Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.27.5 (RNase)
 17,967 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The chapter focuses on the detection of specific mRNA by in situ hybridization (ISH) in synovial tissue specimens. This technique is widely applied, reliable, specific, and sensitive, because even small quantities of mRNA can be detected. Presented here contemporary protocols for ISH using a combined nonradioactive immunohistochemical detection system. In overview, the following steps have to be covered to perform ISH. (1) mRNA probes (sense and antisense) are generated by in vitro transcription of cDNA utilizing digoxigenin-labeled UTP nucleotides, (2) fixed tissue sections are digested with trypsin and treated consecutively with prehybridization solutions, (3) hybridization with labeled riboprobes takes place at 50 degrees C overnight in a humid chamber, (4) unbound riboprobe is removed by incubation with RNase A and additional washing with buffers, (5) stringent washing steps are performed with solutions of different sodium dodecyl sulfate, SSC, and formamide concentrations, (6) digoxigenin-labeled probes are detected immunohistochemically using antidigoxigenin antibodies linked with alkaline phosphatase and NBT/BCIP as detection system.
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PMID:In situ hybridization of synovial tissue. 1795 52

This appendix describes the preparation of selected bacterial media and of buffers and reagents used in the manipulation of nucleic acids and proteins. Recipes for cell culture media and reagents are located elsewhere in the manual. RECIPES: Acids, concentrated stock solutions; Ammonium acetate, 10 M; Ammonium hydroxide, concentrated stock solution; ATP, 100 mM; BCIP, 5% (w/v); BSA (bovine serum albumin), 10% (100 mg/ml); Denhardt solution, 100x; dNTPs: dATP, dTTP, dCTP, and dGTP; DTT, 1 M; EDTA, 0.5 M (pH 8.0); Ethidium bromide solution; Formamide loading buffer, 2x; Gel loading buffer, 6x; HBSS (Hanks balanced salt solution); HCl, 1 M; HEPES-buffered saline, 2x; KCl, 1 M; LB medium; LB plates; Loading buffer; 2-ME, (2-mercaptoethanol)50 mM; MgCl(2), 1 M; MgSO(4), 1 M; NaCl, 5 M; NaOH, 10 M; NBT (nitroblue tetrazolium chloride), 5% (w/v); PCR amplification buffer, 10x; Phosphate-buffered saline (PBS), pH approximately 7.3; Potassium acetate buffer, 0.1 M; Potassium phosphate buffer, 0.1 M; RNase a stock solution (DNase-free), 2 mg/ml; SDS, 20%; SOC medium; Sodium acetate, 3 M; Sodium acetate buffer, 0.1 M; Sodium phosphate buffer, 0.1 M; SSC (sodium chloride/sodium citrate), 20x; SSPE (sodium chloride/sodium phosphate/EDTA), 20x; T4 DNA ligase buffer, 10x; TAE buffer, 50x; TBE buffer, 10x; TBS (Tris-buffered saline); TCA (trichloroacetic acid), 100% (w/v); TE buffer; Terrific broth (TB); TrisCl, 1 M; TY medium, 2x; Urea loading buffer, 2x.
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PMID:Common buffers, media, and stock solutions. 1842 17