Controlled kinetics of non-enzymatic chemiluminescence reactions for simple imaging of DNA and protein.
A robust and sensitive non-enzymatic chemiluminescence (CL) imaging method is presented. In the method a fast-emitting CL reaction is tuned to furnish a slower-emitting reaction suitable for simple CL imaging. Typically, non-enzymatic CL reactions between luminol or fluorescein and oxygen species generated by KCN as catalyst, were rather fast and unsuitable for CL imaging; the speed of the reactions could, however, be reduced substantially by changing KCN for CH(3)CN or benzonitrile. Light emission from the tuned CL reaction was intense and long-lived, and even with a simple arrangement high sensitivity could be achieved. The maximum CL peak was reached after approximately 1.5 min in the presence of 25% acetonitrile, and as little as 16 fmol commercial isoluminol-labeled streptavidin was detected and visualized on either microplate or membrane. The approach was further illustrated by imaging of DNA on a membrane and of antibody on a microplate by use of biotin-streptavidin chemistry. Overall, this simple, economical, and sensitive CL imaging system is expected to be very useful in biochemical analysis, and greatly complements currently used enzyme-based CL imaging methods, especially in routine applications.
Lau
Choiwan
C
School of Pharmaceutical Sciences, Nagasaki University,1-14 Bunkyo-Machi, Nagasaki 852-8521, Japan.
Lu
Jianzhong
J
Yamaguchi
Takashi
T
Kai
Masaaki
M
D004247:DNA / Q000032:analysis*; D007700:Kinetics; D008163:Luminescent Measurements; D011506:Proteins / Q000032:analysis*
D011506:Proteins; D004247:DNA