Fuentes Ricardo, Fernández Juan
Department of Cell and Developmental Biology, University of Pennsylvania, Perelman School of Medicine, Philadelphia, PA, 19104, USA.
Methods Mol Biol. 2014;1211:1-13. doi: 10.1007/978-1-4939-1459-3_1.
A new procedure for improved in situ hybridization of zebrafish whole-mount oocytes, embryos, and early larvae is described. The procedure relies on the simultaneous fixation/permeabilization of samples using formaldehyde as fixative and short C-chain aliphatic carboxylic acids, particularly glacial acetic acid, as permeabilizers. As compared with in situ hybridization performed with routine methods, our procedure is simpler and provides better structural preservation of cells and tissues, equivalent mRNA signals, and similar results in embryos of different developmental stages. It is hypothesized that during aldehyde fixation short C-chain aliphatic carboxylic acids modulate the rate of formation and/or destruction of methylene bridges established between cell proteins.
本文描述了一种用于改进斑马鱼全胚卵母细胞、胚胎和早期幼体原位杂交的新方法。该方法依赖于使用甲醛作为固定剂和短链脂肪族羧酸（特别是冰醋酸）作为通透剂对样品进行同步固定/通透处理。与常规方法进行的原位杂交相比，我们的方法更简单，能更好地保存细胞和组织的结构，mRNA信号相当，并且在不同发育阶段的胚胎中能得到相似的结果。据推测，在醛类固定过程中，短链脂肪族羧酸会调节细胞蛋白质之间形成和/或破坏亚甲基桥的速率。
