Chieco P, Normanni P, Boor P J
Stain Technol. 1984 Jul;59(4):201-11. doi: 10.3109/10520298409113857.
An improvement in the histochemical demonstration of soluble dehydrogenase enzymes has been obtained by preincubating frozen sections in a nitroblue tetrazolium (NBT)/acetone solution, followed by routine incubation in polyvinyl alcohol (PVA) enriched media. Tissue binding properties of NBT were shown clearly to be decreased in histochemical media containing the colloid PVA for soluble enzymes, thus causing loss of the final reaction product (formazan) from the sections. The preincubation step in NBT/acetone allows tetrazolium salt to bind firmly to tissue lipoprotein (substantivity) and diminishes the loss of reduced formazan from heavily reacting tissue sections. The time course of NBT substantivity was examined and it was found that NBT binds rapidly to tissues (liver, kidney, heart) during preincubation, so that a preincubation of 30-60 seconds at room temperature is sufficient to improve the final morphological results greatly. Microspectrophotometric measurements of matched controls and NBT/acetone preincubated sections show that the preincubation step may slightly decrease lactate dehydrogenase (LDH) and glucose-6-phosphate dehydrogenase (G6PDH) activities. This decrease was probably due to increased binding efficiency of formazan to cell lipoproteins but was judged, however, to be irrelevant compared to the morphological advantages produced by the NBT/acetone preincubation procedure.
通过将冰冻切片在氮蓝四唑（NBT）/丙酮溶液中预孵育，然后在富含聚乙烯醇（PVA）的培养基中进行常规孵育，可溶性脱氢酶的组织化学显示得到了改进。在含有用于可溶性酶的胶体PVA的组织化学培养基中，NBT的组织结合特性明显降低，从而导致切片中最终反应产物（甲臜）的损失。NBT/丙酮中的预孵育步骤使四唑盐牢固地结合到组织脂蛋白上（直接性），并减少了反应强烈的组织切片中还原甲臜的损失。研究了NBT直接性的时间进程，发现预孵育期间NBT迅速与组织（肝脏、肾脏、心脏）结合，因此在室温下预孵育30 - 60秒足以大大改善最终的形态学结果。对匹配的对照和NBT/丙酮预孵育切片进行的显微分光光度测量表明，预孵育步骤可能会略微降低乳酸脱氢酶（LDH）和葡萄糖 - 6 - 磷酸脱氢酶（G6PDH）的活性。这种降低可能是由于甲臜与细胞脂蛋白的结合效率提高，但与NBT/丙酮预孵育程序产生的形态学优势相比，这被认为是无关紧要的。
