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2C - Biomarkers (TUP/96) Optimization of Tissue Preparation for the Quantification of Germinal Cells in Histologic Sections of Fathead Minnow Gonads. Dietrich, Daniel1, Wolf, Jeff2, Ellis, Rosanne3, Diamond, Steve4, Friederich, Urs5, Hall, Tilghman6, Klecka, Gary7, Staples, Charles8, 1 Environmental Toxicology, University of Konstanz, Konstanz, BW, Germany2 Experimental Pathology Laboratories, Sterling, VA, USA3 IPMC-TMC GmbH, Neuwilen, TG, Switzerland4 GE Plastics, Huntersville, NC, USA5 Dow Europe, Horgen, ZH, Switzerland6 Bayer Coorporation, Stillwell, KS, USA7 Dow Chemical Co., Midland, MI, USA8 Assessment Technologies Inc., Fairfax, VA, USA ABSTRACT- A study was undertaken to optimize tissue preparation for the differentiation and quantification of cell types in histologic sections of fathead minnow gonads. Sexually mature fish of both sexes were exposed to 10 nM 17- Key words: cell type quantification, Gonad histology, procedure optimization, fathead minnow |
-oestradiol for a period of 10 days. Gonads were excised by microdissection and processed using various fixation, embedding, sectioning and staining procedures. Optimal combinations were: Bouins solution fixation, paraffin embedding, haematoxylin/eosin (H&E) staining for the ovary and glutaraldehyde fixation, glycol methacrylate (GMA) embedding, H&E staining for the testis. The thin sectioning (0.5-1 
m) that was possible with GMA-embedded sections for the testicular tissue greatly enhanced the ability of the pathologist to accurately identify individual cell types and morphologic abnormalities. The histologic sections of ovary and testis were digitally photographed for image analysis. Criteria were established for manually tagging (counting and labeling) individual cells in sections of ovary (> 100 follicles/oocytes per ovary) and testis (> 2000 cells per testis). Compared to gonadal staging, the tagging of individual gonadal cell types is a more objective, and therefore more precise, measurement. In addition, tagging cells via the computer provides a permanent and reviewable record of each analysis. In conjunction with an overall histopathological evaluation of gonadal tissues, the differential quantification of cells in optimally prepared sections adds potentially useful information for the assessment of endocrine effects.