Production and Evaluation of Antibodies and Phage Display-Derived Peptide Ligands for Immunomagnetic Separation of Mycobacterium bovis

This study describes the development and optimization of an immunomagnetic separation (IMS) method to isolate Mycobacterium bovis cells from lymph node tissues. Gamma-irradiated whole M. bovis AF2122/97 cells and ethanol-extracted surface antigens of such cells were used to produce M. bovis-speci?c polyclonal and monoclonal antibodies in rabbits and mice. They were also used to generate M. bovis-speci?c peptide ligands by phage display biopanning. The various antibodies and peptide ligands obtained were used to coat MyOne tosyl-activated Dynabeads (Life Technologies), singly or in combination, and evaluated for IMS. Initially, M. bovis capture from Middlebrook 7H9 broth suspensions (concentration range, 10 to 10⁵ CFU/ml) was evaluated by IMS combined with an M. bovis-speci?c touchdown PCR. IMS-PCR results and, subsequently, IMS-culture results indicated that the beads with greatest immunocapture capability for M. bovis in broth were those coated simultaneously with a monoclonal antibody and a biotinylated 12-mer peptide. These dually coated beads exhibited minimal capture (mean of 0.36% recovery) of 12 other Mycobacterium spp. occasionally encountered in veterinary tuberculosis (TB) diagnostic laboratories. When the optimized IMS method was applied to various M. bovis-spiked lymph node matrices, it demonstrated excellent detection sensitivities (50% limits of detection of 3.16 and 57.7 CFU/ml of lymph node tissue homogenate for IMS-PCR and IMS-culture, respectively). The optimized IMS method therefore has the potential to improve isolation of M. bovis from lymph nodes and hence the diagnosis of bovine tuberculosis.

Luminescent Photoinduced Electron Transfer (PET) Molecules for Sensing and Logic Operations

Chemical species can serve as inputs to supramolecular devices so that a luminescence output is created in a conditional manner. Conditionality is built into these devices by employing the classical photochemical process of photoinduced electron transfer (PET) to compete with luminescence emission. The response of these devices in the analogue regime leads to sensors that can operate in nanometric, micrometric, and millimetric spaces. Some of these devices serve in membrane science, cell physiology, and medical diagnostics. The response in the digital regime leads to Boolean logic gates. Some of these find application in improving aspects of medical diagnostics and in identifying small objects in large populations.

Molecular Logic Gates and Luminescent Sensors Based on Photoinduced Electron Transfer

The competition between Photoinduced electron transfer (PET) and other de-excitation pathways such as fluorescence and phosphorescence can be controlled within designed molecular structures. Depending on the particular design, the resulting optical output is thus a function of various inputs such as ion concentration and excitation light dose. Once digitized into binary code, these input-output patterns can be interpreted according to Boolean logic. The single-input logic types of YES and NOT cover simple sensors and the double- (or higher-) input logic types represent other gates such as AND and OR. The logic-based arithmetic processors such as half-adders and half-subtractors are also featured. Naturally, a principal application of the more complex gates is in multi-sensing contexts.

AND Logic Gates Based on Small Molecules with Chemical Inputs and Luminescence Output

AND logic gate behaviour can be recognized in chemical-responsive luminescence phenomena concerning small molecules. Though initial developments concerned separate and distinguishable chemical species as inputs, consideration of other types of input sets allows substantial expansion of the sub-field. Dissection of these molecular devices into modules, where possible, enables analysis of their logic behaviour according to supramolecular photochemical mechanisms.

Molecular Logic Gate Arrays

Chemists are now able to emulate the ideas and instruments of mathematics and computer science with molecules. The integration of molecular logic gates into small arrays has been a growth area during the last few years. The design principles underlying a collection of these cases are examined. Some of these computing molecules are applicable in medical- and biotechnologies. Cases of blood diagnostics, 'lab-on-a-molecule' systems, and molecular computational identification of small objects are included.

CLOSE—A Data-Driven Approach to Speech Separation

This paper studies single-channel speech separation, assuming unknown, arbitrary temporal dynamics for the speech signals to be separated. A data-driven approach is described, which matches each mixed speech segment against a composite training segment to separate the underlying clean speech segments. To advance the separation accuracy, the new approach seeks and separates the longest mixed speech segments with matching composite training segments. Lengthening the mixed speech segments to match reduces the uncertainty of the constituent training segments, and hence the error of separation. For convenience, we call the new approach Composition of Longest Segments, or CLOSE. The CLOSE method includes a data-driven approach to model long-range temporal dynamics of speech signals, and a statistical approach to identify the longest mixed speech segments with matching composite training segments. Experiments are conducted on the Wall Street Journal database, for separating mixtures of two simultaneous large-vocabulary speech utterances spoken by two different speakers. The results are evaluated using various objective and subjective measures, including the challenge of large-vocabulary continuous speech recognition. It is shown that the new separation approach leads to significant improvement in all these measures.