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Polyethylene glycol (PEG) hydrogel is permeable to biomolecules, but its permeability depends on the molecular weight of monomers and the concentration of monomer solutions. In this study, we show that PEG hydrogel made from 20% to 30% of PEG700 monomer is permeable to amino acids yet impermeable to oligopeptides. Because of its unique permeability, the gel can be used to detect protease by separating amino acids from oligopeptides when proteases cleave the oligopeptides and release amino acids. Based on this principle, an UV crosslinked gel array is fabricated on a chip for simultaneous detection of protease in up to 40 samples with only 1 µl of volume required for each sample. As a proof of concept, the on-chip protease assays are used to detect trypsin in buffer and serum. The detection limits are 1.2 nM in buffer and 17.7 nM in serum, which are comparable to conventional protease assays. Moreover, because only 1 µl of liquid is required, as little as 1.2 fmol of trypsin can be detected by using the on-chip assay. The protease assay also shows good specificity for trypsin and chymotrypsin. The gel array chip could be a useful miniaturized platform for high-throughput detection of different proteases and screening of their inhibitors. [ABSTRACT FROM AUTHOR]

Abstract: We report a functional protease assay in which liquid crystals (LCs) are used as signal reporters to transduce the test results into optical signals. In this assay, an oligopeptide substrate (CLSELDDRADALQAGASQFESSAAKLKRKYWWKNLK) is used as a probe. This oligopeptide can be cleaved by α-chymotrypsin at multiple locations and become smaller fragments after the cleavage. When the original oligopeptide is immobilized on a solid surface, its long flexible oligopeptide chain is able to influence the orientation of a thin layer of LC supported on the surface, as is evident as a bright spot on the surface. In contrast, when the shorter oligopeptide fragments are immobilized on the same surface, their shorter, less flexible chains cannot disrupt the orientation of LC, and a dark spot is observed. On the basis of the dark or bright signal from LC, α-chymotrypsin in buffer solution or complex media such as chicken broth can be detected by using the naked eye. However, when the incubation time is 3h, the limit of detection (LOD) for α-chymotrypsin in buffer solution is 50ng/mL, whereas that in chicken broth is only 500ng/mL. Unlike traditional antibody-based assays which show little difference between active and inactive α-chymotrypsin, only active protease can be detected in this assay. This study shows the potential utility of LCs for detecting functional proteases with good specificity and sensitivity. [Copyright &y& Elsevier]

Abstract: Immunobiosensors are emerging as powerful tools for diagnostic applications. In this paper, we exploit optical properties of liquid crystal (LC) and the specific epitope–antibody interaction to develop an optical immunobiosensor which can be used to detect monoclonal anti-FLAG M2, a model antibody. The sensitive layer of this immunobiosensor is a monolayer of immobilized oligopeptide CDYKDDDDK (FLAG) at an LC/aqueous interface. The linear oligopeptide can function as an epitope which can be recognized by anti-FLAG M2. When the LC is modified with 50μM of FLAG and then exposed to an aqueous solution containing anti-FLAG, anti-FLAG binds to the immobilized FLAG epitope and triggers orientational transition of LC. Because of the optical birefringence of the LC, the orientational transition is accompanied by changes in the optical images of LC from dark to bright under crossed polarizers. The detection limit for the system is 27ng/mL of anti-FLAG, and the response time is <1h. The binding of anti-FLAG to its motif FLAG oligopeptide is also very specific. When a variant epitope CDAKDDDDK is used, no apparent optical response can be observed after the addition of anti-FLAG. [Copyright &y& Elsevier]

Abstract: Semiconducting nanowires have the potential to function as highly sensitive and selective sensors for label-free detection of biological and chemical species. In this paper, we describe the preparation and applications of oligopeptide-modified single crystal silicon nanowire (SiNW) arrays as multichannel metal ion sensors. Because these SiNW arrays were fabricated by using a “top-down” process, a very high density of individually addressable SiNW (180 SiNWs grouped into 36 clusters) can be manufactured on a single chip. Furthermore, after two different SiNW clusters are modified with Pb2+-selective and Cu2+-selective oligopeptide, respectively, concentrations of Pb2+ and Cu2+ in aqueous solutions can be detected simultaneously and selectively in two different channels. By using the SiNW arrays, we successfully achieved a detection limit of 1nM for Pb2+ and 10nM for Cu2+, respectively. These results demonstrate the potential utility of oligopeptide-modified SiNW arrays for highly sensitive and selective chemical and biological sensing applications. [Copyright &y& Elsevier]

Abstract: We report a liquid crystal (LC)-based sensor for real-time and label-free identification of phospholipase-like toxins. Beta-bungarotoxin exhibits Ca2+-dependent phospholipase A2 activity whereas alpha-bungarotoxin and myotoxin II do not exhibit any phospholipase activity. The sensor can selectively identify beta-bungarotoxin, when it hydrolyzes a phospholipid monolayer self-assembled at aqueous–LC interface, through orientational responses of LCs. As a result, optical signals that reflect the spatial and temporal distribution of phospholipids during the hydrolysis can therefore be generated in a real-time manner. The sensor is very sensitive and requires less than 5pg of beta-bungarotoxin for the detection. When phospholipase A2 inhibitors are introduced together with beta-bungarotoxin, no orientational response of LCs can be observed. In addition, the regeneration of the sensor can be done without affecting the sensing performance. This work demonstrates a simple and cost-effective LC-based sensor for identifying phospholipase-like toxins and for screening compound libraries to find potential toxin inhibitors. [Copyright &y& Elsevier]

Abstract: By using phage display library, we identified two highly specific oligopeptide sequences RKRIRRMMPRPS and RNRHTHLRTRPR for binding neonicotinoids such as thiacloprid and imidacloprid. The former shows high affinity for thiacloprid whereas the latter shows high affinity for imidacloprid. Surprisingly, cross binding is minimal despite the similarity of the two molecules. To develop a neonicotinoid biosensor, these two oligopeptides are synthesized and immobilized on the surface of a surface plasmon resonance (SPR) chip with a bare-gold surface. This oligopeptide functionalized SPR biosensor can rapidly detect thiacloprid and imidacloprid in buffer solutions in a real-time manner. The limit of detection (LOD) for thiacloprid and imidacloprid is 1.2μM and 0.9μM, respectively. [Copyright &y& Elsevier]

Abstract: This paper reports the electrical detection of calcium ions by using silicon nanowires (SiNWs) as channels in a chemically gated field-effect-transistor (FET) configuration. To obtain a selective and sensitive layer for calcium sensing, the SiNWs are modified with a biologically relevant amino acid phosphotyrosine (p-Tyr), which is able to complex calcium ions with high affinity. It is found that when the p-Tyr modified SiNWs are exposed to aqueous solutions containing calcium ions, their conductances increase with the increasing of calcium concentration up to 10μM. In contrast, when the SiNWs are exposed to sodium or potassium, or when they are modified with tyrosine (Tyr), no significant increase in the conductance is observed. This finding suggests that the calcium ions complexed with the phosphate group of p-Tyr can act as a positive gate voltage on the FET device comprising of n-type SiNWs, and leads to an increase in their conductances. The FET device is also sensitive to magnesium ions. However, the response is 10 times lower than that of calcium at the same concentration. The study reported here may pave the way for designing an intracellular calcium sensor which permits the monitoring of calcium concentration in real time. [Copyright &y& Elsevier]