Argos has made extensive progress in automating a closed, GMP- compliant system for DC manufacturing. This system involves a computer-controlled workstation that links a GAMBRO Elutra device (to harvest monocytes from pheresis products) to closed-system cell culture process. All media changes and washes are automated and limited operator intervention is required throughout the entire process. A second computer-controlled work station harvests the matured DC, electroporates them with RNA, and aliquots, vials, and freezes individual vaccine doses. Argos has consulted wi...

This platform with its enhanced GMP facilities for production of clinical-grade DC is established in Erlangen DE (P6, Schuler / Schuler-Thurner). However, later it was decided to seek a licence for the production of GMP-grade mRNA that will be used for the (revised) multi-centre clinical trial.

We have compiled and evaluated all published information on clinical trials with DC worldwide. Questionnaires to all authors concerning follow-up of their published patients are about to be sent out. This database will be made available for all DC-THERA participants and will serve as a basis for a critical report on the role of DC based immunotherapy in cancer 2009.

In march 2009 we obtained the new BD ARIA II, which not only allows 14 parameter flow cytometry, but also sorting of even very small antigen specific T cell subpopulations for further molecular analysis in close collaboration with Pierre Coulie´s group in Brussels. [PhD student Steve Voland]

We have a confocal microscope available. This microscope was is used, for instance, for the characterization of antigen depots.

An assay for measuring cytotoxic T lymphocytes was used for the monitoring of T cell activation in the evaluation of the immunotherapy potential of DC pulsed with tumor antigens.

There is know-how on classical MLR assays in our lab. They are used, for instance, for characterization of silenced cells.

In our laboratory, we have experience in performing CBA assays. We use them, e.g., in the monitoring of vaccine induced T cell responses.

A confocal microscope is available in our laboratory.

We have experience in querying DAVID, the Database for Annotation, Visualization and Integrated Discovery (http://david.abcc.ncifcrf.gov/) . For instance, we used DAVID for functional annotation of upregulated and downregulated genes by gene ontology classification and pathway analysis by several bioinformatics approaches. Preliminary results obtained from querying DAVID and Ontologizer (http://www.charite.de/ch/medgen/ontologizer/) show statistically significant enrichment for immunological-relevant signaling pathways and immunology-related GO terms of the upregula...

Our laboratory has a FACS Canto II (Manufacturer: BD Biosciences - URL: http://www.bdbiosciences.com/features/products/display_product.php?keyID=594) at its disposal. For instance, we used multimers kindly provided by Pierre Coulie (Brussels) together with surface markers (such as CD8, CD4, CD3, CD45RA, CCR7, CD25, CD27, CD137, CD127) in different 8-color panels on this FACSCanto II.

We have a FACS Aria cell sorter at our disposition from BD Biosciences (URL: http://www.bdbiosciences.com/features/products/display_product.php?keyID=53). We used this FACS Aria among other things for fluorescence activated cell sorting for proteomic investigations. Using Fluorescence activated cell sorting allowed us to obtain highly pure fractions of CD8+, CD4+ and DN DC subsets. However, because DC are a very rare population, this method is very material- and cost-intensive. Furthermore, it is not surprising that the obtained cell numbers of each subset after isola...

There is a confocal microscope at our disposal for confocal imaging.

The Challenge of Cancer: Individual Cancers and Mutations Every individual´s cancer is unique. Therefore, the most effective treatment is one customized to a particular cancer´s genetic configuration. Genetic mutations and chromosomal abnormalities are commonly associated with human cancers. Unfortunately, since the genetic mutations leading to the development of cancer are often random events, every patient's tumor can contain a unique repertoire of antigens. This characteristic of human cancer requires that each patient's immune system be able to recognize and ...

We have know-how on assays to measure the proportions of Treg cells. For instance, we set up an assay to measure the proportions of Treg cells in tissues or cells that is based on the demethylation of a region of the FOXP3 promoter, which is observed in human Treg cells but not in the other T cells that can express FOXP3 transiently after activation. Starting from DNA, two quantitative PCR are used, which amplify the methylated and unmethylated FOXP3 promoter sequences. The ratio between the expression levels is, in our hands, a faithful Treg marker. We will pursue...

Led by Figdor (P2), this platform was established between four centres that respectively provide expertise and facilities for a variety of high resolution imaging techniques and MRI of humans in Nijmegen NL (P2, Figdor); MRI of mice in Erlangen DE (P6, Lutz); two photon microscopy in Paris FR (P13, Amigorena); and correlation spectroscopy in Marseilles FR (P22, Pierre). The platform organised several high quality courses and workshops in 2006-7 which continued in 2008.

We have developed an automated microarray data analysis (AMDA) software, which provides scientists with an easy and integrated system for the analysis of Affymetrix microarray experiments. AMDA is free and it is available as an R package. It is based on the Bioconductor project that provides a number of powerful bioinformatics and microarray analysis tools. This automated pipeline integrates different functions available in the R and Bioconductor projects with newly developed functions. AMDA covers all of the steps, performing a full data analysis, including image anal...

In our laboratory we perform 51Cr-release assays, for instance in the monitoring of vaccine induced T cell responses.

In our institute, we have a 9-colour FACS cell sorter our disposal. It is used, e.g., for the analysis of various DC populations.

We have expertise in performing toll-like receptor (TLR)-driven luciferase reporter assays. For example, in a study on the activation of DC we applied a TLR-driven luciferase reporter assay which showed dose-dependent activation of TLR2, TLR3, and TLR8, which was independent of the pseudotype, production, or transduction protocol and was abrogated on heat inactivation.