Imaging and cell count in cleared intact cochlea in the Mongolian gerbil using laser scanning confocal microscopy.
Eur Ann Otorhinolaryngol Head Neck Dis
; 134(4): 221-224, 2017 Sep.
Article
en En
| MEDLINE
| ID: mdl-28196606
ABSTRACT
OBJECTIVES:
To draw up a clearing protocol for Mongolian gerbil cochlea, and to assess the feasibility of quantifying and analyzing 3D cell architecture in the transparent cochleae. MATERIALS ANDMETHODS:
Freshly dissected inner ears were prepared on a 13-day protocol fixation, microdissection, post-fixation, decalcification, pretreatment (signal enhancement, permeabilization and blocking), fluorescent labeling (indirect immunolabeling and direct labeling), dehydration, clearing in Spalteholz solution (MSBB methyl salicylate and benzyl benzoate) and mounting. Image acquisition used laser scanning confocal microscopy. ImageJ software was used to measure the length of the organ of Corti thus available for analysis and to count inner and outer hair cells.RESULTS:
Four cochleas underwent imaging. 3D reconstruction enabled organ of Corti length to be measured, at a mean 1269±346µm. Mean inner and outer hair-cell count per organ of Corti length was 142±44 and 400±122, respectively.CONCLUSION:
Cochlear clearing by MSBB was feasible in Mongolian gerbils and provided high-resolution immunofluorescence-labeled inner-ear images. To our knowledge, this was the first application of the technique in this species. Cell count could thus be performed along the organ of Corti length without traumatic dissection.Palabras clave
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Base de datos:
MEDLINE
Asunto principal:
Cóclea
/
Microscopía Confocal
Límite:
Animals
Idioma:
En
Revista:
Eur Ann Otorhinolaryngol Head Neck Dis
Año:
2017
Tipo del documento:
Article