High Throughput Screening Using Enzyme Assay Microarrays

We report a new slide based microarray platform for assaying multiple enzyme activities using fluorogenic substrates. The method enables us to achieve the microfluidic requirements for rapid reaction assembly and compartmentalization. We can thus determine enzymatic activities in individually controlled reaction environments containing cofactors, inhibitors and activators. Fluorogenic substrates in glycerol were arrayed onto glass slides with reaction volumes \u3c 5 nL and feature sizes of \u3c 150 μm. Our method allowed rapid multiple sample deliveries onto the slide (\u3c 3 nL/spot) with no cross contamination between array positions. It enabled us to detect the activation of the fibrinolytic and coagulation proteases namely, thrombin, plasmin, factor Xa, tPa and kallikrein in human plasma. Enzyme-substrate-inhibitor assays using ten caspases were also performed. With over 400 spots/cm2, combinatorial substrate libraries with different proteases can now be rapidly profiled. An assay to detect the dose response of a thrombin inhibitor benzamidine was performed. The inhibitor was arrayed in replicates onto selected positions on the chip. After sequential subnanoliter delivery of the reaction components, the result from the array was analyzed. The expected dose response from benzamidine was seen. A CV of 5.26% was achieved for 232 positions on the array not spiked with the inhibitor. Thus, with potentially several thousand compounds per slide, using rapid sub-nanoliter delivery of components and standard equipment, the true potential of the method is in the field of high throughput screening

PCA Encrypted Short Acoustic Data Inculcated in Digital Color Images

We propose develop a generalized algorithm for hiding audio signal using image steganography. The authors suggest transmitting short audio messages camouflaged in digital images using Principal Component Analysis (PCA) as an encryption technique. The quantum of principal components required to represent the audio signal by removing the redundancies is a measure of the magnitude of the Eigen values. The aforementioned technique follows a dual task of encryption and in turn also compresses the audio data, sufficient enough to be buried in the image. A 57Kb audio signal is decipher from the Stego image with a high PSNR of 47.49 and a correspondingly low mse of 3.3266 × 1

Signals in the Soil: An Introduction to Wireless Underground Communications

In this chapter, wireless underground (UG) communications are introduced. A detailed overview of WUC is given. A comprehensive review of research challenges in WUC is presented. The evolution of underground wireless is also discussed. Moreover, different component of UG communications is wireless. The WUC system architecture is explained with a detailed discussion of the anatomy of an underground mote. The examples of UG wireless communication systems are explored. Furthermore, the differences of UG wireless and over-the-air wireless are debated. Different types of wireless underground channel (e.g., In-Soil, Soil-to-Air, and Air-to-Soil) are reported as well

'Inject-Mix-React-Separate-and-Quantitate' (IMReSQ) method for screening enzyme inhibitors

Many regulatory enzymes are considered attractive therapeutic targets, and their inhibitors are potential drug candidates. Screening combinatorial libraries for enzyme inhibitors is pivotal to identifying hit compounds for the development of drugs targeting regulatory enzymes. Here, we introduce the first inhibitor screening method that consumes only nanoliters of the reactant solutions and is applicable to regulatory enzymes. The method is termed inject-mix-react-separate-and-quantitate (IMReSQ) and includes five steps. First, nanoliter volumes of substrate, candidate inhibitor, and enzyme solutions are injected by pressure into a capillary as separate plugs. Second, the plugs are mixed inside this capillary microreactor by transverse diffusion of laminar flow profiles. Third, the reaction mixture is incubated to form the enzymatic product. Fourth, the product is separated from the substrate inside the capillary by electrophoresis. Fifth, the amounts of the product and substrate are quantitated. In this proof-of-principle work, we applied IMReSQ to study inhibition of recently cloned protein farnesyltransferase from parasite Entamoeba histolytica. This enzyme is a potential therapeutic target for antiparasitic drugs. We identified three previously unknown inhibitors of this enzyme and proved that IMReSQ could be used for quantitatively ranking the potencies of inhibitors

Probing Biology with Small Molecule Microarrays (SMM)

In the continuous drive to increase screening throughput and reduce sample requirement, microarray-based technologies have risen to the occasion. In the past 7 years, a number of new methodologies have been developed for preparing small molecule microarrays from combinatorial and natural product libraries with the goal of identifying new interactions or enzymatic activities. Recent advances and applications of small molecule microarrays are reviewed