Modulation of DNA-Modified Gold-Nanoparticle Stability in Salt with Concatemeric Single-Stranded DNAs for Colorimetric Bioassay Development
✍ Scribed by Dr. M. Monsur Ali; Pushpinder Kanda; Sergio D. Aguirre; Prof. Dr. Yingfu Li
- Publisher
- John Wiley and Sons
- Year
- 2011
- Tongue
- English
- Weight
- 581 KB
- Volume
- 17
- Category
- Article
- ISSN
- 0947-6539
No coin nor oath required. For personal study only.
✦ Synopsis
Gold nanoparticles (AuNPs) exhibit distinct optical properties under specific conditions. For example, spherical AuNPs of 13 nm in diameter appear red when well dispersed and blue or purple when aggregated. [1, 2] Two major methods used to control DNA-modified AuNP aggregation are DNA-templated assembly (also known as cross-linking aggregation) [2, 3] and salt-induced aggregation. [4] The first approach modulates AuNP stability through sequence-specific hybridization and has been widely explored for biosensing applications. [2, 5] The second approach modulates AuNP stability by using divalent metal ions that neutralize the negative charges of DNA on AuNP surfaces, and has also been exploited to develop various colorimetric bioassays. [4, 6] DNA-conjugated AuNPs have also been used to make organized nanostructures with long single-stranded DNA molecules (ssDNAs) produced by rolling circle amplification (RCA). [7] RCA is an isothermal process of amplifying DNA by using a short DNA primer, a circular ssDNA template (DNA circle) and Phi29 DNA polymerase. [8] Because ssDNA products from RCA contain many repeating (concatemeric) units, RCA technique has been widely used to achieve sensitive detection of DNA or other biological targets. [8, 9] In a recent study, AuNPs and RCA have been employed together for the development of a colorimetric assay to detect single-nucleotide polymorphisms through a crosslinking strategy. [10] Herein, we report on the novel use of RCA products to protect AuNPs against salt-induced aggregation. AuNPs modified with a short ssDNA complementary to part of an RCA product (RP) can organize onto the RP through sequence-specific hybridization. We have found that, when MgCl 2 (salt) is added, the AuNP-RP assemblies form unique, globular, island-like nanostructures (Figure 1 a, lower path), which we term "NP islands (NPI)". The color of the NPI-containing solution remains red whereas in the absence of RP, AuNPs form purple aggregates (Figure 1 a, upper path). We have also exploited this finding for the development of a colorimetric assay for ATP detection wherein the action of an ATP-dependant DNAzyme (aptazyme) is coupled to RCA and the resulted RP is used to protect AuNPs from salt-induced aggregation.
AuNPs used in this study (NP1, Figure 1 b) were functionalized with a short ssDNA to a surface density of approximately 50 strands per particle. Optimization of MgCl 2 concentration revealed that evident red-to-purple color transition of NP1 solution occurred at 30 mm of MgCl 2 (along with 20 mm Tris-HCl, pH 7.5 at 23 8C, 100 mm NaCl); thus, this salt condition was used throughout our study. Two RCA products, RP1 (containing NP1-binding site; blue letters in Figure 1 b) and RP2 (which does not bind NP1), were prepared by using the relevant circular DNA templates and primers (for their sequences, see Figure S1 a in the Supporting Information) and analyzed by agarose gel electrophoresis (Figure S1 b). Complex formation between RP1 and NP1 (but not between RP2 and NP1) was also confirmed by agarose gel electrophoresis (Figure S1 c).
Before addition of MgCl 2 , the NP1-only, NP1-RP2, and NP1-RP1 solutions (samples 1-3, respectively) all appeared red (tube inserts in Figure 2 a,123). TEM analysis revealed that gold nanoparticles in samples 1 and 2 were well dispersed (Figure 2 a, 1 and 2) whereas those in sample 3 were clustered (Figure 2 a, 3), suggesting that NP1s in this sample assembled onto RP1 by sequence-specific hybridization. With the addition of 30 mm MgCl 2 , samples 1 and 2 turned purple (tube inserts in Figure 2 a, 4 and 5) whereas sample 3 remained red (tube insert in Figure 2 a, 6). TEM analysis produced interesting results: huge NP aggregates were found in samples 1 and 2 (Figure 2 a, 4 and 5), howev-
[a] Dr.