The Centre of Excellence on Neurodegenerative Diseases which will host the training platform in animal and cell engineering is a multidisciplinary research center composed by a network of laboratories working at the University of Milano. The Centre has generated two major core facilities dedicated to "Genetic, cell and animal engineering" and "Functional imaging".

The Milano platform has the capability to develop protocols for the manipulation and the cloning of DNA sequences and is specialised in the development of transgenic vectors for eucaryotic cells ans animals; in particular, these vestors are studied in order to analyse the functional image. The laboratory also studies genetic expression and has basic bio-informatic programs available for the design of specific oligonucleotides, and for the analysis of databases and the results of analyses with microchips. In the sphere of these activities, both consultancy and technical training services are available.

The laboratory has acquired remarkable experience in the engineering of nerve stem cells as well as differentiated nerve cells with conventional and conditional expression vectors for the study of genetic functions and cellular transplant. Fluorescence images are obtained by performing immunofluorescence experiments or transfection with construct coding for fluorescent proteins (e.g., Green fluorescent Protein,etc.). Images of "in vivo" samples are obtained using a thermostatic video camera which allows the samples to be kept "alive" even for the long periods of recording necessary for time-lapse video microscopy experiments. All acquisition parameters and necessary commands (for the microscope as well as the digital camera and the rotating filter carrier) for the different acquistion protocols are controlled by MetaMorph software. One examples of the use of this sytem of video microcopy are the possibility to observe cytoskeletal filaments, and intracellular organelles like mitochondria, as well as granules of secretion and vesicles of transport departing from the Golgi complex that have been made fluorescent unsing antibodies and/or performing transfection experiments. This observation can take place both in static and in dynamic mode depending on the character of the samples be they fixed or "in vivo". Of particular relevance is the possibility to alternate the observation and recording of images obtained in light and in interferential contrast with images obtained in fluorescence.

The research unit specialised in the generation of trangenic animals has been active since May 2002. The service of generation of transgenic mice is available for all CEND research scientics. The technique of embryonic re-derivation is in preparation. We therefore have all the instrumentation necessary for microijection of cells. In addition, the CEND has reinforced this equipment with:

• Two "Versadoc" for the aquisition and the quantitative analysis of data attained both from gels for nucleic acids and filters; thus it is no longer necessary to use a dark room. Phosphoimagers for the aquisition of data attained from filters prepared with radioactively labelled samples. OCD video camera for the analyses of luminescent and fluorescent samples. Moreover, a dark room is available where this CCD video camera can be used for the quantification of luminescence and flurescence in tissues and "in toto" animals.
• Two real time PCR for the quantitative analysis of cDNA;
• An instrument for the sequencing of DNA that can be used at the Departement of Neurological Sciences (responsible: Giacomo Comi).

One of the aims of this laboratoy is the diffusion of technological know-how and, therefore, it offers refresher courses about technologies used in this lab for students, young graduates and technicians. For longer stays in this lab, grants are available.

The training will be given to groups of max 5 trainees. The entire training activities might take from 2 full man.months and 1 month of laboratory time for the first year, involving both postdoc and technician staff time, to 6 full man.months and 3 months of laboratory time for the following years as trainings sessions will be progressively added over the duration of the project.