Kinetic measurements of DNA hybridization on an oligonucleotide-immobilized 27-MHz quartz crystal microbalance.
A highly sensitive 27-MHz quartz-crystal microbalance, on which a 10-30-mer oligonucleotide was immobilized as a probe molecule, was employed to detect hybridization of complementary oligonucleotides in aqueous solution. From frequency decreases (mass increases due to the hybridization) with passage of time, kinetic parameters such as association constants (K(a)) and binding and dissociation rate constants (k(1) and k(-1)) could be obtained, as well as binding (hybridization) amount at the nanogram level (delta m). Kinetic studies were carried out by changing various parameters: (i) the immobilization method of a probe oligonucleotide on Au electrode, (ii) number of mismatching bases in sequences of target oligonucleotides, (iii) length of both probe and target oligonucleotides, (iv) hybridization temperature, and (v) ionic strength in solution. The obtained results were compared with those obtained by a surface plasmon resonance method using a BIAcore system.
Okahata
Y
Y
Department of Biomolecular Engineering, Tokyo Institute of Technology, Nagatsuda, Midori-ku, Yokohama 226-8507, Japan.
Kawase
M
M
Niikura
K
K
Ohtake
F
F
Furusawa
H
H
Ebara
Y
Y
D001360:Avidin / Q000737:chemistry; D020029:Base Pairing; D001710:Biotin / Q000737:chemistry; D004247:DNA / Q000737:chemistry*; D007700:Kinetics; D009693:Nucleic Acid Hybridization / Q000379:methods*; D015345:Oligonucleotide Probes / Q000737:chemistry; D009841:Oligonucleotides / Q000737:chemistry*; D009994:Osmolar Concentration; D011791:Quartz / Q000737:chemistry; D013696:Temperature
D015345:Oligonucleotide Probes; D009841:Oligonucleotides; D001360:Avidin; D011791:Quartz; D001710:Biotin; D004247:DNA