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Optimizing Nanoplasmonic Biosensor Sensitivity with Orientated Single Domain Antibodies.

Raphael MP, Christodoulides JA, Byers JM, Anderson GP, Liu JL, Turner KB, Goldman ER, Delehanty JB - Plasmonics (2015)

Bottom Line: We demonstrate that orientated single domain antibodies (sdAb) can optimize nanoplasmonic sensitivity by comparing three anti-ricin sdAb constructs to biotin-neutravidin, a model system for small and highly orientated ligand studies.LSPR imaging of electrostatically orientated sdAb exhibited a ricin sensitivity equivalent to that of the biotinylated LSPR biosensors for neutravidin.These results, combined with the facts that sdAb are highly stable and readily produced in bacteria and yeast, build a compelling case for the increased utilization of sdAbs in nanoplasmonic applications.

View Article: PubMed Central - PubMed

Affiliation: Bioelectronics and Sensing, Code 6363, Naval Research Laboratory, 4555 Overlook Ave SW, Washington, DC 20375 USA.

ABSTRACT

Localized surface plasmon resonance (LSPR) spectroscopy and imaging are emerging biosensor technologies which tout label-free biomolecule detection at the nanoscale and ease of integration with standard microscopy setups. The applicability of these techniques can be limited by the restrictions that surface-conjugated ligands must be both sufficiently small and orientated to meet analyte sensitivity requirements. We demonstrate that orientated single domain antibodies (sdAb) can optimize nanoplasmonic sensitivity by comparing three anti-ricin sdAb constructs to biotin-neutravidin, a model system for small and highly orientated ligand studies. LSPR imaging of electrostatically orientated sdAb exhibited a ricin sensitivity equivalent to that of the biotinylated LSPR biosensors for neutravidin. These results, combined with the facts that sdAb are highly stable and readily produced in bacteria and yeast, build a compelling case for the increased utilization of sdAbs in nanoplasmonic applications.

No MeSH data available.


Related in: MedlinePlus

a SPR response to the conjugation of C8-zip sdAbs for a range of ligand concentrations followed by the introduction of 100 nM ricin in (b). The control study in b consisted of a 100-nM ricin solution incubated with 5 μM of C8 for 45 min to block the binding sites. It was then introduced over a 4.0-μg/mL C8 functionalized surface. The vertical dashed line separates the association phase (left) in which the ligand or analyte solution is flowing over the surface from the dissociation phase (right) in which buffer flows over the surface
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Fig1: a SPR response to the conjugation of C8-zip sdAbs for a range of ligand concentrations followed by the introduction of 100 nM ricin in (b). The control study in b consisted of a 100-nM ricin solution incubated with 5 μM of C8 for 45 min to block the binding sites. It was then introduced over a 4.0-μg/mL C8 functionalized surface. The vertical dashed line separates the association phase (left) in which the ligand or analyte solution is flowing over the surface from the dissociation phase (right) in which buffer flows over the surface

Mentions: To optimize the plasmonic sensitivity to ricin, a range of ligand concentrations were introduced over the EDC/Sulfo-NHS activated SPR surface followed by a saturating concentration of ricin (100 nM). Fig. 1a shows the SPR response to C8-zip in which the concentration was varied from 0.4 to 30 μg/mL in PBS, pH 7.0. At the higher ligand concentrations, there were diminishing returns with regards to ricin-binding capacity (Fig. 1b) with the 30 μg/mL exposed surface binding only 8 % more than the surface coated at 10 μg/mL. Exceeding this 30 μg/mL value resulted in a rapid increase in non-specific binding and minimal ricin sensitivity enhancement. C8 and D12f-rhiz exhibited similar surface saturation concentrations, lying between 1 and 30 μg/mL (Supplementary Material). Ligand concentration values such as the 30 μg/mL value for C8-zip were defined as optimal based on these criteria of maximizing sensitivity and minimizing non-specific binding. A control study was also conducted in which a 100-nM ricin solution was first incubated with 5 μM of C8 for 45 min to block the ricin-binding sites. When the blocked ricin was introduced over a C8-functionalized surface, no response was detectable, demonstrating levels of non-specific binding below the SPRi limit of detection (Fig. 1b).Fig. 1


Optimizing Nanoplasmonic Biosensor Sensitivity with Orientated Single Domain Antibodies.

Raphael MP, Christodoulides JA, Byers JM, Anderson GP, Liu JL, Turner KB, Goldman ER, Delehanty JB - Plasmonics (2015)

a SPR response to the conjugation of C8-zip sdAbs for a range of ligand concentrations followed by the introduction of 100 nM ricin in (b). The control study in b consisted of a 100-nM ricin solution incubated with 5 μM of C8 for 45 min to block the binding sites. It was then introduced over a 4.0-μg/mL C8 functionalized surface. The vertical dashed line separates the association phase (left) in which the ligand or analyte solution is flowing over the surface from the dissociation phase (right) in which buffer flows over the surface
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4644190&req=5

Fig1: a SPR response to the conjugation of C8-zip sdAbs for a range of ligand concentrations followed by the introduction of 100 nM ricin in (b). The control study in b consisted of a 100-nM ricin solution incubated with 5 μM of C8 for 45 min to block the binding sites. It was then introduced over a 4.0-μg/mL C8 functionalized surface. The vertical dashed line separates the association phase (left) in which the ligand or analyte solution is flowing over the surface from the dissociation phase (right) in which buffer flows over the surface
Mentions: To optimize the plasmonic sensitivity to ricin, a range of ligand concentrations were introduced over the EDC/Sulfo-NHS activated SPR surface followed by a saturating concentration of ricin (100 nM). Fig. 1a shows the SPR response to C8-zip in which the concentration was varied from 0.4 to 30 μg/mL in PBS, pH 7.0. At the higher ligand concentrations, there were diminishing returns with regards to ricin-binding capacity (Fig. 1b) with the 30 μg/mL exposed surface binding only 8 % more than the surface coated at 10 μg/mL. Exceeding this 30 μg/mL value resulted in a rapid increase in non-specific binding and minimal ricin sensitivity enhancement. C8 and D12f-rhiz exhibited similar surface saturation concentrations, lying between 1 and 30 μg/mL (Supplementary Material). Ligand concentration values such as the 30 μg/mL value for C8-zip were defined as optimal based on these criteria of maximizing sensitivity and minimizing non-specific binding. A control study was also conducted in which a 100-nM ricin solution was first incubated with 5 μM of C8 for 45 min to block the ricin-binding sites. When the blocked ricin was introduced over a C8-functionalized surface, no response was detectable, demonstrating levels of non-specific binding below the SPRi limit of detection (Fig. 1b).Fig. 1

Bottom Line: We demonstrate that orientated single domain antibodies (sdAb) can optimize nanoplasmonic sensitivity by comparing three anti-ricin sdAb constructs to biotin-neutravidin, a model system for small and highly orientated ligand studies.LSPR imaging of electrostatically orientated sdAb exhibited a ricin sensitivity equivalent to that of the biotinylated LSPR biosensors for neutravidin.These results, combined with the facts that sdAb are highly stable and readily produced in bacteria and yeast, build a compelling case for the increased utilization of sdAbs in nanoplasmonic applications.

View Article: PubMed Central - PubMed

Affiliation: Bioelectronics and Sensing, Code 6363, Naval Research Laboratory, 4555 Overlook Ave SW, Washington, DC 20375 USA.

ABSTRACT

Localized surface plasmon resonance (LSPR) spectroscopy and imaging are emerging biosensor technologies which tout label-free biomolecule detection at the nanoscale and ease of integration with standard microscopy setups. The applicability of these techniques can be limited by the restrictions that surface-conjugated ligands must be both sufficiently small and orientated to meet analyte sensitivity requirements. We demonstrate that orientated single domain antibodies (sdAb) can optimize nanoplasmonic sensitivity by comparing three anti-ricin sdAb constructs to biotin-neutravidin, a model system for small and highly orientated ligand studies. LSPR imaging of electrostatically orientated sdAb exhibited a ricin sensitivity equivalent to that of the biotinylated LSPR biosensors for neutravidin. These results, combined with the facts that sdAb are highly stable and readily produced in bacteria and yeast, build a compelling case for the increased utilization of sdAbs in nanoplasmonic applications.

No MeSH data available.


Related in: MedlinePlus