Antigen Retrieval with Urea for Immunoslaining of NOSs in Paraffin Sections of the Rat Brain

概要

論文の詳細を見る
The effects of several antigen retrieval methods on immunostaining of NOSs in paraffin-embedded rat brain tissues were studied in combjnation with the influence of two fixatives, 4% paraformaldehyde (PFA) in 0.1 M phosphate buffer (PH 7.4), and 70% ethanol. The first antibodies for NOSs were purchased from AffinityBioreagents Co., Ltd., New Jersey. Without any antigenretrieval treatment, paraffin sections of the rat brain fixed with both PFA and ethanol were never immunostained. Only the ethanol fixative was useful for antigen retrieval in immunostaining of NOSs. In antigen retrieval with microwave heating, retrieval solutions such as water (except the one containing 6 M of urea), Tris-HCI butters at lower pH (〓7.6) with or without urea, and a citrate buffer had no effect. However, a Tris-HCI butter at pH 9.5 was effective at least for eNOS, and an addition of 5% urea to it enhanced this effect. For both bNOS and eN0S imrrmunostaining, the most effective method was to preincubate sections in an aqueous solution of urea (6 M) for 4 to 12 hr prior to immunostaining. The sections treated with 6M urea solution showed strong immunostaining reactivity for bNOS in neurons dispersively distributed in the cerebral cortex and in nuclei of the brain stem. With the same pretreatment of sections, eNOS immunoreactivity was also clearly detected on the wall of small blood vessels and capillaries in the parenchyma of the brain.
日本組織細胞化学会の論文