Document Detail


Effects of fixative and embedding medium on morphology and immunostaining of the cochlea.
MedLine Citation:
PMID:  18827478     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The localization of proteins by immunostaining is a powerful method to investigate otologic disorders. However, the use of fixatives and embedding media (necessary for the preservation of morphology) can obscure antigens, making it difficult to perform immunoassays. We performed a systematic investigation of the effects of fixative and embedding medium on morphology and immunostaining of the mouse cochlea. Three different fixative solutions [4% formaldehyde (F), 4% formaldehyde + 1% acetic acid (FA), and 4% formaldehyde + 1% acetic acid + 0.1% glutaraldehyde (FGA)] and 3 different embedding media (paraffin, polyester wax, and celloidin) were used. Morphology was assessed using light microscopy. Immunostaining was studied using a panel of 6 antibodies (to prostaglandin D synthase, aquaporin 1, connective tissue growth factor, 200-kDa neurofilament, tubulin and Na(+),K(+)-ATPase). Preservation of morphology was suboptimal with paraffin, adequate with polyester wax and superb with celloidin. Immunostaining was successful using all 6 antibodies in all 3 fixatives and all 3 embedding media. While there were differences in strength of signal and localization of antigen between the 3 fixatives, overall, FA and FGA gave the most uniform results. For a given fixative and antibody, there was surprisingly little difference in the quality of immunostaining between celloidin and paraffin, while results in polyester wax were not as good in some cases. These results suggest that celloidin may be the embedding medium of choice for both morphological and pathological studies, including immunostaining when morphology must be optimized.
Authors:
Jennifer T O'Malley; Saumil N Merchant; Barbara J Burgess; Diane D Jones; Joe C Adams
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural     Date:  2008-10-01
Journal Detail:
Title:  Audiology & neuro-otology     Volume:  14     ISSN:  1421-9700     ISO Abbreviation:  Audiol. Neurootol.     Publication Date:  2009  
Date Detail:
Created Date:  2009-01-26     Completed Date:  2009-03-05     Revised Date:  2014-09-24    
Medline Journal Info:
Nlm Unique ID:  9606930     Medline TA:  Audiol Neurootol     Country:  Switzerland    
Other Details:
Languages:  eng     Pagination:  78-87     Citation Subset:  IM    
Copyright Information:
2008 S. Karger AG, Basel.
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MeSH Terms
Descriptor/Qualifier:
Animals
Cochlea / cytology*,  metabolism
Collodion
Fixatives*
Formaldehyde*
Immunohistochemistry / methods*
Mice
Mice, Inbred CBA
Paraffin Embedding*
Polyesters
Proteins / metabolism
Temporal Bone / cytology,  metabolism
Waxes
Grant Support
ID/Acronym/Agency:
U24 DC0008559/DC/NIDCD NIH HHS; U24 DC008559/DC/NIDCD NIH HHS; U24 DC008559-02/DC/NIDCD NIH HHS
Chemical
Reg. No./Substance:
0/Fixatives; 0/Polyesters; 0/Proteins; 0/Waxes; 1HG84L3525/Formaldehyde; 9004-70-0/Collodion
Comments/Corrections

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