Raman Dye-Labeled Nanoparticle Probes for Proteins

Cao, Y. Charles; Jin, Rongchao; Nam, Jwa Min; Thaxton, C. Shad; Mirkin, Chad A.

doi:10.1021/ja0366235

Cited by 385 publications

(313 citation statements)

References 15 publications

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“…Gold nanoparticles (15,30,50,80,150, and 250 nm) were functionalized with fluorophore labeled, alkanethiol modified oligonucleotides. Nanoparticle concentration was determined using UV-vis spectroscopy.…”

Section: Resultsmentioning

confidence: 99%

Maximizing DNA Loading on a Range of Gold Nanoparticle Sizes

2006

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We have investigated the variables that influence DNA coverage on gold nanoparticles. The effects of salt concentration, spacer composition, nanoparticle size, and degree of sonication have been evaluated. Maximum loading was obtained by salt aging the nanoparticles to ~0.7 M NaCl in the presence of DNA containing a poly(ethylene glycol) (PEG) spacer. In addition, DNA loading was substantially increased by sonicating the nanoparticles during the surface loading process. Lastly, nanoparticles up to 250 nm in diameter were found have ~2 orders of magnitude higher DNA loading than smaller (13-30 nm) nanoparticles, a consequence of their larger surface area. Stable large particles are attractive for a variety of biodiagnostic assays.

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Section: Resultsmentioning

confidence: 99%

Maximizing DNA Loading on a Range of Gold Nanoparticle Sizes

2006

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“…Raman spectra enable fingerprinting of molecules which is of particular interest for bio-applications. Surface enhanced Raman scattering (SERS) provides greater detection sensitivity than conventional Raman spectroscopy [1][2][3], and it is quickly gaining traction in the study of biological molecules adsorbed on a metal surface [4][5][6][7][8][9][10][11][12]. SERS spectroscopy allows for the detection and analysis of minute quantities of analytes because it is possible to obtain high-quality SERS spectra at submonolayer molecular coverage as a result of the large scattering enhancements.…”

Section: Introductionmentioning

confidence: 99%

Adaptive silver films for surface‐enhanced Raman spectroscopy of biomolecules

Drachev

Thoreson

Nashine

et al. 2005

J Raman Spectroscopy

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The interaction of biological molecules with a typical substrate for surface-enhanced Raman scattering (SERS) often leads to their structural and functional changes. In this work we describe SERS substrates, called adaptive silver films (ASFs), in which the biomaterial and the substrate act in concert to produce excellent Raman enhancement through local restructuring of the metal surface while at the same time preserving the properties (such as conformational state and binding activity) of the analyte. These adaptive substrates show great promise for SERS spectroscopy of many different types of biomolecules, and we provide several current examples of their use. IntroductionOne of main advantages of Raman scattering as a detection method for molecule sensing is well known. Raman spectra enable fingerprinting of molecules which is of particular interest for bio-applications. Surface enhanced Raman scattering (SERS) provides greater detection sensitivity than conventional Raman spectroscopy [1][2][3], and it is quickly gaining traction in the study of biological molecules adsorbed on a metal surface [4][5][6][7][8][9][10][11][12]. SERS spectroscopy allows for the detection and analysis of minute quantities of analytes because it is possible to obtain high-quality SERS spectra at submonolayer molecular coverage as a result of the large scattering enhancements. SERS has also been shown to be sensitive to molecular orientation and to the distance from the metal surface [13]. Thus, SERS is well-suited for biomolecule studies in which specificity and sensitivity to the conformational state and orientation of the molecule are very important.The SERS enhancement mechanism originates in part from the large local electromagnetic fields caused by resonant surface plasmons that can be optically excited at certain wavelengths for metal particles of different shapes or closely spaced groups of particles [14][15][16][17][18][19][20][21]. For aggregates of interacting particles, which are often structured as fractals, plasmon resonances can be excited in a very broad spectral range [22]. In addition to electromagnetic field enhancement, metal nanostructures and molecules can form charge-transfer complexes that provide further enhancement for SERS [23][24][25][26][27][28][29]. The resulting overall enhancement depends critically on the particle or aggregate nanostructure morphology [22,[30][31][32][33][34][35][36], and it can be as high as 10 5 to 10 8 for

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“…Previously, SERS was applied for immobilized protein detection 5,6 by coupling small Raman-active dyes to gold nanoparticles functionalized by ligands.…”

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“…20 As a result, sensitivity is not quantitative 5 or is limited to nM range 5 which does not compare favorably with fluorescence methods. For high sensitivity Raman sensing with dye molecules, 7 long acquisition times are needed and molecules are easily subject to degradation under laser radiation.…”

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Protein microarrays with carbon nanotubes as multicolor Raman labels

et al. 2008

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Detection of biomolecules is important in proteomics anddilution. Multiple-color SWNT Raman labels are developed using isotopic SWNTs, and employed for sensing in a multiplexed fashion requiring only a single excitation source. RESULTS Assay design for protein detection by SWNT Raman-tagsA secondary antibody, goat anti-mouse immunoglobulin-G (GaM-IgG), was conjugated to highly water soluble, short (~50-150 nm, see Supplementary Fig. S1 online), macromolecular SWNTs, functionalized with PEGylated phospholipids 26 (PL-PEG, see Methods). GaM-IgG conjugation imparted binding specificity of SWNT-tags to mouse antibodies (Fig. 1). Protein immobilization on substrates was performed in arrayed fashion, by either covalent attachment to Au-coated glass or on commercial microarray slides by standard robotic spotting (see Method). We developed a novel 6-arm-branched carboxylate-terminated PEG, grafted onto Au-coated surfaces for protein 5 immobilization (Fig. 1a), that afforded excellent protein attachment and high resistance to non-specific binding (NSB) effects. Proteins, such as polyclonal mouse IgG or human serum albumin (HSA), were immobilized on the assay surface and either detected byRaman scattering upon binding of GaM-IgG-conjugated SWNTs (using 785 nm excitation laser, see Methods), or were used in sandwich assays, to capture analyte protein (e.g., anti-HSA IgG raised in mouse) from dilute serum (Fig. 1b).To enhance Raman scattering intensity following direct or sandwich assay detection of analyte by SWNT labeled GaM-IgG, we annealed the gold-coated substrate To explore the sensitivity limit of SWNT Raman-based protein detection, human serum albumin (HSA) and GaM-IgG/SWNTs were used as model capture and reporting agents for sandwich assay detection of monoclonal mouse anti-HSA IgG (aHSA) spiked into fetal bovine serum (FBS, Fig. 1). Figure 3a shows Raman mapping images of HSA spots after exposure to various concentrations of aHSA from 100 pM to 1 fM followed by incubation with GaM-IgG/SWNTs. The images were generated by plotting the integrated SWNT G-band intensity at each point (50 μm x 50 μm pixel size) over one quarter of the protein spot. In the maps, uniform SWNT signals were observed within HSA spots exposed to aHSA at concentrations above ~1 pM. At lower concentrations, the SWNT signal was sparse, consistent with a small number of aHSA IgGs captured by the HSA layer. Defining the limit of detection (LOD) as twice the standard deviation above the control (without analyte), we reproducibly obtained aHSA detection sensitivity down to 1 fM, over 8 orders of dynamic range (Fig. 3b, replicate sets of data are shown, obtained on independent assay chips with different batches of GaM-IgG/SWNT conjugates). The data exhibited sigmoidal or S-shape behavior, 33 suggesting saturation and steric hindrance effects for detection at high concentrations, and increased proportional influence of residual NSB effects at the lower detection limit.To compare our SWNT-Raman-based detection to standard fluorescence-based protein mi...

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Raman Dye-Labeled Nanoparticle Probes for Proteins

Cited by 385 publications

References 15 publications

Maximizing DNA Loading on a Range of Gold Nanoparticle Sizes

Maximizing DNA Loading on a Range of Gold Nanoparticle Sizes

Adaptive silver films for surface‐enhanced Raman spectroscopy of biomolecules

Protein microarrays with carbon nanotubes as multicolor Raman labels

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