Abstract
In situ hybridization (ISH) technique with a biotin-labeled probe was established for detecting feline interleukin 1 (IL-1)α mRNA in necropsied specimens. Homology between human IL-1α cDNA used as a probe and feline IL-1α mRNA was confirmed by means of dot blot hybridization using the biotin-labeled probe. Hence, we tried by this biotinylated probe to detect mRNA of IL-1α in paraffin-embedded sections. The following results were obtained for the routine procedures: 1) coating slides with poly-L-lysine and/or heating at 60°C at least for 6 hours gave an excellent result for the adhesion of the tissue sections, 2) 10μg/ml solution of proteinase K treatment for 30 minutes or 50 to 100μg/ml solution of proteinase K treatment for 10 to 30 minutes at 37°C gave the good results in the detection of ISH signal, 3) suitable denaturation time of probes at 70 to 90°C was 5 to 15 minutes, and 4) effective hybridization was obtained by incubation for 24 hours at 4°C, for 18 to 24 hours at 25°C or for 5 to 24 hours at 37°C
