Non-radioactive in situ hybridization. A comparison of several immunocytochemical detection systems using reflection-contrast and electron microscopy.
A number of immunocytochemical detection systems for determining the chromosomal localization of specific nucleic acid sequences by non-radioactive in situ hybridization have been compared. The procedures were: 1. the peroxidase/diaminobenzidine (PO/DAB) combination, either or not gold/silver intensificated; 2. alkaline phosphatase marking using the nitro-blue tetrazolium plus bromochloro-indolyl phosphate substrate combination (AP/NBT + BCIP); and 3. immunogold with or without silver enhancement. The procedures were first tested and optimized in dot blot experiments and then applied to in situ hybridization. As hybridization probes, both a middle-repetitive and a unique sequence (modified with 2-acetylaminofluorene (AAF] were used. The advantages and disadvantages of the various methods for reflection contrast (RC) or transmission electron microscopic (TEM) visualization of hybrids are discussed.
D015073:2-Acetylaminofluorene; D015100:3,3'-Diaminobenzidine; D000469:Alkaline Phosphatase; D002877:Chromosomes, Human; D006651:Histocytochemistry; D006801:Humans; D007120:Immunochemistry; D007211:Indoles; D008853:Microscopy / Q000379:methods; D008854:Microscopy, Electron; D009693:Nucleic Acid Hybridization
D007211:Indoles; D015100:3,3'-Diaminobenzidine; C035455:5-bromo-4-chloro-3-indoxyl phosphate; D015073:2-Acetylaminofluorene; D000469:Alkaline Phosphatase