The NHLBI PEGT Cell Morphology Core (PEGT-CMC) at the University of Pennsylvania provides histopathology services free of charge to NHLBI-supported investigators active in gene transfer research. The PEGT-CMC evaluates viral vector preparations by electron microscopy and quantitates vector (reporter gene) marking in tissue sections using immunohistochemistry, or in situ hybridization. 

For vector analysis, five microliter aliquots are placed on copper grids, stained and examined under an electron microscope. Photographs are taken, developed and printed within the PEGT-CMC. Results are typically available within five days from receipt of the sample, and investigators can receive either the hard copy photographs or digital computer files. Written evaluations are also available.

Vectors are assessed for average density, purity, uniformity and viral particle integrity. For adeno-associated vectors (AAV), contaminating adenoviral particles are noted and AAV particles are assessed for conformation (ring vs. solid particles). Adenoviral particles are assessed for icosahedral shape, penton fiber formation, aggregation and incomplete capsid formation. These basic parameters can be applied to any viral preparation.

For tissue analysis, paraffin, frozen and plastic-embedded samples are used. Routine and special stains, as well as apoptosis and cell proliferation assays, are performed regularly. Immunohistochemistry is performed with a Ventana automated system, using all appropriate positive and negative controls. In situ hybridization for identification of transduced cells uses labeled DNA oligonucleotides. Routine electron microscopy is performed to evaluate phenotypic correction, and immuno-electron microscopy is used for specific subcellular localization of transgenic proteins using gold-labeled antibodies. 

Light microscopic images are taken with a digital camera and can be sent to investigators electronically or on a disc. Image analysis can be performed, quantifying the degree of staining for any target. Slides and tissues are returned to investigators as soon as the work order is completed.

Completion times vary according to each individual project. Vector validation, paraffin sections and frozen sections can usually be completed within two weeks of receipt. GMA specimen preparation and sectioning takes four weeks, and EM specimen preparation and sectioning can take 6 weeks. Development of new immunohistochemistry and in situ hybridization protocols depend on the individual project.

Contact Dr. Peter Bell at pbell@mail.med.upenn.edu for more information.

To apply for services, please download the pdf application form (requires Adobe Acrobat reader) or the Microsoft Word application form and fax to 212-746-8824. An online form will soon be available. Applications will be reviewed on a rolling basis.