An optimized triphenyltetrazolium chloride method for identification of cerebral infarcts

2,3,5-Triphenyltetrazolium chloride (TTC) staining is a convenient procedure for detection of brain infarcts but no standardized procedure is available. We report here an optimized and economic procedure of staining with TTC. Rats were subjected to reversible middle cerebral artery (MCA) occlusion (2-h ischemia and 24-h reperfusion). At the end of reperfusion, brain was isolated and sliced rostro-caudally into serial 2-mm-thick slices. Sets of three serial slices from each brain were incubated for 30 min at 37 degreesC in three different concentrations of TTC-the first slice of the set in 1%, the second in 0.05% and the third in 0.1% TTC-in phosphate-buffered saline. Staining characteristics, optical density (OD) and infarct size were compared between juxtaposing cut surfaces of the slices stained with the three concentrations of TTC. After the first use, 0.05% TTC solution was stored at 4-8 degreesC and reused on the same day or on subsequent days. TTC at 0.05% concentration provided high contrast staining with clear demarcation between normal and infarct tissue. The infarct size in 0.05% TTC-stained slices correlated well with that in 0.1% TTC (r= 0.92)- and 1% TTC (r= 0.93)-stained preparations. 'Nonspecific' staining of corpus callosum and the anterior commissures was minimal with the method. Once-used 0.05% TTC Solution Could be stored at 4-8 degreesC and reused. In conclusion. stainin, with 0.05% TTC provided unproved delineation of brain infarcts, reduced 'nonspecific' staining of white matter and the infarct size correlated well with that measured after 1% TTC stainin,; the method also reduces the costs to 1/20. (C) 2004 Elsevier B.V. All rights reserved.