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Hydrogel encapsulation is a promising carrier for cell and drug delivery due to its ability to protect the encapsulated entities from harsh physiological conditions and enhance their therapeutic efficacy and bioavailability. However, there is not yet consensus on the optimal hydrogel type, encapsulation method, and clinical application. Therefore, a systematic review of hydrogel encapsulation techniques and their potential for clinical application is needed to provide a comprehensive and up-to-date overview. In this systematic review, we searched electronic databases for articles published between 2008 and 2023 that described the encapsulation of cells or drug molecules within hydrogels. Herein, we identified 9 relevant studies that met the inclusion and exclusion criteria of our study. Our analysis revealed that the physicochemical properties of the hydrogel, such as its porosity, swelling behavior, and degradation rate, play a critical role in the encapsulation of cells or drug molecules. Furthermore, the encapsulation method, including physical, chemical, or biological methods, can affect the encapsulated entities' stability, bioavailability, and therapeutic efficacy. Challenges of hydrogel encapsulation include poor control over the release of encapsulated entities, limited shelf life, and potential immune responses. Future directions of hydrogel encapsulation include the development of novel hydrogel and encapsulation methods and the integration of hydrogel encapsulation with other technologies, such as 3D printing and gene editing. In conclusion, this review is useful for researchers, clinicians, and policymakers who are interested in this field of drug delivery and regenerative medicine that can serve as a guide for the future development of novel technologies that can be applied into clinical practice., Competing Interests: The authors declare no competing financial interest., (© 2024 The Authors. Published by American Chemical Society.)

Biocompatible and bioactive carbon-based nanocomposites are ingeniously designed and fabricated with the aim of enhancing drug delivery applicability in breast cancer treatment. Reduced graphene oxide (rGO) and multiwalled carbon nanotubes (MWCNTs) are utilized as nanocarriers for increasing penetrability into cells and the loading capacity. What sets our study apart is the strategic incorporation of the two different complexes of silver (AgL 2 ) and palladium (PdL 2 ) with the carboxamide-based ligand C 9 H 7 N 3 OS (L), which have been synthesized and decorated on nanocarriers alongside doxorubicin (DOX) for stabilizing DOX by π-π interactions and hydrogen bonding. Although DOX is a well-known cancer therapy agent, the efficacy of DOX is hindered owing to drug resistance, poor internalization, and limited site specificity. Aside from stabilizing DOX on nanocarriers, our carbon-based nanocarriers are tailored to act as a precision-guided missile, strategically by adorning with target-sensitive complexes. Based on the literature, carboxamide ligands can connect to overexpressed receptors on cancerous cells and inhibit them from proliferation signaling. Also, the complexes have an antibacterial activity that can control the infection caused by decreasing white blood cells and necrosis of cancerous cells. A high-concentration cytotoxicity assay revealed that decorating PdL 2 on a DOX-containing nanocarrier not only increased cytotoxicity to breast cancerous cell lines (MDA-MB-231 and MCF-7) but also revealed higher cell viability on a normal cell line (MCF-10A). The drug release screening results showed that the presence of PdL 2 led to 72 h correlate release behavior in acidic and physiological pH profiles, while the AgL 2- containing nanocomposite showed an analogue behavior for just 6 h and the release of DOX continued and after about 100 h hit the top., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

Liposomes have been extensively adopted in drug delivery systems with clinically approved formulations. However, hurdles remain in terms of loading multiple components and precisely controlling their release. Herein, we report a vesosomal carrier composed of liposomes encapsulated inside the core of another liposome for the controlled and sustained release of multiple contents. The inner liposomes are made of lipids with different compositions and are co-encapsulated with a photosensitizer. Upon induction of reactive oxygen species (ROS), the contents of the liposomes are released, with each type of liposome displaying distinct kinetics due to the variance in lipid peroxidation for differential structural deformation. In vitro experiments demonstrated immediate content release from ROS-vulnerable liposomes, followed by sustained release from ROS-nonvulnerable liposomes. Moreover, the release trigger was validated at the organismal level using Caenorhabditis elegans . This study demonstrates a promising platform for more precisely controlling the release of multiple components.

Detailed information about molecular interactions and conformational changes of polymeric components in the presence of ionic liquids (ILs) is essential for designing novel polymeric ionic liquid-based biomaterials. In biomaterials science and technology, thermoresponsive polymers (TRPs) are widely viewed as potential candidates for the fabrication of biorelated medical devices. Here, we synthesized thermoresponsive poly( N -vinyl-caprolactam) (PVCL) polymer and investigated the effects of imidazolium-based ILs (1-ethyl-3-methyl imidazolium nitrate and 1-butyl-3-methylimidazolium nitrate) with common anion and different cations on the phase transition behavior of PVCL aqueous solution. The impact of ILs on the phase transition behavior of PVCL was monitored by using UV-visible absorption spectra, steady-state fluorescence spectroscopy, thermal fluorescence spectroscopy, and temperature dependent dynamic light scattering. Results showed significant changes in the absorbance, molecular interactions, agglomeration, and coil to globule transition behaviors of PVCL in the presence of two ILs. PVCL aqueous solution showed significant conformational changes after the addition of ILs., Competing Interests: The authors declare no competing financial interest., (© 2022 The Authors. Published by American Chemical Society.)

Advances in microscale 3D cell culture systems have helped to elucidate cellular physiology, understand mechanisms of stem cell differentiation, produce pathophysiological models, and reveal important cell-cell and cell-matrix interactions. An important consideration for such studies is the choice of material for encapsulating cells and associated extracellular matrix (ECM). This Review focuses on the use of alginate hydrogels, which are versatile owing to their simple gelation process following an ionic cross-linking mechanism in situ, with no need for procedures that can be potentially toxic to cells, such as heating, the use of solvents, and UV exposure. This Review aims to give some perspectives, particularly to researchers who typically work more with poly(dimethylsiloxane) (PDMS), on the use of alginate as an alternative material to construct microphysiological cell culture systems. More specifically, this Review describes how physicochemical characteristics of alginate hydrogels can be tuned with regards to their biocompatibility, porosity, mechanical strength, ligand presentation, and biodegradability. A number of cell culture applications are also described, and these are subcategorized according to whether the alginate material is used to homogeneously embed cells, to micropattern multiple cellular microenvironments, or to provide an outer shell that creates a space in the core for cells and other ECM components. The Review ends with perspectives on future challenges and opportunities for 3D cell culture applications.

The beguiling world of functional polymers is dominated by thermoresponsive polymers with unique structural and molecular attributes. Limited work has been reported on the protein-induced conformational transition of block copolymers; furthermore, the literature lacks a clear understanding of the influence of proteins on the phase behavior of thermoresponsive copolymers. Herein, we have synthesized poly( N -isopropylacrylamide)- b -poly( N -vinylcaprolactam) (PNIPAM- b -PNVCL) by RAFT polymerization using N -isopropylacrylamide and N -vinylcaprolactam. Furthermore, using various biophysical techniques, we have explored the effect of cytochrome c (Cyt c), myoglobin (Mb), and hemoglobin (Hb) with varying concentrations on the aggregation behavior of PNIPAM- b -PNVCL. Absorption and steady-state fluorescence spectroscopy measurements were performed at room temperature to examine the copolymerization effect on fluorescent probe binding and biomolecular interactions between PNIPAM- b -PNVCL and proteins. Furthermore, temperature-dependent fluorescence spectroscopy and dynamic light scattering studies were performed to get deeper insights into the lower critical solution temperature (LCST) of PNIPAM- b -PNVCL. Small-angle neutron scattering (SANS) was also employed to understand the copolymer behavior in the presence of heme proteins. With the incorporation of proteins to PNIPAM- b -PNVCL aqueous solution, LCST has been varied to different extents owing to the preferential, molecular, and noncovalent interactions between PNIPAM- b -PNVCL and proteins. The present study can pave new insights between heme proteins and block copolymer interactions, which will help design biomimetic surfaces and aid in the strategic fabrication of copolymer-protein bioconjugates.

Selenium-enriched nickel selenide (NiSe-Se) nanotubes supported on highly conductive nickel foam (NiSe-Se@Ni foam) were synthesized using chemical bath deposition with the aid of lithium chloride as a shape-directing agent. The uniformly grown NiSe-Se@Ni foam, with its large number of electroactive sites, facilitated rapid diffusion and charge transport. The NiSe-Se@Ni foam electrode exhibited a superior specific capacitance value of 2447.46 F g -1 at a current density value of 1 A g -1 in 1 M aqueous KOH electrolyte. Furthermore, a high-energy-density pouch-type hybrid supercapacitor (HSC) device was fabricated using the proposed NiSe-Se@Ni foam as the positive electrode, activated carbon on Ni foam as the negative electrode, and a filter paper separator soaked in 1 M KOH electrolyte solution. The HSC delivered a specific capacitance of 84.10 F g -1 at a current density of 4 mA cm -2 with an energy density of 29.90 W h kg -1 at a power density of 594.46 W kg -1 for an extended operating voltage window of 1.6 V. In addition, the HSC exhibited excellent cycling stability with a capacitance retention of 95.09% after 10,000 cycles, highlighting its excellent potential for use in the hands-on applications. The real-life practicality of the HSC was tested by using it to power a red light-emitting diode.

A graphene aerogel (GA) with a three-dimensional (3D) structure, ultra-lightweight nature, and high hydrophobicity was simply fabricated by the one-step pyrolysis of glucose and ammonium chloride. The as-synthesized GA exhibited a 3D interconnected microporous architecture with a high surface area of ∼2860 m 2 g -1 and pore volume of 2.24 cm 3 g -1 . The hydrophobic GA (10 mg 100 mL -1 ) demonstrated rapid and excellent adsorption performance for the removal of food toxins such as various biogenic amines (histamine, cadaverine, and spermine) and the hazardous bacterium Staphylococcus aureus (a food contaminant and a cause of poor wound healing) from a liquid matrix with a maximum simultaneous adsorption capacity for multiple biogenic amines of >85.19% (histamine), 74.1% (cadaverine), and 70.11% (spermidine) and a 100% reduction in the viable cell count of S. aureus within 80 min of interaction. The outstanding adsorption capacity can be attributed to a highly interconnected porous network in the 3D architecture and a high surface-to-volume ratio. A case study using soy sauce spiked with multiple biogenic amines showed successful removal of toxins with excellent recyclability without any loss in absorption performance. Biocompatibility of the GA in terms of cell viability was observed even at high concentrations (83.46% and 75.28% at 25 and 50 mg mL -1 , respectively). Confirmatory biocompatibility testing was conducted via live/dead cell evaluation, and the morphology of normal lung epithelial cells was examined via scanning electron microscopy showed no cellular shrinkage. Moreover, GA showed excellent removal of live colonies of S. aureus from the food matrix and immunoblotting analysis showed elevated protein expression levels of β-catenin and α-SMA (α-smooth muscle actin). The biocompatible sugar-based GA could simultaneously adsorb multiple biogenic amines and live bacteria and was easy to regenerate via simple separation due to its high floatability, hydrophobicity, surface area, and porosity without any structural and functional loss, making it especially relevant for food safety and biomedical applications.

Although sodium hybrid capacitors (NHCs) have emerged as one of the most promising next-generation energy storage systems, further advancement is delayed primarily by the absence of high-performance battery-type anodes. Herein, we report a nature-inspired synthesis route to prepare hard carbon anodes with high capacity, rate capability, and cycle stability for dual-carbon NHCs. Shape- and size-controllable crystal aggregates of inexpensive triazine molecules are utilized as reactive templates that perform triple duties of structure-directing agent, porogen, and nitrogen source. This enables the fine control of microstructure/morphology/composition and thereby electrochemical reactions toward Na-ion. The resulting hard carbon optimized in terms of lateral size, interlayer spacing, and surface affinity of graphene-like layers achieves a specific capacity of ∼380 mAh/g after 100 cycles at a current density of 250 mA/g mainly via intercalation, the current record of hard carbons. Combined with a commercial microporous carbon fiber cathode, the full cell is able to deliver a volumetric energy density of 2.89 mWh/cm 3 and a volumetric power density of 160 mW/cm 3 , outperforming NHCs based on inorganic Na-ion anode materials. More importantly, such performance could not only be retained for 10000 cycles (4.5 F/cm 3 at 10 mA/cm 3 ) with 0.000 028 6% loss per cycle at >97% Coulombic efficiency but also successfully transferred to flexible pouch cells without significant performance loss after 300 bending cycles or during wrapping at a 10R condition. Simple preparation of hard carbon anodes using organic crystal reactive templates, therefore, demonstrates great potential for the manufacture of high-performance flexible NHCs using only carbon electrode materials.

The utilization of a sustainable and lightweight graphene aerogel (GA), synthesized from crude biomass, as a cell growth promoter and an adsorbent for the efficient removal of histamine (HIS), a food toxicant, from the real food matrix has been explored. Due to the self-supported three-dimensional nanoporous honeycomb-like structure of the graphene framework and the high surface area, the synthesized GA achieved an 80.69 ± 0.89% removal of HIS from red wine (spiked with HIS) after just 60 min under both acidic (3.0) and neutral (7.4) pH conditions. Furthermore, simple cleaning with 50% ethanol and deionized water, without any change in weight, allowed them to be reused more than 10 times with a still significant HIS removal ability (more than 71.6 ± 2.57%). In vitro cell culture experiments demonstrated that the synthesized GA had nontoxic effects on the cell viability (up to 80.35%) even at higher concentrations (10 mg mL -1 ), as determined via the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and lactate dehydrogenase assays using human lung bronchial epithelial cells. Interestingly, GA promotes the wound-healing ability on the scratched epithelial cell surfaces via enhancing the cell migrations as also validated by the western blot analysis via expression levels of epithelial β-catenin and E-cadherin proteins. The distinct structural advantage along with the nontoxicity of the green synthesized GA will not only facilitate the economic feasibility of the synthesized GA for its practical real-life applications in liquid toxin and pollutant removal from the food and environment but also broaden its applicability as a promising biomaterial of choice for biomedical applications.

The qualitative and quantitative evaluation of the physicochemical parameters associated with the pathogenicity of high-aspect-ratio nanomaterials is important for comprehensive regulation efforts and safety-by-design approaches. Here, we report quantitative data on the correlations between the rigidity of these nanomaterials and toxicity endpoints in vitro and in vivo. As measured by new ISO standards published in 2017, rigidity shows a strong positive correlation with inflammogenic potential, as indicated by inflammatory cell counts and IL-1β (a biomarker for frustrated phagocytosis) levels in both the acute and chronic phases. In vitro experiments using differentiated THP-1 cells find that only highly rigid multiwalled carbon nanotubes (MWCNTs) and asbestos fibers lead to piercing and frustrated phagocytosis. Thus, this study suggests a bending ratio of 0.97 and a static bending persistence length of 1.08 as threshold rigidity values for asbestos-like pathogenicity. However, additional research using MWCNTs with rigidity values that lie between those of non-inflammogenic ( D b = 0.66 and SBPL = 0.87) and inflammogenic fibers ( D b = 0.97 and SBPL = 1.09) is required to identify more accurate threshold values, which would be useful for comprehensive regulation and safety-by-design approaches based on MWCNTs.

Metal nanoparticle-containing porous organic polymers have gained great interest in chemical and pharmaceutical applications owing to their high reactivity and good recyclability. In the present work, a palladium nanoparticle-decorated triazine-urea-based porous organic polymer (Pd@TU-POP) was designed and synthesized using 1,3-bis(4-aminophenyl)urea with cyanuric chloride and palladium acetate. The porous structure and physicochemical properties of the electrode material Pd@TU-POP were observed using a range of standard techniques. The Pd@TU-POP material on the electrode surface showed superior sensing ability for rutin (RT) because the Pd dispersion facilitated the electrocatalytic performance of TU-POP by reducing the overpotential of RT oxidation dramatically and improving the stability significantly. Furthermore, TU-POP provides excellent structural features for loading Pd nanoparticles, and the resulting Pd@TU-POP exhibited enhanced electron transfer and outstanding sensing capability in a linear range between 2 and 200 pM having a low detection value of 5.92 × 10 -12 M (S/N = 3). The abundant porous structure of Pd@TU-POP not only provides electron transport channels for RT diffusion but also offers a facile route for quantification sensing of RT with satisfactory recoveries in aqueous electrolyte containing human plasma and red wine. These data reveal that the synthetic Pd@TU-POP is an excellent potential platform for the detection of RT in biological samples.

Curcumin-conjugated gold clusters (CUR-AuNCs) were synthesized using a "green" procedure and utilized as an anticancer and a bioimaging agent. Curcumin is a well-known anticancer agent, which forms a cluster when reacting with a gold precursor under mild alkali condition. A fluorescence spectroscopy analysis showed that the CUR-AuNCs emitted red fluorescence (650 nm) upon visible light (550) irradiation. Fourier transform infrared spectroscopy analysis confirmed the stretching and bending nature between the gold atoms and curcumin. Meanwhile, transmission electron microscopy analysis showed a cluster of approximately 1-3 nm with a uniform size. Time-resolved fluorescence analysis demonstrated that the red fluorescence was highly stable. Moreover, laser confocal imaging and atomic force microscopy analysis illustrated that a cluster was well distributed in the cell. This cluster exhibited less toxicity in the mortal cell line (COS-7) and high toxicity in the cervical cancer cell line (HeLa). The results demonstrated the conjugation of curcumin into the fluorescent gold cluster as a potential material for anticancer therapy and bioimaging applications.

Here, we report the fabrication of palladium nanoparticles on porous aromatic frameworks (Pd/PAF-6) using a facile chemical approach, which was characterized by various spectro- and electrochemical techniques. The differential pulse voltammetry (DPV) response of Pd/PAF-6 toward the vanillin (VA) sensor shows a linear relationship over concentrations (10-820 pM) and a low detection limit (2 pM). Pd/PAF-6 also exhibited good anti-interference performance toward 2-fold excess of ascorbic acid, nitrophenol, glutathione, glucose, uric acid, dopamine, ascorbic acid, 4-nitrophenol, glutathione, glucose, uric acid, dopamine, and 100-fold excess of Na(+), Mg(2+), and K(+) during the detection of VA. The developed electrochemical sensor based on Pd/PAF-6 had good reproducibility, as well as high selectivity and stability. The established sensor revealed that Pd/PAF-6 could be used to detect VA in biscuit and ice cream samples with satisfactory results.

A microporous covalent triazine polymer (CTP) network with a high surface area was synthesized via the Friedel-Crafts reaction and employed as a potential transport system for drug delivery and controlled release. The CTP was transformed to the nanoscale region by intense ultrasonication followed by filtration to yield nanoscale CTP (NCTP). This product showed excellent dispersibility in physiological solution while maintaining its chemical structure and porosity. An anticancer drug, doxorubicin (DOX), was loaded onto the NCTP through hydrophobic and π-π interactions, and its release was controlled at pH 4.8 and 7.4. The NCTP showed no toxicity toward cancer or normal cells, but the NCTP-DOX complex showed high efficacy against both types of cells in vitro. In-vitro cell imaging revealed that NCTP is a potential material for bioimaging. The potency of NCTP on cellular senescence was confirmed by the expression of senescence associated marker proteins p53 and p21. These results suggest that NCTP can be used as a new platform for drug delivery and imaging with potential applications in diagnosis and therapy.

We have developed a simple approach for the large-scale synthesis of water-soluble green carbon nanodots (G-dots) from many kinds of large food waste-derived sources. About 120 g of G-dots per 100 kg of food waste can be synthesized using our simple and environmentally friendly synthesis approach. The G-dots exhibit a high degree of solubility in water because of the abundant oxygen-containing functional groups around their surface. The narrow band of photoluminescence emission (400-470 nm) confirms that the size of the G-dots (∼4 nm) is small because of a similar quantum effects and emission traps on the surfaces. The G-dots have excellent photostability; their photoluminescence intensity decreases slowly (∼8%) under continuous excitation with a Xe lamp for 10 days. We carried out cell viability assay to assess the effect of cytotoxicity by introducing G-dots in cells such as Chinese hamster ovary cells (CHO-K1), mouse muscle cells (C2C12), and African green monkey kidney cells (COS-7), up to a concentration of 2 mg mL(-1) for 24 h. Due to their high photostability and low cytotoxicity, these G-dots are excellent probes for in vitro bioimaging. Moreover, the byproducts (not including G-dots) of G-dot synthesis from large food-waste derived sources promoted the growth and development of seedlings germinated on 3DW-supplemented gauze. Because of the combined advantages of green synthesis, high aqueous stability, high photostability, and low cytotoxicity, the G-dots show considerable promise in various areas, including biomedical imaging, solution state optoelectronics, and plant seed germination and/or growth.

We report a morphotropic phase transformation in vanadium dioxide (VO2) nanobeams annealed in a high-pressure hydrogen gas, which leads to the stabilization of metallic phases. Structural analyses show that the annealed VO2 nanobeams are hexagonal-close-packed structures with roughened surfaces at room temperature, unlike as-grown VO2 nanobeams with the monoclinic structure and with clean surfaces. Quantitative chemical examination reveals that the hydrogen significantly reduces oxygen in the nanobeams with characteristic nonlinear reduction kinetics which depend on the annealing time. Surprisingly, the work function and the electrical resistance of the reduced nanobeams follow a similar trend to the compositional variation due mainly to the oxygen-deficiency-related defects formed at the roughened surfaces. The electronic transport characteristics indicate that the reduced nanobeams are metallic over a large range of temperatures (room temperature to 383 K). Our results demonstrate the interplay between oxygen deficiency and structural/electronic phase transitions, with implications for engineering electronic properties in vanadium oxide systems.

Development of nanostructured materials with enhanced redox reaction capabilities is important for achieving high energy and power densities in energy storage systems. Here, we demonstrate that the nanohybridization of ionic liquids (ILs, 1-butyl-3-methylimidazolium tetrafluoroborate) and cobalt hydroxide (Co(OH)2) through ionothermal synthesis leads to a rapid and reversible redox reaction. The as-synthesized IL-Co(OH)2 has a favorable, tailored morphology with a large surface area of 400.4 m(2)/g and a mesopore size of 4.8 nm. In particular, the IL-Co(OH)2-based electrode exhibits improvement in electrochemical characteristics compared with bare Co(OH)2, showing a high specific capacitance of 859 F/g at 1 A/g, high-rate capability (∼95% retention at 30 A/g), and excellent cycling performance (∼96% retention over 1000 cycles). AC impedance analysis demonstrates that the introduction of ILs on Co(OH)2 facilitates ion transport and charge transfer: IL-Co(OH)2 shows a higher ion diffusion coefficient (1.06 × 10(-11) cm(2)/s) and lower charge transfer resistance (1.53 Ω) than those of bare Co(OH)2 (2.55 × 10(-12) cm(2)/s and 2.59 Ω). Our density functional theory (DFT) calculations reveal that the IL molecules, consisting of anion and cation groups, enable easier hydrogen desorption/adsorption process, that is, a more favorable redox reaction on the Co(OH)2 surface.

The palladium-catalyzed diacetoxylation and trifluoromethanesulfonic acid-catalyzed diacetoxylation using inexpensive and environmentally friendly hydrogen peroxide and peracetic acid were successfully conducted with the help of microchemical technology. Excellent yield and selectivity were achieved in significantly shortened reaction times without the decomposition of explosive oxidants and further transformation of unstable products, offering a safe and efficient alternative to traditional methods for alkene diacetoxylation.

In order to develop energy storage devices with high power and energy densities, electrodes should hold well-defined pathways for efficient ionic and electronic transport. Herein, we demonstrate high-performance supercapacitors by building a three-dimensional (3D) macroporous structure that consists of chemically modified graphene (CMG). These 3D macroporous electrodes, namely, embossed-CMG (e-CMG) films, were fabricated by using polystyrene colloidal particles as a sacrificial template. Furthermore, for further capacitance boost, a thin layer of MnO(2) was additionally deposited onto e-CMG. The porous graphene structure with a large surface area facilitates fast ionic transport within the electrode while preserving decent electronic conductivity and thus endows MnO(2)/e-CMG composite electrodes with excellent electrochemical properties such as a specific capacitance of 389 F/g at 1 A/g and 97.7% capacitance retention upon a current increase to 35 A/g. Moreover, when the MnO(2)/e-CMG composite electrode was asymmetrically assembled with an e-CMG electrode, the assembled full cell shows remarkable cell performance: energy density of 44 Wh/kg, power density of 25 kW/kg, and excellent cycle life.

Single nucleotide polymorphisms (SNPs) are one of the key diagnostic markers for genetic disease, cancer progression, and pharmcogenomics. The ligase detection reaction (LDR) is an excellent method to identify SNPs, combining low detection limits and high specificity. We present the first multiplex LDR-surface enhanced Raman spectroscopy (SERS) SNP genotyping scheme. The platform has the advantage in that the diagnostic peaks of Raman are more distinct than fluorescence, and in theory, a clinically significant number of markers can be multiplexed in a single sample using different SERS reporters. Here we report LDR-SERS multiplex SNP genotyping of K-Ras oncogene alleles at 10 pM detection levels, optimization of DNA labeling as well as Raman conditions, and the linear correlation of diagnostic peak intensity to SNP target concentration in heterozygous samples. Genomic DNA from typed cells lines was obtained and scored for the K-Ras genotype. These advances are significant as we have further developed our new SNP genotyping platform and have demonstrated the ability to correlate genotype ratios directly to diagnostic Raman peak signal intensity.

Genomics provides a comprehensive view of the complete genetic makeup of an organism. Individual sequence variations, as manifested by single nucleotide polymorphisms (SNPs), can provide insight into the basis for a large number of phenotypes and diseases including cancer. The ability rapidly screen for SNPs will have a profound impact on a number of applications, most notably personalized medicine. Here we demonstrate a new approach to SNP detection through the application of surface-enhanced Raman scattering (SERS) to the ligase detection reaction (LDR). The reaction uses two LDR primers, one of which contains a Raman enhancer and the other a reporter dye. In LDR, one of the primers is designed to interrogate the SNP. When the SNP being interrogated matches the discriminating primer sequence, the primers are ligated and the enhancer and dye are brought into close proximity enabling the dye's Raman signature to be detected. By detecting the Raman signature of the dye rather than its fluorescence emission, our technique avoids the problem of spectral overlap which limits number of reactions which can be carried out in parallel by existing systems. We demonstrate the LDR-SERS reaction for the detection of point mutations in the human K-ras oncogene. The reaction is implemented in an electrokinetically active microfluidic device that enables physical concentration of the reaction products for enhanced detection sensitivity and quantization. We report a limit of detection of 20 pM of target DNA with the anticipated specificity engendered by the LDR platform.