...t the target of FcgammaRs with the antigen induces the internalization of the immune complex, leading to the maturation of CD11c+CD11b+ conventional DCs. These DCs become thus efficient to present antigenic peptides to helper CD4 T cells and cytotoxic CD8 T cells via cross-presentation pathway.

...vaccine induced specific anti-tumor T cell responses (thymidine-incorporation, ELISA, LDA and ELISPOT assays). We have set the staining conditions for the HLA-DR*1101 tetramers loaded with tetanus toxoid and MAGE-3 peptides corresponding promiscuous CD4+ T cell epitopes. Antigen-specific CD4+ T cell...

Monocytes were cultured in serum free medium in the presence of IL-4 and GM-CSF, and concentrated cytokines were added.

The cultured cells were subsequently pulsed with tumor antigen derived peptides (one per culture) at a concentration of 5 µg/mL in approximately 5 mL and at a cell density of 20 x 10e6/mL for 1 h.

The tumor antigen derived peptide pulsed monocyte-derived DC were diluted in medium containing 20 ng/mL TNF-?, plus IL-4 and GM-CSF.

The mature peptide pulsed DC were frozen at 5 x 10e6 cells per vial in freezing medium for subsequent stimulation of autologous T cells.

The autologous T lymphocytes were stimulated with (frozen) MoDC at a ratio of 1 DC per 10 T cells.

The majority of monocytes (1 x 10e9) were frozen in 90% A-plasma and 10% GMP-grade DMSO.

All lymphocytes (6,4 x 10e9) were frozen in 90% A-plasma and 10% GMP-grade DMSO.

The autologous DC loaded with KLH, as immunological tracer, and an allogeneic peptidome (i.e. natural tumour peptides, NTPs), obtained from melanoma cell line SK-Mel24, are used for vaccination.

The T blasts are used to immunize MAGE-3 positive patients....

From the CD14- PBMCs, untouched naive CD4+ T cells were isolated with a combination of magnetic sorting.

MoDC were stimulated with yeast.

The dead cells were stained and used in the uptake of dead cells by DC.

The D2SC1/Flt3-DC were stained (Mini67-1KT or Mini26-1KT (PKH67/PKH26 green/red fluorescent cell linker mini kit for general cell membrane labelling) from Sigma) for a process of dead cell uptake.

The dead cells were stained (Mini67-1KT or Mini26-1KT (PKH67/PKH26 green/red fluorescent cell linker mini kit for general cell membrane labelling) from Sigma) for a process of dead cell uptake by DC.

These stained were stained for a process of dead cell uptake by DC.

These cells were stained with anti-CD11c antibodies for a process of dead cell uptake by DC.

On these cells, transcriptional profiling was performed and a comparative pathway-based analysis was done.

These macrophages were used to perform a transcriptional profiling. Stimulation was done with different stimuli at different time points in the presence or absence of a kinase inhibitors.