کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
6270458 1614656 2007 10 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Detection of two different mRNAs in a single section by dual in situ hybridization: A comparison between colorimetric and fluorescent detection
موضوعات مرتبط
علوم زیستی و بیوفناوری علم عصب شناسی علوم اعصاب (عمومی)
پیش نمایش صفحه اول مقاله
Detection of two different mRNAs in a single section by dual in situ hybridization: A comparison between colorimetric and fluorescent detection
چکیده انگلیسی

We have compared the performance of two methods designed to simultaneously detect two different mRNAs within a single brain section by dual ISH. Specific mRNA riboprobes labeled with biotin and digoxigenin were simultaneously hybridized and visualized using either brightfield or fluorescence microscopy. For brightfield visualization, the biotin-labeled riboprobe was detected with a peroxidase chromogen, whereas, an alkaline phosphatase substrate was used for the detection of the digoxigenin-labeled riboprobe. Dual fluorescent ISH involved the detection of the biotin-labeled riboprobe with an Alexa®488-conjugated streptavidin followed by the visualization of the digoxigenin-labeled riboprobe with the red fluorescent substrate HNPP. The dual ISH protocols presented here offer sensitive methods to detect the expression of two mRNAs of interest, with both colorimetric and fluorescent ISH each having its strengths and limitations. For example, dual colorimetric ISH has proven to be particularly useful to study the distribution of two mRNAs in different brain nuclei, whereas, dual fluorescent ISH has provided better results when studying the co-localization of two different mRNAs in single neurons. The comprehensive step-by-step procedure is presented, together with a troubleshooting section in which the advantages and limitations of these procedures are reviewed in depth. Moreover, alternative protocols for dual ISH were also compared to those presented here.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Journal of Neuroscience Methods - Volume 162, Issues 1–2, 15 May 2007, Pages 119-128
نویسندگان
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