TY - JOUR
T1 - Transglutaminase-mediated in situ hybridization (TransISH) system
T2 - A new methodology for simplified mRNA detection
AU - Kitaoka, Momoko
AU - Mitsumori, Masayuki
AU - Hayashi, Kounosuke
AU - Hiraishi, Yoshiyuki
AU - Yoshinaga, Hisao
AU - Nakano, Koji
AU - Miyawaki, Katsuyuki
AU - Noji, Sumihare
AU - Goto, Masahiro
AU - Kamiya, Noriho
PY - 2012/7/17
Y1 - 2012/7/17
N2 - Figure Persented: Detection and localization of specific DNA or RNA sequences in cells and tissues are of great importance for biological research, diagnosis, and environmental monitoring. However, the most common procedure for in situ hybridization employs laborious immunostaining techniques. In the present study, we report proof-of-concept for a new RNA-enzyme conjugated probe for the detection of mRNA on tissue sections with a simple procedure. An RNA probe modified with a specific dipeptide substrate of transglutaminase was prepared. Alkaline phosphatase was then covalently and site-specifically combined to the dipeptide-labeled RNA using microbial transglutaminase. The new RNA probe labeled with alkaline phosphatase was validated by in situ hybridization (ISH) and proved to be a sensitive and sequence specific probe for mRNA detection in tissues. The new transglutaminase-mediated ISH (TransISH) strategy is free from antigen-antibody reaction, leads to one-step signal amplification after hybridization, and thus will be widely applicable for highly sensitive nucleic acid detection.
AB - Figure Persented: Detection and localization of specific DNA or RNA sequences in cells and tissues are of great importance for biological research, diagnosis, and environmental monitoring. However, the most common procedure for in situ hybridization employs laborious immunostaining techniques. In the present study, we report proof-of-concept for a new RNA-enzyme conjugated probe for the detection of mRNA on tissue sections with a simple procedure. An RNA probe modified with a specific dipeptide substrate of transglutaminase was prepared. Alkaline phosphatase was then covalently and site-specifically combined to the dipeptide-labeled RNA using microbial transglutaminase. The new RNA probe labeled with alkaline phosphatase was validated by in situ hybridization (ISH) and proved to be a sensitive and sequence specific probe for mRNA detection in tissues. The new transglutaminase-mediated ISH (TransISH) strategy is free from antigen-antibody reaction, leads to one-step signal amplification after hybridization, and thus will be widely applicable for highly sensitive nucleic acid detection.
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U2 - 10.1021/ac2034198
DO - 10.1021/ac2034198
M3 - Article
C2 - 22746100
AN - SCOPUS:84863968503
SN - 0003-2700
VL - 84
SP - 5885
EP - 5891
JO - Analytical Chemistry
JF - Analytical Chemistry
IS - 14
ER -