Download Topic: “Flow cytometric measurement of intracellular proteins”

Topic: Flow cytometric measurement of intracellular proteins
Andrzej Eljaszewicz, MSc
Form of classes: 1h tutorial, 3h practical clasess
Self-study topics:
1. Phenotypic and functional characteristic of human peripheral blood: lymphocytes (T
cells and B cells), monocytes, NK cells and granulocytes.
2. The presentation of antigen to T lymphocytes –MHC class I restriction; MHC class II
restriction, characteristic of superantigens
3. Markers of the cell activation
Basic literature:
Janeway’s Immunobiology.
available online at
http://www.ncbi.nlm.nih.gov/books/NBK10757/ (free)
http://www.ncbi.nlm.nih.gov/sites/entrez?db=books (free)
Additional literature:
1. Manjula Reddy, et al; “Comparative analysis of lymphocyte activation
marker expression and cytokine secretion profile in stimulated human
peripheral blood mononuclear cell culture: an in vitro model to monitor
cellular immune function” Journal of Immunological Methods 2004.
293:127-142
2. Marzio R. et al.; „CD69 and regulation of the immune function“;
Immunopharmacology and immunotoxicology 1999. 21:565-582
3. Claman Prussin, Dean D. Metcalfe; “Detection in intracytoplasmic
cytokine using flow cytometry and directly conjugated anti-cytokine
antibodies”; Journal of Immunological Methods 1995. 188:117-128
4. Sewell W.A., et al.; ”Determination of intracellular cytokines by flowcytometry following whole-blood culture”; Journal of Immunological
Methods 1997. 209:67-74
5. Beate Mascher; et al.; “Expression and kinetic of cytokines
determinated by intracellular staining using flow cytometry”; Journal
of Immunological Methods 1999. 223:115-121
Practical skills:
Students know how to:
1. stain cells for intracellular markers with the fixation and permeabilization reagents
(methanol and saponine based methods)
2. analyze flow cytometric data obtained from prepared samples
Form of crediting:
1. At the beginning of the classes a 10-point quiz will be presented to students to test
their understanding of the material from the current week (self study topics)
2. Participation in practical exercises.