Association of gold nanorods in water solutions: Influence of globular proteins

By the absorption spectroscopy method, optical properties of gold nanorods (10 × 38 nm) and their interaction with globular protein bovine hemoglobin and bovine serum albumin were investigated. Nanorod behavior was studied in water solution and in solution of 97 mM NaCl under ultrasound action during 90 min and results were then compared. In water solutions, nanorod coagulation (aggregation) was observed with reduced optical density of the longitudinal plasmon band widening at λ > 800 nm. In NaCl solution, absorption spectra evolution had a complex character and was in some degree analogous to the result that was obtained for two-dimensional grids of gold nanoparticles when changing the distance between them. By interacting with serum albumin, stabilization of colloid solution and dissociation of nanorod aggregates were observed.     

Preparation and properties of the water-soluble chlorophyll-bovine serum albumin complexes

By mixing chlorophyll (Chl) a or b with a dense bovine serum albumin solution, the water-soluble Chl-bovine serum albumin complexes were prepared. These complexes, eluted near the void volume on a gel filtration, were separated well from unreacted bovine serum albumin, indicating an aggregation of such molecules in the complexes. Preparation of chlorophyllide (Chlide) a- or Chlide b-bovine serum albumin complex was unsuccessful, while the phytol-, and beta-carotene-bovine serum albumin complexes could be obtained. Chls in the Chl-bovine serum albumin complexes had the following characteristics. Main absorption peak of Chl a or b in the red region occurred at 675 nm or 652 nm, respectively. The Chl a-bovine serum albumin complex having absorption peak at 740 nm was also prepared. As compared with the stabilities of Chl a and b in Triton X-100. Both Chls in the bovine serum albumin-complexes were stable against oxidative stresses, such as photobleaching, Fenton reagent, peroxidase-H2O2 system. But they were easily hydrolyzed by chlorophyllase. These properties of Chls in the bovine serum albumin-complexes were similar to those of Chls in the isolated light-harvesting Chl a/b protein complex. A possible localization of Chls within the bovine serum albumin complexes was suggested that the porphyrin moiety of Chl was buried in bovine serum albumin; however, the hydrophilic edge of porphyrin ring, adjacent to the phytol group, occurred in the hydrophilic region of a bovine serum albumin molecule.     

Preparation and properties of the water-soluble chlorophyll-bovine serum albumin complexes

By mixing chlorophyll (Chl) a or b with a dense bovine serum albumin solution, the water-soluble Chl-bovine serum albumin complexes were prepared. These complexes, eluted near the void volume on a gel filtration, were separated well from unreacted bovine serum albumin, indicating an aggregation of such molecules in the complexes. Preparation of chlorophyllide (Chlide) a- or Chlide b-bovine serum albumin complex was unsuccessful, while the phytol-, and β-carotene-bovine serum albumin complexes could be obtained. Chls in the Chl-bovine serum albumin complexes had the following characteristics. (i) Main absorption peak of Chl a or b in the red region occurred at 675 nm or 652 nm, respectively. The Chl a-bovine serum albumin complex having absorption peak at 740 nm was also prepared. As compared with the stabilities of Chl a and b in Triton X-100. (ii) Both Chls in the bovine serum albumin-complexes were stable against oxidative stresses, such as photobleaching, Fenton reagent, peroxidase-H₂O₂ system. But (iii) they were easily hydrolyzed by chlorophyllase. These properties of Chls in the bovine serum albumin-complexes were similar to those of Chls in the isolated light-harvesting Chl a/b protein complex. A possible localization of Chls within the bovine serum albumin complexes was suggested that the porphyrin moiety of Chl was buried in bovine serum albumin; however, the hydrophilic edge of porphyrin ring, adjacent to the phytol group, occurred in the hydrophilic region of a bovine serum albumin molecule.     

Fluorescence enhancing of 5,10,15,20-tetraphenylporphyrin in complex with human serum albumin and gold nanorods

Using absorption and fluorescence spectroscopy methods we obtained the results demonstrating alterations in spectral characteristics in supramolecular system composed of gold nanorods (AuNR) (10 × 38 nm) and complexes of human serum albumin (HSA) and 5,10,15,20-tetraphenylporphyrin (TPP). TPP fluorescence (λ_max = 636 and 658 nm) was found to enhance. The dependence of fluorescence enhancing in time was of nonlinear nature. Maximum TPP fluorescence enhancing value was as high as 16% and it was achieved in 7 min after mixing the components. Simultaneously with TPP fluorescence enhancing we observed a decrease in HSA own fluorescence (λ_max = 340 nm) and optical density reduction in maximum of longitudinal localized plasmon band of AuNR (λ_max = 752 nm).     

Resonance Rayleigh-scattering method for the determination of proteins with gold nanoparticle probe

Gold nanoparticles with a 12-nm diameter were used as probes for the determination of proteins by resonance Rayleigh-scattering techniques. In weak acidic solution, large amounts of citrate anions will self-assemble on the surface of positively charged gold nanoparticles to form supermolecular compounds with negative charges. Below the isoelectric point, proteins with positive charges such as human serum albumin (HSA), bovine serum albumin (BSA), and ovalbumin (Ova) can bind gold nanoparticles to form larger volume products (the diameter of the binding product of gold nanoparticles with HSA is 23 nm.) through electrostatic force, hydrogen bonds, and hydrophobic effects, which can result in a red shift of the maximum absorption wavelength, the remarkable enhancement of the resonance Rayleigh-scattering intensity (RRS), and the appearance of the RRS spectra. At the same time, the second-order-scattering (SOS) and frequency-doubling-scattering (FDS) intensities are also enhanced. The binding products of gold nanoparticles with different proteins have similar spectral characteristics and the maximum wavelengths are located near 303 nm for RRS, 540 nm for SOS, and 390 for FDS, respectively. The scattering enhancement (DeltaI) is directly proportional to the concentration of proteins. Among them, the RRS method has the highest sensitivity and the detection limits are 0.38 ng/ml for HSA, 0.45 ng/ml for BSA, and 0.56 ng/ml for Ova, separately. The methods have good selectivity. A new RRS method for the determination of trace proteins using a gold nanoparticle probe has been developed. Because gold nanoparticle probes do not need to be modified chemically in advance, the method is very simple and fast.     

A surface plasmon resonance probe with a novel integrated reference sensor surface

A surface plasmon resonance (SPR) sensor probe with integrated reference surface is described. In order to fabricate the integrated reference surface, two dielectric layers with different thickness were deposited on the single gold SPR sensor surface via plasma polymerization of hexamethyldisiloxane. The working sensor surface was a 34 nm dielectric layer with immobilized bovine serum albumin (BSA) antigen and an adjacent thin 1 nm dielectric layer without BSA provided reference surface. A specific immunoreaction of anti-BSA antibody was detected after immersion of the SPR probe into sample solution. Simultaneous observation of reference and working surface response enabled determination of the immunoreaction without the need for the baseline measurement. Moreover, compensation of nonspecific adsorption could be confirmed using anti-human serum albumin antibody.     

Plasmonic Au-MoO<Subscript>3</Subscript> Colloidal Nanoparticles by Reduction of HAuCl<Subscript>4</Subscript> by Blue MoO<Subscript>x</Subscript> Nanosheets and Observation of the Gasochromic Property

Defective colloids of blue MoO_x nanosheets were prepared by anodizing exfoliation method in water. This colloidal solution exhibits an optical plasmonic absorption band in the infrared region at about 760 nm. Merely mixing HAuCl₄ solution with the MoO_x leads to loss of the blue color, decaying of 760 nm plasmonic peak and simultaneous formation of the gold plasmon absorption peak at 550–570 nm. Some spectral variations in gold plasmonic peak and MoO_x optical band gap were observed for Mo:Au ratio of 10:1, 20:1, 30:1, and 40:1. The size of the gold nanoparticles was in the 5–6 nm range with fcc crystalline structure. X-ray photoelectron spectroscopy (XPS) revealed that the initial solution contains Mo⁵⁺ states and hydroxyl groups, which after reduction, hydroxyl groups are eliminated and the Mo⁵⁺ states converted to Mo⁶⁺. The obtained Au-MoO₃ colloids have a gasochromic property in which they are colored back to blue in the presence of hydrogen gas and the molybdenum oxide absorption peak recovered again. Furthermore, it was observed that both gold and Mo oxide plasmonic peaks redshift by insertion of hydrogen gas which is attributed to change in solution refractive index and formation of defect concentration.     

Controlled-release system of single-stranded DNA triggered by the photothermal effect of gold nanorods and its in vivo application

Gold nanorods have strong absorption bands in the near-infrared region, in which light penetrates deeply into tissues. The absorbed light energy is converted into heat by gold nanorods, the so-called 'photothermal effect'. Hence, gold nanorods are expected to act not only as on-demand thermal converters for photothermal therapy but also as controllers of a drug-release system responding to irradiation by near-infrared light. To achieve a controlled-release system that can be triggered by light irradiation, double-stranded DNA (dsDNA) was modified on gold nanorods. When the dsDNA-modified gold nanorods were irradiated by near-infrared light, the single-stranded DNA (ssDNA) was released from gold nanorods due to the photothermal effect. The amount of released ssDNA was dependent upon the power and exposure time of light irradiation. Release of ssDNA was also observed in tumors grown on mice after light irradiation. Such a controlled-release system of oligonucleotide triggered by the photothermal effect could expand the applications of gold nanorods that have unique optical characteristics in medicinal fields.     

Optical Saturable Absorption in Gold Nanoparticles

In this work, we present the experimental study of the nonlinear absorption of gold nanospheres and nanorods in aqueous suspension, using picosecond white-light supercontinuum open-aperture Z-scan. We demonstrate a saturable absorption effect in all particle suspensions at low-pulse energy. In the high-pulse energy regime, the apparent reverse-saturable absorption, observed in gold nanorods, was determined to be induced by photodegradation. Using the Lorentzian deconvolution method for the absorption spectra, we explain the variations on nonlinear optical effects and prove that saturable absorption only occurs within the plasmonic bands.     

Gold Nanowire and Nanorod Plasmonic Mechanisms for Increasing Ultra-Thin Organic Photovoltaic Active Layer Absorption

We theoretically investigate the effect of incorporating gold cylindrical- and ellipsoidal-shaped nanowires and gold nanorods situated centrally within the active layer of organic bulk-heterojunction photovoltaic devices, on the optical absorption performance using finite element electromagnetic simulations. Gold cylindrical nanowire-embedded devices show increased active layer absorption enhancement with increasing radius; however, this effect decreases with the introduction of a polystyrene dielectric capping layer around the nanowires. Active layer absorption, with respect to changes in the orientation, aspect ratio, periodicity, and spacing between ellipsoidal nanowires were optimized. A maximum absorption enhancement weighted by AM 1.5 solar spectrum of 17 % is predicted for gold ellipsoidal nanowires of aspect ratio of 1.167 with in-plane horizontal orientation and arranged with periodicity of 35 nm within a 30-nm thin active layer. We attribute this enhancement primarily to interparticle electromagnetic coupling between adjacent nanowires, where, a maximum spatial and spectral overlap of the electromagnetic field with the absorption band of the active layer material is achieved. This effect increases with decreasing aspect ratio as well as decreasing periodicity with a trade-off observed between nanowire packing density and the active layer absorption enhancement. For gold nanorod-embedded organic photovoltaic devices, the inter-particle electromagnetic coupling effects are weaker and longitudinal surface–plasmon resonances supported by the nanorods are more pronounced. However, since the longitudinal surface–plasmon resonances occur at wavelengths greater than the absorption edge of the photovoltaic active layer, a mere 3.4 % increase in absorption enhancement is achieved for the photovoltaic device incorporating gold nanorods with aspect ratio of 1.167 and periodicity of 35 nm.     

Analysis of colloidal gold probes by isoelectric focusing in agarose gels

Colloidal gold particles of different size (3-20 nm in diameter) were prepared by tannic acid-citrate and citrate reduction methods. From these colloids, different probes were prepared using sheep anti-rabbit antiserum, sheep anti-rabbit IgG, bovine serum albumin, polyethylene glycol, and protein A as the primary stabilizers and polyethylene glycol and/or bovine serum albumin as secondary and tertiary stabilizers, in different combinations. The probes were analyzed by isoelectric focusing in agarose gels, which allow the migration of particles in the size range 3-20 nm. (P. Sewer and S. J. Hayes, 1986, Anal. Biochem. 158, 72-78). Isoelectric focusing revealed that the surface charge of colloidal gold probes is dependent upon the size of the gold particle, the reduction method used, the primary ligand, and the pH at which this is adsorbed, as well as upon the secondary and tertiary stabilizers used. It is proposed that such differences in surface charge may underlie the different results which may sometimes be observed in colloidal gold labeling, especially when novel ligands are used.     

氨苄青霉素标记的胶体金颗粒的制备及活性研究

采用微波加热法制得15 nm的胶体金颗粒,并将小分子抗原氨苄青霉素分别以戊二醛和碳化二亚胺为偶联剂与牛血清蛋白偶联制成全抗原,再通过最佳标记量的确定分别将氨苄青霉素、戊二醛法全抗原、碳化二亚胺法全抗原同胶体金进行结合,红外检测和杯碟法的抑菌试验表明采用仅有戊二醛作为偶联剂的全抗原能够很好地与胶体金结合,并保持良好抗原活性。     

Zonal heterogeneity of rat hepatocytes in the in vivo uptake of 17 nm colloidal gold granules

The in vivo uptake in hepatocytes of intravenously injected colloidal gold granules with a diameter of 17 nm or 79 nm and coated with bovine serum albumin or with polyvinyl-pyrrolidone was studied. Irrespective of coating only the 17 nm granules were taken up in hepatocytes. Perivenous hepatocytes did take up much more gold granules than periportal hepatocytes. The gold granules were found in lysosomes around bile canaliculi. Two hours after injection hepatocytes contained the maximal amount of granules. At least a portion of the granules was discharged into the bile. The observed zonal gradient in the uptake of 17 nm gold granules might be caused by the greater supply of granules to the perivenous hepatocytes as a combined result of the higher porosity of the endothelial lining and the smaller number of Kupffer cells with a low endocytic activity in this zone.     

Zonal heterogeneity of rat hepatocytes in the in vivo uptake of 17 nm colloidal gold granules

Summary The in vivo uptake in hepatocytes of intravenously injected colloidal gold granules with a diameter of 17 nm or 79 nm and coated with bovine serum albumin or with polyvinyl-pyrrolidone was studied. Irrespective of coating only the 17 nm granules were taken up in hepatocytes. Perivenous hepatocytes did take up much more gold granules than periportal hepatocytes. The gold granules were found in lysosomes around bile canaliculi. Two hours after injection hepatocytes contained the maximal amount of granules. At least a portion of the granules was discharged into the bile. The observed zonal gradient in the uptake of 17 nm gold granules might be caused by the greater supply of granules to the perivenous hepatocytes as a combined result of the higher porosity of the endothelial lining and the smaller number of Kupffer cells with a low endocytic activity in this zone.     

Nonlinear Optical Properties of Large-Sized Gold Nanorods

The nonlinear optical properties of single gold nanorods (GNRs) with a large diameter of ～200 nm and a long length of ～800 nm were investigated by using a focused femtosecond (fs) laser light with tunable wavelength. While the linear and nonlinear optical properties of small-sized GNRs have been extensively studied, the nonlinear optical properties of large-sized GNRs and the effects of high-order surface plasmon resonances remain unexplored. Second harmonic generation (SHG) or/and two-photon-induced luminescence (TPL) were observed in the nonlinear response spectra, and their dependences on excitation wavelength and polarization were examined. The scattering and absorption spectra of the small- and large-sized GNRs were compared by using the discrete dipole approximation method. It was found that the extinction of large-sized GNRs is dominated by scattering rather than absorption, which is dominant in small-sized GNRs. In addition, it was revealed that the excitation wavelength-dependent SHG of a GNR is governed by the linear scattering of the GNR and the maximum SHG is achieved at the valley of the scattering spectrum. In comparison, the excitation wavelength dependence of TPL is determined by the absorption spectrum of the GNR. The polarization-dependent SHG of a GNR exhibits a strong dependence on the dimension of the GNR, and it may appear as bipolar distributions parallel or perpendicular to the long axis of the GNR or multipole distributions.     

Preparation of protein colloidal gold complexes in the presence of commonly used buffers

In the presence of various commonly used buffers, phosphate-buffered saline (PBS), tris-buffered saline (TBS), Na-cacodylate, bovine serum albumin and a wide range of cytochemically active proteins (monoclonal and polyclonal IgG, concanavalin A, Ricinus communis lectin I, Helix pomatia lectin, protein A) were complexed to colloidal gold of different particle sizes (6 nm, 9 nm, 22 nm). The resulting complexes were active in cytochemical labelling. Complex-formation in the presence of electrolyte opens the possibilities of: maintenance of ionic environment during complexing of proteins sensitive to low ionic strength, pH control by addition of buffers to the protein solution or to the gold sol, direct coupling of protein supplied in PBS or saline avoiding dialysis against low ionic strength buffers. Using the electron microscope to estimate the protein amounts needed for stabilization provided a sensitive and economical method to obtain aggregate-free protein-gold complexes.     

Absorption and fluorescence spectra of hypericin sodium salt in complexes with albumins

The spectra of absorption and fluorescence of hypericin sodium salt (Na-Hy) in organic solvents and in complexes with human serum albumin, bovine serum albumin, and lipoproteins of low and high density have been studied. It was shown that, as the proton donor properties of the solvent enhance, the absorption and fluorescence maxima shift toward the blue region, and as the proton-accepting properties increase, the maxima shift toward the red region. The absorption spectra of complexes of Na-Hy with bovine serum albumin significantly differ from those of complexes of this ligand with human serum albumin, which is evidenced by a lesser width of absorption bands and a lower value of the Stokes shift. The positions of the absorption and fluorescence maxima and the value of the Stokes shift for the complex of Na-Hy with human serum albumin increases when D₂O instead of common water is used as a solvent. It was concluded that H-bonds of hypericin play a significant role in the interaction with human serum albumin.     

Controlled release of PEG chain from gold nanorods: Targeted delivery to tumor

Gold nanorods exhibit strong absorbance of light in the near infrared region, which penetrates deeply into tissues. Since the absorbed light energy is converted into heat, gold nanorods are expected to act as a contrast agent for in vivo bioimaging and as a thermal converter for photothermal therapy. To construct a gold nanorod targeted delivery system for tumor a peptide substrate for urokinase-type plasminogen activator (uPA), expressed specifically on malignant tumors, was inserted between the PEG chain and the surface of the gold nanorods. In other words, we constructed PEG–peptide-modified gold nanorods. After mixing the gold nanorods with uPA, the PEG chain was released from the surface of the gold and subsequently nanorod aggregation took place. The formation of the aggregation was monitored as a decrease in light absorption at 900 nm. Tumor homogenate induced a significant decrease in this absorption. Larger amount of the PEG–peptide-modified gold nanorods bound to cells expressing uPA in vitro compared with control gold nanorods, which had scrambled sequence of the peptide. The PEG–peptide-modified gold nanorods showed higher accumulation in tumor than the control after they were injected intravenously into tumor-bearing mice, however, the density of the peptide on the surface of the gold nanorods was a key factor of their biodistributions. This targeted delivery system, which responds to uPA activity, is expected to be a powerful tool for tumor bioimaging and photothermal tumor therapy.     

Peptide-conjugated gold nanorods for nuclear targeting

Resonant electron oscillations on the surface of noble metal nanoparticles (Au, Ag, Cu) create the surface plasmon resonance (SPR) that greatly enhances the absorption and Rayleigh (Mie) scattering of light by these particles. By adjusting the size and shape of the particles from spheres to rods, the SPR absorption and scattering can be tuned from the visible to the near-infrared region (NIR) where biologic tissues are relatively transparent. Further, gold nanorods greatly enhance surface Raman scattering of adsorbed molecules. These unique properties make gold nanorods especially attractive as optical sensors for biological and medical applications. In the present work, gold nanorods are covalently conjugated with a nuclear localization signal peptide through a thioalkyl-triazole linker and incubated with an immortalized benign epithelial cell line and an oral cancer cell line. Dark field light SPR scattering images demonstrate that nanorods are located in both the cytoplasm and nucleus of both cell lines. Single cell micro-Raman spectra reveal enhanced Raman bands of the peptide as well as molecules in the cytoplasm and the nucleus. Further, the Raman spectra reveal a difference between benign and cancer cell lines. This work represents an important step toward both imaging and Raman-based intracellular biosensing with covalently linked ligand-nanorod probes.     

Surface Plasmon Dynamics of High-Aspect-Ratio Gold Nanorods

Ultrafast transient absorption studies are reported for high-aspect-ratio gold nanorods that were fabricated by electrochemical deposition in polycarbonate templates. The nanorods are 60 nm in diameter with distribution of lengths of up to 6 μm. The average aspect ratio was ∼50, resulting in a longitudinal surface plasmon resonance (SPR_L) band in the mid-IR, as well as a transverse (SPR_T) band in the visible. The rods were excited at 400 nm and probed at a range of wavelengths from the visible to the mid-IR to interrogate both SPR bands. The dynamics observed, including the electron–phonon coupling time and coherent acoustic breathing mode oscillations, closely resemble those previously reported for gold spherical nanoparticles and smaller-aspect-ratio nanorods. The electron–phonon coupling time was similarly determined to be 3.3 ± 0.2 ps for both of the SPR bands. Also, oscillations with a 32-ps period were observed for probing near the SPR_T band in the visible region due to impulsive coherent excitation of the acoustic breathing mode, which are consistent with the 60-nm diameter of the nanorods determined by scanning electron microscopy. The results demonstrate that the dynamics for long gold nanorods are similar to those for smaller nanoparticles. Gerald M. Sando is a NRL-ASEE Research Associate