A simple and efficient method for preparation of isolated ovarian follicles for transmission electron microscopy.
OBJECTIVE
A simple method for preparation of isolated ovarian follicles for transmission electron microscopy (TEM) using transwell inserts is described.


METHODS
Pre-antral follicles were enzymatically isolated from mouse ovaries and cultured overnight on transwell insert polyester membranes. The following day, isolated ovarian follicles were processed for TEM by moving the transwell insert through successive wells containing the fixation and embedding reagents. After polymerization of the resin, the polyester membrane with the follicles embedded in the resin was disengaged from the transwell unit. The resin was sectioned. Semi-thin sections were stained with toluidine blue while ultra-thin sections were stained by uranyl acetate and examined by light microscopy and TEM, respectively.


RESULTS
Isolated ovarian follicles were easily processed in groups for TEM. Follicles were well embedded and there appeared to be no loss of tissue during processing. The ultra-structure of processed isolated ovarian follicles was well preserved with little evidence of processing artifacts.


CONCLUSIONS
In situ processing and preparation of isolated ovarian follicles by first allowing attachment on transwell insert membranes was shown to be a simple, rapid and effective method for TEM.
Department of OB-GYN, Women's Health Institute, Cleveland Clinic Foundation, Cleveland, OH, USA. desain@ccf.org
Nina
N
Desai
Faten
F
AbdelHafez
Judith
J
Drazba
James
J
Goldfarb
Tommaso
T
Falcone
D000818:Animals; D002448:Cell Adhesion; D003433:Crosses, Genetic; D005260:Female; D008567:Membranes, Artificial; D051379:Mice; D008810:Mice, Inbred C57BL; D008854:Microscopy, Electron; D009924:Organ Culture Techniques / Q000295:instrumentation / Q000379:methods; D006080:Ovarian Follicle / Q000648:ultrastructure*; D011091:Polyesters; D013048:Specimen Handling; D013194:Staining and Labeling / Q000379:methods; D016610:Tissue Embedding / Q000379:methods; D016707:Tissue Fixation / Q000379:methods
D008567:Membranes, Artificial; D011091:Polyesters