A novel method for monitoring, in real time, the formaldehyde
(FA)-induced DNA–protein cross-linking process with the piezoelectric
quartz crystal impedance (PQCI) technique is proposed. The method was used
to monitor FA-induced DNA–lysozyme cross-link formation. Lysozyme was
directly immobilized on the silver electrode surface of a piezoelectric
quartz crystal by adsorption. The lysozyme-coated piezoelectric sensor was
in contact with FA and DNA solutions. The time courses of the resonant
frequency and equivalent circuit parameters of the sensor during the
cross-linking were simultaneously obtained and are discussed in detail. On
the basis of the feature of the multi-dimensional information provided by
the PQCI technique, it was concluded that the observed frequency decrease
could be mainly ascribed to the mass increase resulting from the
cross-linking. According to the frequency decrease with time, the kinetics
of the cross-linking process were quantitatively studied. A piezoelectric
response model for the cross-linking was theoretically derived. Fitting the
experimental data to the model, the kinetic parameters, such as the binding
and dissociation rate constants (k1 and
k−1) and the cross-linking equilibrium constant
(Ka), were determined. At 37 °C, the
k1, k−1 and
Ka values obtained were 7.0 (±0.1) ×
10−5 (μg ml−1)−1
s−1, 6.6 (±0.1) × 10−3
s−1 and 1.06 (±0.02) × 10−2
(μg ml−1)−1, respectively.
