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.

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The complementary guest binding process onto the cyanurate lipid 1 monolayer at the air-water interface was observed by using a highly sensitive 27 MHz quartz-crystal microbalance (QCM), which was attached horizontally on the monolayer from the air phase. Binding amount (∆m), association constants (Ka), as well as binding and dissociation rate constants (k1 and k-1) could be obtained from time courses of the frequency decrease (mass increase) of the QCM. A highly sensitive 27 MHz QCM was used to detect small mass changes of Langmuir adsorption of small guest molecules instead of our conventional 9 MHz QCM. Guests such as 2,4,6-triaminopyrimidine (A) possessing three complementary hydrogen bonding sites were selectively bound to the monolayer 1 showing a 1:2 host-guest binding ratio, k 1 ) 2.8 M -1 s -1 , k-1 ) 7.2 × 10 -4 s -1 , and Ka ) 3900 M -1 . The Ka value was consistent with that obtained by NMR spectra in bulk CDCl3. Guest molecules of A hardly bound to monolayers 2-4 with -NH2, -CONH2, and -OH head groups. Binding constants of guest molecules to the monolayer 1 were of the following order: 2,4,6triaminopyrimidine (A) > 2,6-diaminopyridine (B) possessing three hydrogen bonds > 2-aminopyridine (C) forming two hydrogen bonds > pyridine (D) with only one hydrogen bond ≈ barbituric acid (E) having no complementary hydrogen binding sites. The obtained binding kinetics at the water interface were compared with molecular mechanics calculations of binding energy in vacuum.

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Complementary molecular recognition in the gas phase onto self-assembled monolayers bearing thymine or adenine bases as terminal groups was studied by using a highly sensitive 63 MHz quartz-crystal microbalance. Association constants (K a), binding and dissociation rate constants (k 1 and k - 1), and binding amount in nanogram level (Δm) could be obtained from time coarses of frequency decrease (mass increase) at various gaseous guest concentrations. Kinetic parameters were corrected to avoid the effect of different adsorption ability of guest molecules. Binding of 2-aminopyridine (an adenine model) onto a thymine monolayer showed the 50−100 times larger binding constant than that of a noncomplementary guest molecule, γ-butyrolactam. On the other hand, an adenine monolayer preferentially bound the thymine model of γ-butyrolactam over 2-aminopyridine, the adenine model. When a simple hydrocarbon monolayer of 1-decanethiol was used, all of these guest molecules were hardly bound. The selectivity between complementary and noncomplementary nucleobase pairs observed in the gas/thin film surface was comparable to the selectivity seen in organic solvents. The formation energy of hydrogen bonding obtained from K a values in the gas/thin film surface was consistent with that calculated by molecular mechanics as in a vacuum. K a values of the guest molecules were dependent on the nucleobase content in the mixed monolayer of nucleobase and alkyl chains.