Abstract
Fixation time plays a more critical role in quantitative computerized immunohistochemistry than in routine qualitative studies. This study assessed the significance of fixation time in methanol-Carnoy solution (MC) and 10% neutral buffered formalin (NBF) relative to computerized quantitation of immunohistochemical staining with 2 different antibodies.Normal skin specimens were fixed in NBF or MC solution for times ranging from 4 hr to 1 week. Paraffin sections were stained immunohistochemically with a polyclonal antibody for transforming growth factor-β (TGF-β) or a monoclonal antibody for thrombomodulin (TM). Extracellular matrix-associated TGF-β immunoreactivity and TM-positive vessels were identified and assessed with quantitative computerized image analysis.For both fixatives and antibodies, staining was superior in specimens fixed for 24 hr. TGF-β and TM stainings were stronger and more specific in MC-fixed tissues than in NBF-fixed tissues. We concluded that 24 hr in NBF and MC is the optimal fixation time for quantitative image analysis. MC is superior to NBF for immunohistochemical staining of TGF-β and TM. (The J Histotechnol 22:109, 1999)
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