Efficient production of uniform gold nanoparticles via a streamlined low-cost, semi-automated, open-source platform.

In the quest to discover dependable and repeatable methods for producing noble metal nanospheres, both commercial and academic scientists have shown great interest. The challenge of precisely controlling the size of these nanospheres is critical, as variations can alter their optical characteristics, leading to complications in subsequent applications. In this context, we present the design and validation of an affordable, semi-automated device that synthesizes gold nanoparticles using the Turkevich method. This device, named 'NanoSynth Mini' and powered by Raspberry Pi, demonstrates the capability to generate gold nanoparticles with diameters ranging from 15 to 60 nanometers with minimal variability. Its design allows for seamless integration into lab processes, providing consistent support for extensive research initiatives.

3D Printed SERS-Active Thin-Film Substrates Used to Quantify Levels of the Genotoxic Isothiazolinone.

Several reports present methods to fabricate thin-film substrates capable of surface-enhanced Raman scattering (SERS). Substrates synthesized by displacing silver onto copper using facile synthesis methods such as galvanic displacement can generate high levels of SERS enhancement rivaling commercially available substrates manufactured by lithographic methods. Here, we describe the optimization of a novel set of SERS-active thin-film substrates synthesized via the electroless displacement of Ag onto the surface of three-dimensional (3D) printed disks composed of the copper/polymer (PLA) composite filament. The effect of AgNO3 concentration on the deposition, morphology, and overall SERS activity of the substrates has been carefully studied. Two commonly used Raman reporters, 4-mercaptobenzoic acid (MBA) and malachite green isothiocyanate (MGITC), were used to measure the SERS output of the substrates. Good SERS signal reproducibility (RSD ∼16.8%) was measured across the surface of replicate substrates and high-sensitivity detection of MBA was achieved (10-12 M). To test the real-world application of our substrates, we opted to detect 5-chloro-2-methyl-4-isothiazolin-3-one (CMIT), which is a genotoxic, biocide common in many household products, known to leach into water supplies. Our newly developed SERS-active substrates could detect CMIT down to 10 ppm when spiked in simulated lake water samples, which is well within current agency standards.

Synthesis of SERS-active core-satellite nanoparticles using heterobifunctional PEG linkers.

Surface-enhanced Raman scattering (SERS) is a sensitive analytical technique capable of magnifying the vibrational intensity of molecules adsorbed onto the surface of metallic nanostructures. Various solution-based SERS-active metallic nanostructures have been designed to generate substantial SERS signal enhancements. However, most of these SERS substrates rely on the chemical aggregation of metallic nanostructures to create strong signals. While this can induce high SERS intensities through plasmonic coupling, most chemically aggregated assemblies suffer from poor signal reproducibility and reduced long-term stability. To overcome these issues, here we report for the first time the synthesis of gold core-satellite nanoparticles (CSNPs) for robust SERS signal generation. The novel CSNP assemblies consist of a 30 nm spherical gold core linked to 18 nm satellite particles via linear heterobifunctional thiol-amine terminated PEG chains. We explore the effects that the varying chain lengths have on SERS hot-spot generation, signal reproducibility and long-term activity. The chain length was varied by using PEGs with different molecular weights (1000 Da, 2000 Da, and 3500 Da). The CSNPs were characterized via UV-Vis spectrophotometry, transmission electron microscopy (TEM), ζ-potential measurements, and lastly SERS measurements. The versatility of the synthesized SERS-active CSNPs was revealed through characterization of optical stability and SERS enhancement at 0, 1, 3, 5, 7 and 14 days.

Epigallocatechin Gallate-Gold Nanoparticles Exhibit Superior Antitumor Activity Compared to Conventional Gold Nanoparticles: Potential Synergistic Interactions.

Epigallocatechin gallate (EGCG) possesses significant antitumor activity and binds to laminin receptors, overexpressed on cancer cells, with high affinity. Gold nanoparticles (GNPs) serve as excellent drug carriers and protect the conjugated drug from enzymatic metabolization. Citrate-gold nanoparticles (C-GNPs) and EGCG-gold nanoparticles (E-GNPs) were synthesized by reduction methods and characterized with UV-visible spectroscopy, transmission electron microscopy (TEM), and dynamic light scattering (DLS). Cytotoxicity of citrate, EGCG, C-GNPs, and E-GNPs was evaluated by the water-soluble tetrazolium salt (WST-1) assay. Nanoparticle cellular uptake studies were performed by TEM and atomic absorption spectroscopy (AAS). Dialysis method was employed to assess drug release. Cell viability studies showed greater growth inhibition by E-GNPs compared to EGCG or C-GNPs. Cellular uptake studies revealed that, unlike C-GNPs, E-GNPs were taken up more efficiently by cancerous cells than noncancerous cells. We found that E-GNP nanoformulation releases EGCG in a sustained fashion. Furthermore, data showed that E-GNPs induced more apoptosis in cancer cells compared to EGCG and C-GNPs. From the mechanistic standpoint, we observed that E-GNPs inhibited the nuclear translocation and transcriptional activity of nuclear factor-kappaB (NF-κB) with greater potency than EGCG, whereas C-GNPs were only minimally effective. Altogether, our data suggest that E-GNPs can serve as potent tumor-selective chemotoxic agents.

Drug-loaded exosomal preparations from different cell types exhibit distinctive loading capability, yield, and antitumor efficacies: a comparative analysis.

BACKGROUND: Despite tremendous advancement, cancer still remains one of the leading causes of death worldwide. Inefficiency of current drug delivery regimens is one important factor that limits the therapeutic efficacy of existing drugs, thus contributing to cancer mortality. To address this limitation, synthetic nanotechnology-based delivery systems have been developed; however, they raise concern of inducing adverse immunogenic reactions. Exosomes (Exos) are nonimmunogenic nanosized vesicles that have received significant attention as efficient drug delivery system. METHODS: Drug loading in Exos were achieved by incubating different cell types viz pancreatic cancer cells (PCCs), pancreatic stellate cells (PSCs), and macrophages (MØs) with Doxorubicin (DOX). Differential ultracentrifugation was performed to isolate exosome and their size was determined by dynamic light scattering analysis. The efficacy of drug packaging into Exos was evaluated by HPLC. Flow cytometry was performed to examine the apoptosis. Cell viability was determined using the WST-1 assay. RESULTS: PCCs shed the most Exos and were the most efficient in drug loading followed by MØs and PSCs as examined by HPLC quantification. However, when compared for antitumor efficacy, MØ-derived Exos loaded with DOX (MØ-Exo-DOX) showed highest activity followed by PSCs and PCCs. CONCLUSION: These varying antitumor activities likely resulted from nondrug contents of Exos since we did not observe any significant differences in their uptake by the cancer cells. Altogether, our data suggest that donor cell-specific differences exist in Exos, which could influence their utility as drug carrier for therapeutic purposes.

Development of a SERS Probe for Selective Detection of Healthy Prostate and Malignant Prostate Cancer Cells Using ZnII.

Even in the 21st century, prostate cancer remains the second leading cause of cancer-related death for men. Since a normal prostate gland has a high ZnII content and there are huge differences in ZnII content between healthy and malignant prostate cancer cells, mobile zinc can be used as a biomarker for prostate cancer prediction. A highly efficient surface enhanced Raman spectroscopy (SERS) probe using a p-(imidazole)azo)benzenethiol attached gold nanoparticle as a Raman reporter, which has the capability to identify prostate cancer cells based on ZnII sensing, has been designed. A facile synthesis, characterization and evaluation of a ZnII sensing Raman probe are described. Reported data indicate that after binding with ZnII , Raman reporter attached to a gold nanoparticle forms an assembly structure, which allows selective detection of ZnII even at 100 ppt concentration. Theoretical full-wave finite-difference time-domain (FDTD) simulations have been used to understand the enhancement of the SERS signal. The SERS probe is highly promising for in vivo sensing of cancer, where near-IR light can be easily used to avoid tissue autofluorescence and to enhance tissue penetration depth. Reported data show that the SERS probe can distinguish metastatic cancer cells from normal prostate cells very easily with a sensitivity as low as 5 cancer cells mL-1 . The probe can be used as a chemical toolkit for determining mobile ZnII concentrations in biological samples.

Designing a multicolor long range nanoscopic ruler for the imaging of heterogeneous tumor cells.

Tumor heterogeneity is one of the biggest challenges in cancer treatment and diagnosis. A multicolor optical ruler is essential to address the heterogeneous tumor cell complexity. Driven by this need, the current article reports the design of a multicolor long range nanoscopic ruler for screening tumor heterogeneity by accurately identifying epithelial cells and cancer stem cells (CSCs) simultaneously. A nanoscopic surface energy transfer (NSET) ruler has been developed using blue fluorescence polymer dots (PDs) and red fluorescence gold cluster dots (GCDs) as multicolor fluorescence donor and plasmonic gold nanoparticle (GNP) acts as an excellent acceptor. Reported experimental results demonstrated that the multicolor nanoscopic ruler's working window is above 35 nm distances, which is more than three times farther than that of Förster resonance energy transfer (FRET) distance limit. Theoretical modeling using Förster dipole-dipole coupling and dipole to nanoparticle surface energy transfer have been used to discuss the possible mechanism for multicolor nanoscopic ruler's long-range capability. Using RNA aptamers that are specific for the target cancer cells, experimental data demonstrate that the nanoscopic ruler can be used for screening epithelial and CSCs simultaneously from a whole blood sample with a detection capability of 10 cells per mL. Experimental data show that the nanoscopic ruler can distinguish targeted cells from non-targeted cells.

Development of Multifunctional Fluorescent-Magnetic Nanoprobes for Selective Capturing and Multicolor Imaging of Heterogeneous Circulating Tumor Cells.

Circulating tumor cells (CTC) are highly heterogeneous in nature due to epithelial-mesenchymal transition (EMT), which is the major obstacle for CTC analysis via "liquid biopsy". This article reports the development of a new class of multifunctional fluorescent-magnetic multicolor nanoprobes for targeted capturing and accurate identification of heterogeneous CTC. A facile design approach for the synthesis and characterization of bioconjugated multifunctonal nanoprobes that exhibit excellent magnetic properties and emit very bright and photostable multicolor fluorescence at red, green, and blue under 380 nm excitation is reported. Experimental data presented show that the multifunctional multicolor nanoprobes can be used for targeted capture and multicolor fluorescence mapping of heterogeneous CTC and can distinguish targeted CTC from nontargeted cells.

Multimodal Nonlinear Optical Imaging of Live Cells Using Plasmon-Coupled DNA-Mediated Gold Nanoprism Assembly.

Multiphoton excitation microscopy techniques are the emerging nonlinear optical (NLO) imaging methods to watch the biological world due its ability to penetrate deep into living tissues. Driven by the need to develop multimodal NLO imaging probe, current article reports the design of DNA-mediated gold nanoprisms assembly based optical antennas to enhance multiphoton imaging capability in biological II window. Reported experimental data show a unique way to enhance second harmonic generation (SHG) and two-photon fluorescence (TPF) properties by several orders of magnitudes via plasmon coupled organization into gold nanoprism assembly structures. Experimental and theoretical modeling data using finite difference time domain (FDTD) simulations indicate that huge enhancement of SHG and TPF properties are mainly due to the electric quadrupole contribution and electric field enhancement. Using 1100 nm biological II window light, reported results demonstrated that antibody conjugated assembly structures are capable of exhibiting highly selective and very bright multimodal SHG and TPF imaging of human Hep G2 liver cancer cells.

Hybrid Theranostic Platform for Second Near-IR Window Light Triggered Selective Two-Photon Imaging and Photothermal Killing of Targeted Melanoma Cells.

Despite advances in the medical field, even in the 21st century cancer is one of the leading causes of death for men and women in the world. Since the second near-infrared (NIR) biological window light between 950 and 1350 nm offers highly efficient tissue penetration, the current article reports the development of hybrid theranostic platform using anti-GD2 antibody attached gold nanoparticle (GNP) conjugated, single-wall carbon nanotube (SWCNT) for second near-IR light triggered selective imaging and efficient photothermal therapy of human melanoma cancer cell. Reported results demonstrate that due to strong plasmon-coupling, two-photon luminescence (TPL) intensity from theranostic GNP attached SWCNT materials is 6 orders of magnitude higher than GNP or SWCNT alone. Experimental and FDTD simulation data indicate that the huge enhancement of TPL intensity is mainly due to strong resonance enhancement coupled with the stronger electric field enhancement. Due to plasmon coupling, the theranostic material serves as a local nanoantennae to enhance the photothermal capability via strong optical energy absorption. Reported data show that theranostic SWCNT can be used for selective two-photon imaging of melanoma UACC903 cell using 1100 nm light. Photothermal killing experiment with 1.0 W/cm(2) 980 nm laser light demonstrates that 100% of melanoma UACC903 cells can be killed using theranostic SWCNT bind melanoma cells after just 8 min of exposure. These results demonstrate that due to plasmon coupling, the theranostic GNP attached SWCNT material serves as a two-photon imaging and photothermal source for cancer cells in biological window II.

Antimicrobial Peptide-Conjugated Graphene Oxide Membrane for Efficient Removal and Effective Killing of Multiple Drug Resistant Bacteria.

According to the World Health Organization (WHO), multiple drug-resistant (MDR) bacterial infection is a top threat to human health. Since bacteria evolve to resist antibiotics faster than scientists can develop new classes of drugs, the development of new materials which can be used, not only for separation, but also for effective disinfection of drug resistant pathogens is urgent. Driven by this need, we report for the first time the development of a nisin antimicrobial peptide conjugated, three dimensional (3D) porous graphene oxide membrane for identification, effective separation, and complete disinfection of MDR methicillin-resistant Staphylococcus aureus (MRSA) pathogens from water. Experimental data show that due to the size differences, MRSA is captured by the porous membrane, allowing only water to pass through. SEM, TEM, and fluorescence images confirm that pathogens are captured by the membrane. RT-PCR data with colony counting indicate that almost 100% of MRSA can be removed and destroyed from the water sample using the developed membrane. Comparison of MDR killing data between nisin alone, the graphene oxide membrane and the nisin attached graphene oxide membrane demonstrate that the nisin antimicrobial peptide attached graphene oxide membrane can dramatically enhance the possibility of destroying MRSA via a synergestic effect due to the multimodal mechanism.

Hybrid Graphene Oxide Based Plasmonic-Magnetic Multifunctional Nanoplatform for Selective Separation and Label-Free Identification of Alzheimer's Disease Biomarkers.

Despite intense efforts, Alzheimer's disease (AD) is one of the top public health crisis for society even at 21st century. Since presently there is no cure for AD, early diagnosis of possible AD biomarkers is crucial for the society. Driven by the need, the current manuscript reports the development of magnetic core-plasmonic shell nanoparticle attached hybrid graphene oxide based multifunctional nanoplatform which has the capability for highly selective separation of AD biomarkers from whole blood sample, followed by label-free surface enhanced Raman spectroscopy (SERS) identification in femto gram level. Experimental ELISA data show that antibody-conjugated nanoplatform has the capability to capture more than 98% AD biomarkers from the whole blood sample. Reported result shows that nanoplatform can be used for SERS "fingerprint" identification of ß-amyloid and tau protein after magnetic separation even at 100 fg/mL level. Experimental results indicate that very high sensitivity achieved is mainly due to the strong plasmon-coupling which generates huge amplified electromagnetic fields at the "hot spot". Experimental results with nontargeted HSA protein, which is one of the most abundant protein components in cerebrospinal fluid (CSF), show that multifunctional nanoplatform based AD biomarkers separation and identification is highly selective.

Multifunctional biocompatible graphene oxide quantum dots decorated magnetic nanoplatform for efficient capture and two-photon imaging of rare tumor cells.

Circulating tumor cells (CTCs) are extremely rare cells in blood containing billions of other cells. The selective capture and identification of rare cells with sufficient sensitivity is a real challenge. Driven by this need, this manuscript reports the development of a multifunctional biocompatible graphene oxide quantum dots (GOQDs) coated, high-luminescence magnetic nanoplatform for the selective separation and diagnosis of Glypican-3 (GPC3)-expressed Hep G2 liver cancer tumor CTCs from infected blood. Experimental data show that an anti-GPC3-antibody-attached multifunctional nanoplatform can be used for selective Hep G2 hepatocellular carcinoma tumor cell separation from infected blood containing 10 tumor cells/mL of blood in a 15 mL sample. Reported data indicate that, because of an extremely high two-photon absorption cross section (40530 GM), an anti-GPC3-antibody-attached GOQDs-coated magnetic nanoplatform can be used as a two-photon luminescence platform for selective and very bright imaging of a Hep G2 tumor cell in a biological transparency window using 960 nm light. Experimental results with nontargeted GPC3(-) and SK-BR-3 breast cancer cells show that multifunctional-nanoplatform-based cell separation, followed by two-photon imaging, is highly selective for Hep G2 hepatocellular carcinoma tumor cells.

Aptamer-conjugated graphene oxide membranes for highly efficient capture and accurate identification of multiple types of circulating tumor cells.

Tumor metastasis is responsible for 1 in 4 deaths in the United States. Though it has been well-documented over past two decades that circulating tumor cells (CTCs) in blood can be used as a biomarker for metastatic cancer, there are enormous challenges in capturing and identifying CTCs with sufficient sensitivity and specificity. Because of the heterogeneous expression of CTC markers, it is now well understood that a single CTC marker is insufficient to capture all CTCs from the blood. Driven by the clear need, this study reports for the first time highly efficient capture and accurate identification of multiple types of CTCs from infected blood using aptamer-modified porous graphene oxide membranes. The results demonstrate that dye-modified S6, A9, and YJ-1 aptamers attached to 20-40 µm porous garphene oxide membranes are capable of capturing multiple types of tumor cells (SKBR3 breast cancer cells, LNCaP prostate cancer cells, and SW-948 colon cancer cells) selectively and simultaneously from infected blood. Our result shows that the capture efficiency of graphene oxide membranes is ~95% for multiple types of tumor cells; for each tumor concentration, 10 cells are present per milliliter of blood sample. The selectivity of our assay for capturing targeted tumor cells has been demonstrated using membranes without an antibody. Blood infected with different cells also has been used to demonstrate the targeted tumor cell capturing ability of aptamer-conjugated membranes. Our data also demonstrate that accurate analysis of multiple types of captured CTCs can be performed using multicolor fluorescence imaging. Aptamer-conjugated membranes reported here have good potential for the early diagnosis of diseases that are currently being detected by means of cell capture technologies.

Long-range two-photon scattering spectroscopy ruler for screening prostate cancer cells.

Optical rulers have served as a key tool for scientists from different disciplines to address a wide range of biological activity. Since the optical window of state of the art FRET rulers is limited to a 10 nm distance, developing long range optical rulers is very important to monitor real life biological processes. Driven by this need, the current manuscript reports for the first time the design of long-range two-photon scattering (TPS) spectroscopy rulers using gold nano-antenna separated by a bifunctional rigid double strand DNA molecule, which controls the spectroscopy ruler length. Reported data demonstrate that the TPS spectroscopy ruler's working window is a within a 25 nm distance, which is more than twice that of well recognized FRET optical ruler. A possible mechanism for the two-photon spectroscopy ruler's long range capability have been discussed using angle-resolved TPS measurement and FDTD simulations. Solution-phase experimental data demonstrated that a long-range TPS ruler using A9 aptamer can be used for the screening of prostate-specific membrane antigen (PSMA) (+) prostate cancer cells even at 5 cells per mL level. Reported result with PSMA (-) normal skin HaCaT cells indicate that TPS ruler based assay has the capability to enable distinction from non-targeted cell lines. Ultimately, the long range TPS ruler can be used towards better understanding of chemical and biological processes.

Aptamer-conjugated theranostic hybrid graphene oxide with highly selective biosensing and combined therapy capability.

Cancer is a life-threatening disease, which is rapidly becoming a global pandemic. Driven by this need, here we report for the first time an aptamer-conjugated theranostic magnetic hybrid graphene oxide-based assay for highly sensitive tumor cell detection from blood samples with combined therapy capability. AGE-aptamer-conjugated theranostic magnetic nanoparticle-attached hybrid graphene oxide was developed for highly selective detection of tumor cells from infected blood samples. Experimental data indicate that hybrid graphene can be used as a multicolor luminescence platform for selective imaging of G361 human malignant melanoma cancer cells. The reported results have also shown that indocyanine green (ICG)-bound AGE-aptamer-attached hybrid graphene oxide is capable of combined synergistic photothermal and photodynamic treatment of cancer. Targeted combined therapeutic treatment using 785 nm near-infrared (NIR) light indicates that the multimodal therapeutic treatment is highly effective for malignant melanoma cancer therapy. The reported data show that this aptamer-conjugated theranostic graphene oxide-based assay has exciting potential for improving cancer diagnosis and treatment.

Highly efficient and excitation tunable two-photon luminescence platform for targeted multi-color MDRB imaging using graphene oxide.

Multiple drug-resistance bacteria (MDRB) infection is one of the top three threats to human health according to the World Health Organization (WHO). Due to the large penetration depth and reduced photodamage, two-photon imaging is an highly promising technique for clinical MDRB diagnostics. Since most commercially available water-soluble organic dyes have low two-photon absorption cross-section and rapid photobleaching tendency, their applications in two-photon imaging is highly limited. Driven by the need, in this article we report extremely high two-photon absorption from aptamer conjugated graphene oxide (σ2PA = 50800 GM) which can be used for highly efficient two-photon fluorescent probe for MDRB imaging. Reported experimental data show that two-photon photoluminescence imaging color, as well as luminescence peak position can be tuned from deep blue to red, just by varying the excitation wavelength without changing its chemical composition and size. We have demonstrated that graphene oxide (GO) based two-photon fluorescence probe is capable of imaging of multiple antibiotics resistance MRSA in the first and second biological transparency windows using 760-1120 nm wavelength range.

Extremely High Two-Photon Absorbing Graphene Oxide for Imaging of Tumor Cells in the Second Biological Window.

Cancer, a life-threatening disease, has become a global pandemic. Targeted tumor imaging using near-infrared (NIR) light is the key to improve the penetration depth and it is highly promising for clinical tumor diagnostics. Driven by this need, in this Letter we have reported aptamer conjugated graphene oxide-based two-photon imaging of breast tumor cells selectively. Reported data indicate that there is an extremely high two-photon absorption from aptamer conjugated graphene oxide (σ2PA = 46890 GM). Experimental data show that two-photon luminescence signal remains almost unchanged even after 2 h of illuminations. Reported results show that S6 RNA aptamers conjugated graphene oxide-based two-photon fluorescence can be used for selective two-photon imaging of SK-BR-3 breast tumor cell in second biological transparency windows using 1100 nm wavelength. Experimental data demonstrate that it is highly capable of distinguishing targeted breast cancer SK-BR-3 cells from other nontargeted MDA-MB-231 breast cancer cells.

Accurate Identification and Selective Removal of Rotavirus Using a Plasmonic-Magnetic 3D Graphene Oxide Architecture.

According to the World Health Organization, even in the 21st century, more than one million children die each year due to the rotavirus contamination of drinking water. Therefore, accurate identification and removal of rotavirus are very important to save childrens' lives. Driven by the need, in this Letter, we report for the first time highly selective identification and removal of rotavirus from infected water using a bioconjugated hybrid graphene oxide based three-dimensional (3D) solid architecture. Experimental results show that due to the presence of a high intensity of "hot spots" in the 3D network, an antibody-attached 3D plasmonic-magnetic architecture can be used for accurate identification of rotavirus using surface-enhanced Raman spectroscopy (SERS). Reported data demonstrate that the antibody-attached 3D network binds strongly with rotavirus and is capable of highly efficient removal of rotavirus, which has been confirmed by SERS, fluorescence imaging, and enzyme-linked immunosorbent assay (ELISA) data. We discuss a possible mechanism for accurate identification and efficient removal of rotavirus from infected drinking water.