CYTOCHEMISTRY AND ELECTRON MICROSCOPY
CYTOCHEMISTRY AND ELECTRON MICROSCOPY
复制标题
细胞化学和电子显微镜
作者：
D. Sabatini;K. Bensch;R. Barrnett
本文所介绍的醛类及其一般用作固定剂的较适宜浓度为：戊二醛4~6.5%、乙二醛4%、羟基己二醛12.5%、巴豆醛10%、异丁醛5%、乙醛10%和异丁烯醛5%。将其制备为二甲胂酸盐或磷酸盐缓冲溶液（0.1至0.2 M，pH 6.5至7.6），除戊二醛外，其含有蔗糖（0.22至0.55 M）。固定0.5小时至24小时后，将块储存在冷（4 ° C）缓冲液（0.1M）加蔗糖（0.22M）中。该材料用于酶组织化学，用于Epon包埋后的电子显微镜检查（有或没有用1%或2%的四氧化锇进行第二次固定），以及用于两种技术的组合。在醛固定后，细胞的膜分化不明显，核结构不同于通常观察到的四氧化锇。在四氧化锇的后固定，即使经过长时间的储存，开发的图像，值得注意的是在戊二醛的情况下，在很大程度上是无法区分的最佳条件下固定的组织与四氧化锇单独。脂肪酸酯酶，乙酰胆碱酯酶，碱性磷酸酶，酸性磷酸酶，5-核苷酸酶，腺苷三磷酸酶，DPNH和TPNH黄递酶的活动后，大多数的固定剂组化。细胞色素氧化酶，琥珀酸脱氢酶，葡萄糖-6-磷酸酶保留后，羟基醛，并在较小程度上，乙二醛固定后。上述几种酶的活性的最终产物相对于精细结构被定位。为此目的，使用双重固定程序，在每种情况下选择适当的醛。
The aldehydes introduced in this paper and the more appropriate concentrations for their general use as fixatives are: 4 to 6.5 per cent glutaraldehyde, 4 per cent glyoxal, 12.5 per cent hydroxyadipaldehyde, 10 per cent crotonaldehyde, 5 per cent pyruvic aldehyde, 10 per cent acetaldehyde, and 5 per cent methacrolein. These were prepared as cacodylate- or phosphate-buffered solutions (0.1 to 0.2 M, pH 6.5 to 7.6) that, with the exception of glutaraldehyde, contained sucrose (0.22 to 0.55 M). After fixation of from 0.5 hour to 24 hours, the blocks were stored in cold (4°C) buffer (0.1 M) plus sucrose (0.22 M). This material was used for enzyme histochemistry, for electron microscopy (both with and without a second fixation with 1 or 2 per cent osmium tetroxide) after Epon embedding, and for the combination of the two techniques. After fixation in aldehyde, membranous differentiations of the cell were not apparent and the nuclear structure differed from that commonly observed with osmium tetroxide. A postfixation in osmium tetroxide, even after long periods of storage, developed an image that—notable in the case of glutaraldehyde—was largely indistinguishable from that of tissues fixed under optimal conditions with osmium tetroxide alone. Aliesterase, acetylcholinesterase, alkaline phosphatase, acid phosphatase, 5-nucleotidase, adenosine triphosphatase, and DPNH and TPNH diaphorase activities were demonstrable histochemically after most of the fixatives. Cytochrome oxidase, succinic dehydrogenase, and glucose-6-phosphatase were retained after hydroxyaldipaldehyde and, to a lesser extent, after glyoxal fixation. The final product of the activity of several of the above-mentioned enzymes was localized in relation to the fine structure. For this purpose the double fixation procedure was used, selecting in each case the appropriate aldehyde.