follow both the
transferred cells and the endogenous repertoire. Nevertheless, using the tetramer-based
enrichment to follow transferred cells only, does allow the transfer of fewer transgenic T
cells, which maintains “physiologic” conditions in many cases.
Additional limitations exist when trying to compare other approaches with results about
endogenous repertoire obtained after enrichment. For example, techniques such as in vivo
imaging are not yet sensitive enough to look at the low number of cells that can be detected
by tetramers and still require the transfer of high amounts of TCR-transgenic T cells in order
to have a consistent read-out. These concerns must be taken into account when combining
different approaches to address a question.
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