Use of fluorochrome-labeled rRNA targeted oligonucleotide probe and tyramide signal amplification to improve sensitivity of fluorescence in situ hybridization.
A tyramide signal amplification (TSA) system was used in combination with a conventional fluorochrome-labeled 16S rRNA oligonucleotide probe to increase the sensitivity of fluorescence in situ hybridization. TSA was performed after hybridization resulted in a low fluorescence signal intensity. In contrast to the horseradish peroxidase-tyramide signal amplification (HRP-TSA) system and biotin-tyramide signal amplification (biotin-TSA) system, no additional expensive probe labeling was required. A whole cell hybridization technique was used to compare the fluorescence signal obtained using a monolabeled probe with that obtained using the TSA system. The fluorescence signal of the probe obtained using the TSA system was much higher than that obtained using the monolabeled probe. The technique was successfully applied to the in situ detection of microbial communities in anaerobic sludge. It was demonstrated that TSA resulted in an increased in sensitivity, as the fluorescence signal intensity was much higher than that obtained using a conventional probe.
Jupraputtasri
Wiwat
W
School of Bioresources, King Mongkut University of Technology Thonburi, Bangkhuntien, Bangkok 10150, Thailand.
Cheevadhanarak
Supapon
S
Chaiprasert
Pawinee
P
Tanticharoen
Morakot
M
Techkarnjanaruk
Somkiet
S