Fabrication of DNA microarrays on polydopamine-modified gold thin films for SPR imaging measurements

Abstract

Polydopamine (PDA) films were fabricated on thin film gold substrates in a single-step polymerization-deposition process from dopamine solutions and then employed in the construction of robust DNA microarrays for the ultrasensitive detection of biomolecules with nanoparticle-enhanced surface plasmon resonance (SPR) imaging. PDA multilayers with thicknesses varying from 1 to 5 nm were characterized with a combination of scanning angle SPR and AFM experiments, and 1.3 ± 0.2 nm PDA multilayers were chosen as an optimal thickness for the SPR imaging measurements. DNA microarrays were then fabricated by the reaction of amine-functionalized single-stranded DNA (ssDNA) oligonucleotides with PDA-modified gold thin film microarray elements, and were subsequently employed in SPR imaging measurements of DNA hybridization adsorption and protein-DNA binding. Concurrent control experiments with non-complementary ssDNA sequences demonstrated that the adhesive PDA multilayer was also able to provide good resistance to the nonspecific binding of biomolecules. Finally, a series of SPR imaging measurements of the hybridization adsorption of DNA-modified gold nanoparticles onto mixed sequence DNA microarrays were used to confirm that the use of PDA multilayer films is a simple, rapid, and versatile method for fabricating DNA microarrays for ultrasensitive nanoparticle-enhanced SPR imaging biosensing.

Figures

Figure 1

Real-time SPRI measurement of PDA film growth. Inset shows the linear relationship between growth time and reflectivity change. Timeexp denotes the cumulated time that the array surface has been exposed to dopamine solution.

Figure 2

Scanning angle measurements show the shifts in angle minima as PDA deposition time lengthens. Inset displays the linear dependence of angle on film growth time.

Figure 3

SPRI measurement of ssDNA hybridizing with its complementary counterpart (sequence A, solid curve) on the DNA microarray, giving 2.5 Δ%R. The data also shows no nonspecific adsorption of ssDNA to the noncomplementary, control DNA (sequence B, dotted curve) on the microarray surface. The inset is the real-time SPRI difference image.

Figure 4

SPRI measurement of SSB binding onto ssDNA attached to PDA layer on the microarray.

Figure 5

Schematic of CT ssDNA-modified AuNPs hybridizing with D ssDNA. The mixed, two component spots were constructed by attaching ssDNA to the PDA layer. The numbers on the microarray represent the percentage of D ssDNA on the surface. Also shown is the real-time difference image of ssDNA microarray after signal saturation from CT-AuNPs hybridization.

Figure 6

SPRI measurements of CT ssDNA modified AuNPs hybridization onto ssDNA microarray built from PDA attachment chemistry. Inset shows the CT -AuNPs hybridization onto the surfaces with 0% to 1% D ssDNA.