Advertisement

Immunohistochemical Staining of Cyclooxygenases with Monoclonal Antibodies

  • Ghassan M. Saed
Part of the Methods In Molecular Biology book series (MIMB, volume 477)

Abstract

Immunohistochemistry is an important tool that is often used for the diagnosis of several diseases in the pathology laboratory. The quality and sensitivity of immunohistochemical staining is affected by formalin fixation, which results in variable loss of antigenicity, known as a masking effect. While the sensitivity of immunohistochemistry is excellent for certain antigens, other antigens such as COX-1 and COX-2 are difficult to identify, especially in formalin-fixed, paraffin sections. Antigen retrieval is a technique that re-exposes epitopes and allows detection of masked antigens with standard immunohistochemical procedures. One common method involves partial, enzymatic pre-digestion with trypsin or pepsin while other, nonenzymatic procedures or heat-mediated antigen retrieval methods include pressure-cookers, hot plates, or microwave (MW) irradiation of tissue sections in water or a variety of antigen-retrieval solutions. In this chapter, we will describe a technique that provides a more reliable, much simpler approach for the demonstration of cyclooxygenase-1 and cyclooxygenase-2 expression in frozen, vibratome or paraffin sections, and/or cells in cultures.

Key words

COX-1 COX-2 Immunofluorescence Post-operative adhesions Ovarian cancer 

References

  1. 1.
    Vane J. Towards a better aspirin. Nature 1994;367:215–6.CrossRefPubMedGoogle Scholar
  2. 2.
    DuBois RN, Awad J, Morrow J, Roberts LJ, 2nd, Bishop PR. Regulation of eicosanoid production and mitogenesis in rat intestinal epithelial cells by transforming growth factor-alpha and phorbol ester. J Clin Invest 1994;93:493–8.CrossRefPubMedGoogle Scholar
  3. 3.
    Kujubu DA, Fletcher BS, Varnum BC, Lim RW, Herschman HR. TIS10, a phorbol ester tumor promoter-inducible mRNA from Swiss 3T3 cells, encodes a novel prostaglandin synthase/cyclooxygenase homologue. J Biol Chem 1991;266:12866–72.PubMedGoogle Scholar
  4. 4.
    Chen G, Wilson R, McKillop JH, Walker JJ. The role of cytokines in the production of prostacyclin and thromboxane in human mononuclear cells. Immunol Invest 1994;23:269–79.CrossRefPubMedGoogle Scholar
  5. 5.
    Schwab JM, Schluesener HJ, Laufer S. COX-3: just another COX or the solitary elusive target of paracetamol? Lancet 2003;361:981–2.CrossRefPubMedGoogle Scholar
  6. 6.
    Shaftel SS, Olschowka JA, Hurley SD, Moore AH, O’Banion MK. COX-3: a splice variant of cyclooxygenase-1 in mouse neural tissue and cells. Brain Res Mol Brain Res 2003;119:213–5.CrossRefPubMedGoogle Scholar
  7. 7.
    Ockleford CD. An atlas of antigens: fluorescence microscope localisation patterns in cells and tissues. New York, NY: Stockton Press, 1990.Google Scholar
  8. 8.
    Kupper H, Storz H. Double staining technique using a combination of indirect and direct immunofluorescence with monoclonal antibodies. Acta Histochem 1986;78:185–8.PubMedGoogle Scholar
  9. 9.
    Wheatley SP, Wang YL. Indirect immunofluorescence microscopy in cultured cells. Methods Cell Biol 1998;57:313–32.CrossRefPubMedGoogle Scholar
  10. 10.
    Tornehave D, Hougaard DM, Larsson L. Microwaving for double indirect immunofluorescence with primary antibodies from the same species and for staining of mouse tissues with mouse monoclonal antibodies. Histochem Cell Biol 2000;113:19–23.CrossRefPubMedGoogle Scholar

Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • Ghassan M. Saed
    • 1
  1. 1.Departments of Obstetrics and Gynecology & Cell Biology and AnatomyWayne State University School of MedicineDetroitUSA

Personalised recommendations