Affiliations: Whitaker Biomedical Imaging Laboratory, Department of Biomedical Engineering | Department of Clinical Pathology, The Cleveland Clinic Foundation, Cleveland, Ohio | Biomedical Engineering Center, The Ohio State University, Columbus, Ohio
Note: [] Corresponding author: D. Geoffrey Vince, Department of Biomedical Engineering, Mail Drop Wb3, The Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland, OH 44195, USA. Tel.: +1 216 444 1211; Fax: +1 216 444 9198; Email: vince@ bme.ri.ccf.org.
Abstract: Formaldehyde fixatives have traditionally been used to preserve tissues as they impart excellent morphological preservation. Formaldehyde fixes tissue by cross linking, a process which can reduce the antigenicity of tissue and weakens the intensity of immunohistochemical stains. Preliminary studies have shown that Histochoice tissue fixative offers equal or greater staining intensity than neutral buffered formalin (NBF). This study compares these fixatives quantitatively and presents the results in unambiguous statistical terms. Tissue samples were collected, bisected, and fixed in NBF or Histochoice. The sections were stained with a total of 21 antibodies, and color images were collected. The hue, saturation, and value were determined for each positive pixel and an ANOVA performed. Small differences in hue were noted in 8 of 21 cases. Histochoice‐fixed tissue gave a greater mean saturation than NBF with 57.1% of the antibodies tested. No significant difference in the saturation was detected in 28.6% of the cases; NBF gave higher mean saturation levels with only 14.3% of the antibodies. Histochoice‐fixed tissue was found to give lower values in 66.7% of cases than those prepared with NBF, indicating darker staining. These results show that Histochoice produces staining intensity that is comparable, and in many cases superior, to formalin.
Keywords: Image analysis, fixation, immunohistochemistry, color analysis
