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On/off-switchable LSPR nano-immunoassay for troponin-T

View Article: PubMed Central - PubMed

ABSTRACT

Regeneration of immunosensors is a longstanding challenge. We have developed a re-usable troponin-T (TnT) immunoassay based on localised surface plasmon resonance (LSPR) at gold nanorods (GNR). Thermosensitive poly(N-isopropylacrylamide) (PNIPAAM) was functionalised with anti-TnT to control the affinity interaction with TnT. The LSPR was extremely sensitive to the dielectric constant of the surrounding medium as modulated by antigen binding after 20 min incubation at 37 °C. Computational modelling incorporating molecular docking, molecular dynamics and free energy calculations was used to elucidate the interactions between the various subsystems namely, IgG-antibody (c.f., anti-TnT), PNIPAAM and/or TnT. This study demonstrates a remarkable temperature dependent immuno-interaction due to changes in the PNIPAAM secondary structures, i.e., globular and coil, at above or below the lower critical solution temperature (LCST). A series of concentrations of TnT were measured by correlating the λLSPR shift with relative changes in extinction intensity at the distinct plasmonic maximum (i.e., 832 nm). The magnitude of the red shift in λLSPR was nearly linear with increasing concentration of TnT, over the range 7.6 × 10−15 to 9.1 × 10−4 g/mL. The LSPR based nano-immunoassay could be simply regenerated by switching the polymer conformation and creating a gradient of microenvironments between the two states with a modest change in temperature.

No MeSH data available.


Representative figures used for free energy calculation by computational analysis; (a) coil-PNIPAAM, (b) globular-PNIPAAM, (c) anti-TnT-coil-PNIPAAM complex; (d) anti-TnT-globular-PNIPAAM complex; (e) TnT-coil-PNIPAAM complex and (f) TnT-globular-PNIPAAM complex.
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f5: Representative figures used for free energy calculation by computational analysis; (a) coil-PNIPAAM, (b) globular-PNIPAAM, (c) anti-TnT-coil-PNIPAAM complex; (d) anti-TnT-globular-PNIPAAM complex; (e) TnT-coil-PNIPAAM complex and (f) TnT-globular-PNIPAAM complex.

Mentions: Where, the first term refers to the molecular mechanics energy and includes van der Waals, electrostatic and internal energies of the system. The second term refers to the desolvation free energy that again has polar and non-polar contributions. The third term refers to the entropic contributions to the total free energy. In most of the cases, the entropic contributions do not majorly dictate the binding affinity and so the discussion is usually based on the total contributions from the first two terms. The nonpolar contribution to the desolvation free energy is computed using the solvent accessible surface area. The polar contribution is computed using either generalised Born approach (in GB approach) or Poisson Boltzmann approach (as in PB approach)434445. Overall, these two approaches differ with respect to the polar term in the desolvation free energy. In this study, the whole discussion of free energies is based only on a generalised Born approach. We have presented the individual and total free energies (Supporting InformationTables S1–5) for five different complexes namely, anti-TnT:TnT, anti-TnT:c-PNIPAAM, anti-TnT:g-PNIPAAM, TnT:c-PNIPAAM and TnT:g-PNIPAAM as shown in Fig. 5.


On/off-switchable LSPR nano-immunoassay for troponin-T
Representative figures used for free energy calculation by computational analysis; (a) coil-PNIPAAM, (b) globular-PNIPAAM, (c) anti-TnT-coil-PNIPAAM complex; (d) anti-TnT-globular-PNIPAAM complex; (e) TnT-coil-PNIPAAM complex and (f) TnT-globular-PNIPAAM complex.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5382532&req=5

f5: Representative figures used for free energy calculation by computational analysis; (a) coil-PNIPAAM, (b) globular-PNIPAAM, (c) anti-TnT-coil-PNIPAAM complex; (d) anti-TnT-globular-PNIPAAM complex; (e) TnT-coil-PNIPAAM complex and (f) TnT-globular-PNIPAAM complex.
Mentions: Where, the first term refers to the molecular mechanics energy and includes van der Waals, electrostatic and internal energies of the system. The second term refers to the desolvation free energy that again has polar and non-polar contributions. The third term refers to the entropic contributions to the total free energy. In most of the cases, the entropic contributions do not majorly dictate the binding affinity and so the discussion is usually based on the total contributions from the first two terms. The nonpolar contribution to the desolvation free energy is computed using the solvent accessible surface area. The polar contribution is computed using either generalised Born approach (in GB approach) or Poisson Boltzmann approach (as in PB approach)434445. Overall, these two approaches differ with respect to the polar term in the desolvation free energy. In this study, the whole discussion of free energies is based only on a generalised Born approach. We have presented the individual and total free energies (Supporting InformationTables S1–5) for five different complexes namely, anti-TnT:TnT, anti-TnT:c-PNIPAAM, anti-TnT:g-PNIPAAM, TnT:c-PNIPAAM and TnT:g-PNIPAAM as shown in Fig. 5.

View Article: PubMed Central - PubMed

ABSTRACT

Regeneration of immunosensors is a longstanding challenge. We have developed a re-usable troponin-T (TnT) immunoassay based on localised surface plasmon resonance (LSPR) at gold nanorods (GNR). Thermosensitive poly(N-isopropylacrylamide) (PNIPAAM) was functionalised with anti-TnT to control the affinity interaction with TnT. The LSPR was extremely sensitive to the dielectric constant of the surrounding medium as modulated by antigen binding after 20 min incubation at 37 °C. Computational modelling incorporating molecular docking, molecular dynamics and free energy calculations was used to elucidate the interactions between the various subsystems namely, IgG-antibody (c.f., anti-TnT), PNIPAAM and/or TnT. This study demonstrates a remarkable temperature dependent immuno-interaction due to changes in the PNIPAAM secondary structures, i.e., globular and coil, at above or below the lower critical solution temperature (LCST). A series of concentrations of TnT were measured by correlating the λLSPR shift with relative changes in extinction intensity at the distinct plasmonic maximum (i.e., 832 nm). The magnitude of the red shift in λLSPR was nearly linear with increasing concentration of TnT, over the range 7.6 × 10−15 to 9.1 × 10−4 g/mL. The LSPR based nano-immunoassay could be simply regenerated by switching the polymer conformation and creating a gradient of microenvironments between the two states with a modest change in temperature.

No MeSH data available.