THE HUMAN PROTEIN ATLAS BLOG
Tissue Atlas: production and staining
Today, we start a "mini-series" about our Tissue Atlas here at the blog. Join us on a tour through the lab, meet some of the people working there, and see some really nice images produced by the scientists.
All the work on our Tissue Atlas is done at our Uppsala site, with Cecilia Lindskog as site director. You can learn all about her in one of our previous blog posts.
First we meet research engineer IngMarie Olsson who is group leader for the Tissue Microarray Production, Immunohistochemistry, and Scanning-group.
– I have been working in the project since 2003 and am now responsible for production of tissue microarrays, immunohistochemical staining and scanning of stained slides.
Tissue microarrays are used in the basic protein profiling for the Human Protein Atlas. Each protein expression profile is based on 8 tissue microarrays including normal tissues from 144 different individuals (triplicate samples of 44 different tissue types) and cancer tissues from 216 different patients (duplicate samples of tumor tissues representing the 20 most common forms of human cancer). In total, 576 tissue cores are immunostained and analyzed for each antibody. (For more information of our antibodies, and the production of them, see this blog post).
Formalin fixed, paraffin embedded tissue specimens are collected from the Department of Pathology, Uppsala University Hospital. Representative areas to include on the tissue microarrays are defined for each tissue specimen by visual inspection of a corresponding hematoxylin-eosin stained section under a microscope. Tissue cores from these selected areas are then removed and used for tissue microarray production.
IngMarie Olsson shows a prepared tissue microarray that consists of 72 tissue cores.
– After production the tissue microarray is cut into 4 micrometer thick sections that are placed on a special object glass, to ensure that the tissue attaches well. From one array you can get out about 250 sections, Ingmarie Olsson explains.
After drying the glass can be stored at -20C for up to two years, awaiting staining.
For the immunohistochemical staining, antibodies are titrated using a specially designed test-tissue microarray, representing a limited selection of tissues and cells. The primary working dilution is based on the protein concentration for each antibody.
– For this process we use instruments and commercially available detection kits to ensure standardization and reproducibility of the staining.
IngMarie Olsson is responsible for ensuring that there are arrays for all tissues at all times, a job that requires constant updating and database searching.
– I have to know what we have waiting in the freezer, how many arrays we have, and that the staining is in a good pace so that the persons doing the microscopy have just the right amount of work to do. When we run out of tissue to take from I have to find it in the pathology database, and go into their archive to find it.
At all times, IngMarie has the whole process in the back of her head, and once a month she prepares statistics to see how the lab is doing.
– Right now we have a buffer of arrays for about two years.
Stay tuned for next week´s blog post, where you will meet some of the people responsible for microscopy and annotation! And this Friday, Image of the Week will be from this group!
Frida Henningson Johnson