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Evaluation of vascular endothelial staining by various antibodies in human and xenograft tumors using three different methods of fixation
Author(s) -
Breckenkamp Ann E,
Mintze Karen,
Barnes Versie,
Vance Tasha,
Pierson Nicholas,
Pangallo Jenna,
Sandusky George
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.1006.8
Subject(s) - staining , stain , pathology , antibody , fixative , paraformaldehyde , cd31 , immunohistochemistry , cd34 , fixation (population genetics) , chemistry , biology , microbiology and biotechnology , medicine , immunology , biochemistry , stem cell , gene
Light microscopy was used to compare the staining of various antibodies against the vascular endothelial markers CD31, CD34, CD105, EphB2, and EphB4 in human and xenograft tissues that were fixed in three different fixatives: 4% paraformaldehyde (PF), 10% Neutral Buffered Formalin (NBF), or Zinc‐Tris (ZT). Since the ability of an antibody to bind to its target molecule depends on the fixation method, it would be useful to identify antibodies that stain tumor vasculature regardless of which fixative was used to preserve the tissue. Each antibody evaluated was optimized in ZT fixed tissues, and the protocol was used to stain all samples, except for two antibodies that did not stain ZT fixed tissues. Bethyl Laboratories Rabbit Anti‐Human CD31 (#IHC‐00055) and Cell Signaling Mouse Anti‐Human CD34 (#3569) gave excellent staining in human tissues prepared in 4% PF, 10% NBF, and ZT, in contrast to the other antibodies evaluated. The Bethyl antibody also gave moderate staining in HCT116 xenografts prepared in all three fixatives, making it the only antibody evaluated that was useful for staining vasculature in human breast, colon, and lung tissues and xenografts for all three fixatives.