Serotype-specific identification of Dengue virus by silicon nanowire array biosensor.

In this work, we demonstrated a silicon nanowire (SiNW) biosensing platform capable of simultaneously identifying different Dengue serotypes on a single sensing chip. Four peptide nucleic acids (PNAs), specific to each Dengue serotypes (DENV-1 to DENV-4), were spotted on different areas of the SiNW array surface, and the covalently immobilized PNA probes were then interacted with different Dengue serotypes target to establish the specificity of detection. Detection scheme is based on the changes in resistances due to accumulation of negative charges contributed by the hybridized DNA target. The results show that resistance changes only occur in regions where the Dengue target hybridizes with its complementary probe. What is more, a mixture of two different Dengue serotypes obtained from a one-step duplex RT-PCR was applied to the multiplex SiNW surface to validate SiNW capability to identify multiple Dengue serotypes on a single sensing platform. Through this study, we have established the multiplex SiNW biosensor as a promising device to detect multiple Dengue infections with high specificity.

Label-free detection of carbohydrate-protein interactions using nanoscale field-effect transistor biosensors.

Carbohydrate-protein interactions play a significant role in cell communication, cell adhesion, cell trafficking, and immune responses. Many efforts have been made to demonstrate detection of carbohydrate-protein interactions. However, the existing methods are still tedious and expensive. Therefore, the detection of carbohydrate-protein interactions is of great significance, and new, efficient methods are required for fast and sensitive recognition testing. In this report, we, for the first time, developed the silicon nanowire (SiNW)-based biosensor capable of label-free electrical detection of carbohydrate-protein interactions with high selectivity and sensitivity by covalently immobilizing unmodified carbohydrates on the sensor surface. We fabricated new SiNW sensor chips with more SiNW arrays for potential detection of multiple analytes. In order to realize the immobilization of the unmodified carbohydrates on the SiNW surface, we used X-ray photoelectron spectra and fluorescence microscopy to verify the successful surface functionalization on the silicon surface. Furthermore, we demonstrated real-time detection of carbohydrate-protein interactions using the carbohydrate-modified SiNW sensor chips. The results show good specificity between galactose-lectin EC and mannose-Con A, which is in good agreement with that reported previously. Finally, the results also show that we are able to use the galactose-modified SiNW biosensor to detect lectin EC as low as 100 fg/mL, which is 4 orders of magnitude lower than that reported by other technologies. We believe that the developed SiNW biosensor paves a novel way for studying carbohydrate-protein interactions.

An integrated chip for rapid, sensitive, and multiplexed detection of cardiac biomarkers from fingerprick blood.

Cardiovascular diseases are the major cause of death among adults worldwide. Electrocardiogram (ECG) is a first test when a patient suffering from chest pain sees a doctor, however, it is lack of the required sensitivity. Standard assays to detect cardiac biomarkers, like enzyme-linked immunosorbent assay (ELISA) are sensitive, but suffer from important sample and reagent consumption in large-scale studies. Moreover they are performed in central laboratories of clinics and hospitals and take a long time, which is highly incompatible with the quick decisions needed to save a heart attack patient. Herein, we describe an integrated chip allowing rapid, sensitive, and simultaneous analysis of three cardiac biomarkers in fingerprick blood. The integrated chip is composed of a filtration chip for plasma separation from blood and a silicon nanowire (SiNW) array sensor chip for protein detection. These two chips are fabricated separately and bonded to form a single unit after alignment. The integrated chip is capable of reducing the dead volume of the sample by eliminating the tubing between the two chips. After the plasma is filtrated by the filtration chip, the SiNW sensor, spotted with three different antibodies, enabled us to detect three cardiac biomarkers, troponin T (cTnT), creatine kinase MM (CK-MM) and creatine kinase MB (CK-MB), simultaneously. The integrated chip is able to attain a low detection limit of 1 pg/ml for the three cardiac biomarkers from 2 µl blood in 45 min.

Self-assembled monolayer-assisted silicon nanowire biosensor for detection of protein-DNA interactions in nuclear extracts from breast cancer cell.

The large number of estrogen receptor (ER) binding sites of various sequence patterns requires a sensitive detection to differentiate between subtle differences in ER-DNA binding affinities. A self-assembled monolayer (SAM)-assisted silicon nanowire (SiNW) biosensor for specific and highly sensitive detection of protein-DNA interactions, remarkably in nuclear extracts prepared from breast cancer cells, is presented. As a typical model, estrogen receptor element (ERE, dsDNA) and estrogen receptor alpha (ERα, protein) binding was adopted in the work. The SiNW surface was coated with a vinyl-terminated SAM, and the termination of the surface was changed to carboxylic acid via oxidation. DNA modified with amine group was subsequently immobilized on the SiNW surface. Protein-DNA binding was finally investigated by the functionalized SiNW biosensor. X-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM) were employed to characterize the stepwise functionalization of the SAM and DNA on bare silicon surface, and to visualize protein-DNA binding on the SiNW surface, respectively. We observed that ERα had high sequence specificity to the SiNW biosensor which was functionalized with three different EREs including wild-type, mutant and scrambled DNA sequences. We also demonstrate that the specific DNA-functionalized SiNW biosensor was capable of detecting ERα as low as 10 fM. Impressively, the developed SiNW biosensor was able to detect ERα-DNA interactions in nuclear extracts from breast cancer cells. The SAM-assisted SiNW biosensor, as a label-free and highly sensitive tool, shows a potential in studying protein-DNA interactions.

Highly sensitive and reversible silicon nanowire biosensor to study nuclear hormone receptor protein and response element DNA interactions.

To thoroughly understand the role that estrogen receptors partake in regulation of gene expression, characterization of estrogen receptors (ERs) and estrogen-response elements (EREs) interactions is essential. In the work, we present a highly sensitive and reusable silicon nanowire (SiNW) biosensor to study the interactions between human ER proteins (ER, α and ß subtypes) and EREs (dsDNA). The proteins were covalently immobilized on the SiNW surface. Various EREs including wild-type, mutant and scrambled DNA sequences were then applied to the protein-functionalized SiNW surface. Due to negatively charged dsDNA, binding of the EREs to the ERs on the n-type SiNW biosensor leads to the accumulation of negative charges on the surface, thereby inducing increase in resistance. The results show that the specificity of the ERE-ERα binding is higher than that of the ERE-ERß binding, what is more, the mutant ERE reduces the binding affinity for both ERα and ERß. By applying various concentrations of wild-type ERE to the bound ERα, a very low concentration of 10 fM wild-type ERE was found to be able to bind to the ERα. The reversible association and dissociation between ERα and wt-ERE was achieved, pointing to a reusable biosensor for protein-DNA binding. Through the study, we have established the SiNW biosensor as a promising method in providing comprehensive study for hormone receptor-response element interactions.

Morpholino-functionalized silicon nanowire biosensor for sequence-specific label-free detection of DNA.

We investigated Morpholino-functionalized silicon nanowires (SiNWs) as a novel gene chip platform for the sequence-specific label-free detection of DNA. Morpholino attachment and subsequent Morpholino-DNA hybridization on silicon surface was characterized by X-ray photoelectron spectroscopy and fluorescence microscopy. The resultant Morpholino-modified surfaces showed high specificity of recognition for DNA. Subsequently, by using the same protocol, the surface of the SiNW biosensor was functionalized with Morpholino, and this was used for label-free Morpholino-DNA hybridization detection. Real-time measurements of the Morpholino-functionalized SiNW biosensor exhibited a decrease in a time-dependent conductance when complementary and mutant DNA samples were added. Furthermore, identification of fully complementary versus mismatched DNA samples was carried out by the Morpholino-functionalized SiNW biosensor. We demonstrated that DNA detection using the Morpholino-functionalized SiNW biosensor could be carried out to the hundreds of femtomolar range. The Morpholino-functionalized SiNWs show a novel biosensor for label-free and direct detection of DNA with good selectivity, and a promising application in gene expression.