Recent RNA research has revealed the close involvement of various RNAs in cellular functions. RNAs are becoming the inevitable target molecules for research into details of gene expression. RNA and its related complexes are also promising targets for disease diagnosis. Multi cellular specimens such as organ tissues, histopathological specimens, and embryos are among the possible targets of RNA-based diagnostic techniques. In this report, we focused on a method that would provide such spatial and temporal information. We demonstrated that an RNA-specific probe (OMUpy2) was not only applicable to the detection of a specific mRNA in Drosophila embryos in a temporal and spatial manner but was also relatively quick and easy to use. The probe, OMUpy2, could be applied to other multi cellular systems for RNA-based diagnosis and research. The promising results of this manuscript show the great potential of RNA-based detection for both biological research and diagnostic medicine.RNA-specific probe (OMUpy2) was found to be applicable to the detection of specific mRNA in Drosophila embryos from the view point of temporal and spatial localization in multi cellar specimens.

In situ visualization of c-fosmRNA was shown both in fixed cells and in living cells using bispyrene-modified 2′-O-methyl RNA probes (OMUpy2) or phosphorothioate modified OMUpy2 (OMUpy2-S), which was RNA-specific with high sensitivity, and a system for time-lapse imaging of c-fosmRNA was successfully developed.

2′-O-Psoralen-conjugated antisense oligonucleotide was able to recognize a point mutation of mRNA. It had outstanding ability to photo-cross-link only to oligoribonucleotides (ORN) having a point mutation. This type of antisense molecule is the only one of its kind so far. To give high photo-cross-linking efficiency and sequence selectivity to antisense molecules, we synthesized novel photo-reactive oligonucleotides (2′-Ps-xom) containing psoralen at the 2′-O-position adenosine via an ethoxymethylene (2′-Ps-eom), propoxymethylene (2′-Ps-pom) and butoxymethylene (2′-Ps-bom) linker, respectively. We evaluated the photo-cross-linking efficiency and sequence selectivity in photo-cross-linking of 2′-Ps-xom to the complementary ORN and to an ORN having a mismatch base. Among them, 2′-Ps-eom exhibited superior photo-cross-linking efficiency with high sequence selectivity.We developed 2′-O-psoralenylmethoxyalkyl adenosine conjugated antisense oligonucleotides that photo-cross-link to an oligoribonucleotide having a point mutation site with high sequence selectivity and high photo-cross-linking efficiency upon UVA irradiation.

This paper describes the cultivation of primary cells in a microchamber and the real-time monitoring of small amounts of antibody secretion after the introduction of a minute amount of stimulus by a microinjector using time-resolved fluorescence anisotropy analysis.

This article presents a real-time monitoring system for cellular analysis using micro total analysis systems technology. Time-resolved luminescence anisotropy analysis was adopted for real-time detection of small amounts of a target protein produced by a small number of cells. The system was tested by real-time monitoring of the antibody secretion by hybridomas. The cells were successfully cultivated in a micro-incubation chamber (240 nl) fabricated on a microchip. The quantification of the antibody was achieved using the Ru(II) complex-labeled Staphylococcus aureus protein A probe, which can bind specifically to the Fc region of the antibody. Using this system, we detected as little as 24 fmol of immunoglobulin G under physiological conditions without the bound/free separation protocol. We successfully achieved real-time and quantitative monitoring of small amounts of antibody production by approximately 200 hybridoma cells. This method could be applied to various cellular analyses using small numbers of cells.

In situ visualization of c-fosmRNA was shown both in fixed cells and in living cells using bispyrene-modified 2′-O-methyl RNA probes (OMUpy2) or phosphorothioate modified OMUpy2 (OMUpy2-S), which was RNA-specific with high sensitivity, and a system for time-lapse imaging of c-fosmRNA was successfully developed.