RNAscope® technology provided by Advanced Cell Diagnostics (ACD) allows the detection and evaluation of coinciding mRNA expression profiles in the same or adjacent cells in unprecedented quantitative detail using multicolor fluorescent in situ hybridization (FISH). While already extensively used in thinly sectioned material of various pathological tissues and, to a lesser extent, in some whole mounts, we provide here a detailed approach to use the fluorescent RNAscope method in the mouse inner ear and thick brain sections by modifying and adapting existing techniques of whole mount fluorescent in situ hybridization (WH-FISH). We show that RNAscope WH-FISH can be used to quantify local variation in overlaying mRNA expression intensity, such as neurotrophin receptors along the length of the mouse cochlea. We also show how RNAscope WH-FISH can be combined with immunofluorescence (IF) of some epitopes that remain after proteinase digestion and, to some extent, with fluorescent protein markers such as tdTomato. Our WH-FISH technique provides an approach to detect cell-specific quantitative differences in developing and mature adjacent cells, an emerging issue revealed by improved cellular expression profiling. Further, the presented technique may be useful in validating single-cell RNAseq data on expression profiles in a range of tissue known or suspected to have locally variable mRNA expression levels.

• Klíčová slova
• Fluorescent proteins, Immunofluorescence, In situ hybridization, RNA quantification, RNAscope, Whole mount FISH,
• MeSH
• fluorescenční protilátková technika metody   MeSH
• hybridizace in situ fluorescenční MeSH
• kochlea metabolismus   MeSH
• messenger RNA genetika   metabolismus   MeSH
• myši inbrední BALB C MeSH
• myši inbrední C57BL MeSH
• neurotrofin 3 metabolismus   MeSH
• receptor trkB genetika   metabolismus   MeSH
• receptor trkC genetika   metabolismus   MeSH
• regulace genové exprese MeSH
• zobrazování trojrozměrné MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• messenger RNA MeSH
• neurotrofin 3 MeSH
• receptor trkB MeSH
• receptor trkC MeSH

Textile fabrics are highly anisotropic, so that their mechanical properties including strengths are a function of direction. An extreme case is when a woven fabric sample is cut in such a way where the bias angle and hence the tension loading direction is around 45° relative to the principal directions. Then, once loaded, no yarn in the sample is held at both ends, so the yarns have to build up their internal tension entirely via yarn-yarn friction at the interlacing points. The overall fabric strength in such a sample is a result of contributions from the yarns being pulled out and those broken during the process, and thus becomes a function of the bias direction angle θ, sample width W and length L, along with other factors known to affect fabric strength tested in principal directions. Furthermore, in such a bias sample when the major parameters, e.g. the sample width W, change, not only the resultant strengths differ, but also the strength generating mechanisms (or failure types) vary. This is an interesting problem and is analysed in this study. More specifically, the issues examined in this paper include the exact mechanisms and details of how each interlacing point imparts the frictional constraint for a yarn to acquire tension to the level of its strength when both yarn ends were not actively held by the testing grips; the theoretical expression of the critical yarn length for a yarn to be able to break rather than be pulled out, as a function of the related factors; and the general relations between the tensile strength of such a bias sample and its structural properties. At the end, theoretical predictions are compared with our experimental data.

• Klíčová slova
• bias directions, fabric strength, failure types, sample width effect, yarn pullout and breakage,
• Publikační typ
• časopisecké články MeSH