Characterization of monoclonal antibodies against the glaucoma-associated protein myocilin

Mohamed Karim Ezzat, Kyle G. Howell, Cindy K. Bahler, Thomas G. Beito, Nils Loewen, Eric M. Poeschla, Michael P Fautsch

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

Although the glaucoma-associated protein myocilin has been the focus of intensive research, its biological function is still unknown. One of the limiting factors has been the lack of well-characterized antibodies, particularly monoclonal antibodies. We describe the development of six monoclonal antibodies specific to myocilin and characterize their suitability in Western blot and immunohistochemical applications. Three of the six monoclonal antibodies recognize the N-terminus of myocilin (amino acids 33-214), two antibodies recognize the middle third of the protein (amino acids 215-368), and one antibody recognizes the C-terminus (amino acids 369-504). Isotyping revealed that all antibodies are of the IgG1κ class except one, which is IgG2bκ. Purified myocilin monoclonal antibodies were able to recognize myocilin in human aqueous humor separated on denatured/reduced and native gels, and human trabecular meshwork lysate by Western blot. Myocilin was also detected by immunohistochemistry in trabecular meshwork, ciliary body, iris, cornea, sclera, choroid, and retinal pigment epithelial cells.

Original languageEnglish (US)
Pages (from-to)376-384
Number of pages9
JournalExperimental Eye Research
Volume87
Issue number4
DOIs
StatePublished - Oct 2008

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Glaucoma
Monoclonal Antibodies
Trabecular Meshwork
Proteins
Amino Acids
Antibodies
Western Blotting
Ciliary Body
Sclera
Retinal Pigments
Choroid
Aqueous Humor
Immunoglobulin Isotypes
Iris
Cornea
trabecular meshwork-induced glucocorticoid response protein
Immunoglobulin G
Gels
Epithelial Cells
Immunohistochemistry

Keywords

  • aqueous humor
  • glaucoma
  • monoclonal antibodies
  • myocilin
  • TIGR
  • trabecular meshwork

ASJC Scopus subject areas

  • Ophthalmology
  • Cellular and Molecular Neuroscience
  • Sensory Systems

Cite this

Characterization of monoclonal antibodies against the glaucoma-associated protein myocilin. / Ezzat, Mohamed Karim; Howell, Kyle G.; Bahler, Cindy K.; Beito, Thomas G.; Loewen, Nils; Poeschla, Eric M.; Fautsch, Michael P.

In: Experimental Eye Research, Vol. 87, No. 4, 10.2008, p. 376-384.

Research output: Contribution to journalArticle

Ezzat, Mohamed Karim ; Howell, Kyle G. ; Bahler, Cindy K. ; Beito, Thomas G. ; Loewen, Nils ; Poeschla, Eric M. ; Fautsch, Michael P. / Characterization of monoclonal antibodies against the glaucoma-associated protein myocilin. In: Experimental Eye Research. 2008 ; Vol. 87, No. 4. pp. 376-384.
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