Biology Reference
In-Depth Information
Figure 9.17.
Commercial dual screen-printed carbon electrode. See also
Color Insert.
The target sequences have been chosen so that the same primer
is able to generate the PCR products of both bacteria. Thus, with
a unique screen-printed strip it is possible to identify the causing
bacteria of the disease.
The dual screen-printed electrode used in this section is shown
in Fig. 9.17.
9.4.4.1 Genosensor design
As it has been commented, the genosensor design is the same that
was used in section 9.4.3.2, withsome variations
Gold nanostructuration is carried out by applying a constant
current of
−
5
μ
A for 2 minutes in an acidic medium containing
AuCl
4
−
1 mM. Then theelectrode is generously rinsedwith water.
A4
μ
L aliquot of 50 nM thiolated probes is dropped in each
working electrode for 10 minutes. One working electrode supports
the probe corresponding to
S. pneumoniae
, and the other supports
the probe corresponding to
M. pneumoniae
. Then, the electrode is
rinsed with 0.1 M Tris-HNO
3
bufferpH7.2,andablockingstepis
carried out with a 40-
μ
L aliquot of casein 2% for 20 minutes and
rinsed with0.1 M Tris bufferpH 7.2
Hybridizationstepiscarriedoutatroomtemperaturein2
×
SSC
buffer by dropping a 40
μ
L aliquot of the biotinylated target for 1
hourandrinsingwithTrisbufferpH7.2.Afterthat40
μ
Lof5
×
10
−
10
M S-AP are dropped on the electrode for 1 hour. Then the electrode
is rinsed with Tris buffer pH 9.8 and enzymatic reaction with 3-IP
and silver ions, and detection step is carried out as mentioned in
previous sections.
