研究了纳米颗粒的多酶活性,包括过氧化物酶和过氧化氢酶,以及在葡萄糖高灵敏检测中的应用.
Prussian Blue Modi ed Ferritin as Peroxidase Mimetics and Its Applications in Biological DetectionTable I. Kinetic parameters of PB-Ft NPs. Catalyst PB-Ft E/M 7 22× 10 10 7 39× 10 10 7 22× 10 10 7 39× 10 10 3 09× 10 10 3 09× 10 10 Substrate TMB H2 O2 (TMB) ABTS H2 O2 (ABTS) TMB H2 O2 Km (mM) 1.594 11.984 0.532 0.537 0.307 323.6 Vmax (Ms 1 1 92× 10 7 7 20× 10 7 3 13× 10 8 6 14× 10 9 1 06× 10 6 1 17× 10 6 kcat/s 1 2 66× 102 3 13× 101 4 32× 101 3 35× 101 3 43× 103 3 79× 103
Zhang et al.
(kcat/Km/M 1 s 1 1 7× 105 2 6× 103 8 6× 104 6 2× 104 1 1× 106 1 2× 104
PBMNPs9
Notes: E is the nanoparticle concentration, Km is the Michaelis constant, Vmax is the maximal reaction velocity and Kcat is the catalytic constant, where kcat= Vmax/[E]. Note that the kcat value shows the catalytic ef ciency per nanoparticle. PBMNPs represent PB coated Fe2 O3 magnetic nanoparticles reported in our previous work.9
provides a lower detective concentration of glucose compared to carboxyl-modi ed graphene oxide (GO-COOH) (1 mol/L)39 and Fe3 O4 nanoparticles (5 mol/L).40 And the slope of the tted line can be regarded as the sensitivity to glucose, which is also much larger than the values in other literatures, indicating PB-Ft NPs are quite sensitive in glucose detection. 3.4. Enzyme-Linked Immunosorbent Assay (ELISA) The principle of ELISA is the use of an enzyme to deliver a signal that a particular antigen-antibody reaction has occurred and to what extent. In our experiment, PB-Ft NPs were used as a probe with the simultaneous combination of the peroxidase-like activity of PB and the
speci city of ferritin. Compared to the previously repored PBMNPs-SPA bioconjugates, PB-Ft NPs can effectively inhibit the non-speci c adsorption owning to native ferritin shell. Figure 8(a) shows that the absorbance increases with nanoparticle concentration and reaches a pleatue when the concentration of PB-Ft NPs is approximately 4 0× 10 11 mol/L, which was thus selected as a optimal probe concentration for anti-HoSF antibody detection. As shown in Figure 8(b), the HoSF group displays an absorb
ance slightly higher than the PBS group (control-1), the reason is that the iron oxide cores of ferritins presented weak catalytic activity, as indicated above. The competition group shows an absorbance higher than the HoSF group (control-2), a possible reason is the incubated PBFt NPs could not be totally washed off and gave rise to(b) 120
RESEARCH ARTICLE
(a)120 substrate: TMB
substrate: ABTS
Relatively activity(%)
100 80 60 40 20 0 2 4 6 8 10
Relatively activity (%)
100 80 60 40 20 0 2 4 6 8 10
pH (c)100
pH (d) 1.0substrate: ABTS 0.9
Absorbance (415 nm)
Relatively activity(%)
80 60 40 20 0 10 20 30 40 50 60 70 80
0.8 0.7 0.6 0.5 0.4 0.3 0.2 1 2 3 4 5 6 7 8 y=0.0796x+0.3727 R2=0.9896
Temperature (ºC)Fig. 6.
Concentration of PB-Ft NPs (µg/mL)
The peroxidase-like activity of the PB-Ft NPs is pH (a, b), temperature (c), PB-Ft NPs concentration (d) dependent.
6
J. Nanosci. Nanotechnol. 12, 1–8, 2012