Biomedical Engineering Reference
In-Depth Information
3
3
2.5
2.5
2
2
1.5
1.5
1
1
0.5
0.5
0
0
0
20
40
60
80
100
120
0
20
40
60
80
100
120
B
Figure 11.16
Binding and dissociation of RNA synthesized on a (a) 420 nM template and a (b) 42 nM template
(
Blair et al., 2007
).
Whe
n only a solid line (--) is used then a single-fractal analysis applies. When
both a dashed (- - -) and a solid (--) line are used then the dashed line represents a single-fractal
analysis and the solid line represents a dual-fractal analysis.
A
Time (min)
Time (min)
(
Tyagi and Kramer, 1996; Leone et al., 1998; Guilietti et al., 2001; Summerer and Marx,
2002; Marras et al., 2006
).
Blair et al. (2007
) have developed a novel hybridization-based assay for the real-time moni-
toring of RNA synthesis. In their method complementary nucleotides were used to quantify
the amount of RNA production by T7 polymerase.
Figure 11.16a
shows the binding and dis-
sociation of RNA synthesized on a 420 nM template. The RNA concentration was deter-
mined by conversion using a calibration curve. A dual-fractal analysis is required to
adequately describe the binding kinetics. A single-fractal analysis is adequate to describe
the dissociation kinetics. The values of (a) the binding rate coefficient,
k
, and the fractal
dimension,
D
f
, for a single-fractal analysis, (b) the binding rate coefficients,
k
1
and
k
2
, and
the fractal dimensions,
D
f1
and
D
f2
, for a dual-fractal analysis, and (c) the dissociation rate
coefficient,
k
d
, and the fractal dimension for the dissociation phase,
D
fd
, for a single-fractal
analysis are given in
Tables 11.9
and
11.10
. It is of interest to note that for a dual-fractal
analysis as the fractal dimension increases by a factor of 1.35 from a value of
D
f1
equal to
1.6862 to
D
f2
r equal to 2.2812, the binding rate coefficient increases by a factor of 2.79 from
a value of
k
1
equal to 0.1909 to
k
2
equal to 0.5324. The changes in the degree of heterogene-
ity on the sensor chip surface and in the binding rate coefficient are in the same direction.
Table 11.9: Binding and dissociation rate coefficients for (a) RNA synthesized on a 42 nM and a
420 nM template (
Blair et al., 2007
).
Template
Concentration (nM)
k
k
1
k
2
k
d
420
0.2546
0.0121
0.1909
0.0121
0.5324
0.0120
0.00204
0.00031
42
0.02317 0.00234
na
na
na
