Quantification of in situ hybridization signals in rat testes
Quantification of in situ hybridization signals in rat testes
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DOI：
10.1369/jhc.4a6249.2004
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发表时间：
2004-06-01
影响因子：
3.2
通讯作者：
Sasaki, J
中科院分区：
文献类型：
--
作者：
Kimura, T;Kosaka, J;Sasaki, J
我们进行了基础研究定量原位杂交（ISH）信号在大鼠睾丸，一个合适的器官的量化，因为生殖细胞进行同步发展，并显示阶段特异性基因表达。在该模型实验中，选择rRNA作为石蜡切片中的可氢化RNA。用布因固定剂固定并与地高辛标记探针杂交的标本可以通过图像的“后处理”容易地进行定量分析。与探针杂交的rRNA量在早期初级精母细胞中最大，其次是粗线期初级精母细胞，然后是双线期精母细胞，最后是次级精母细胞和精子细胞。在减数分裂的中期、后期和末期以及第一步精子细胞的早期，其含量达到较低水平，然后在精子发生过程中略有增加。ISH rRNA染色是用于评价组织切片中mRNA定量分析和可杂交RNA水平的有用参数。
We performed basic research into quantifying in situ hybridization (ISH) signals in rat testis, a suitable organ for the quantification because germ cells undergo synchronized development and show stage-specific gene expression. In this model experiment, rRNA was selected as the hyloridizable RNA in paraffin sections. Specimens fixed with Bouin's fixative and hybridized with digoxygenin-labeled probes could easily be analyzed quantitatively through "posterization" of the images. The amount of rRNA hybridized with the probe was greatest in early primary spermatocytes, followed by pachytene primary spermatocytes, then diplotene spermatocytes, and finally by secondary spermatocytes and spermatids. The amounts reached low levels in metaphase, anaphase, and telophase of meiotic division and early step 1 spermatids, and then slightly increased during spermiogenesis. ISH rRNA staining was a useful parameter for evaluation of the quantitative analysis of mRNA and the levels of hybridizable RNA in tissue sections.