California Institute of Technology

Adrian Ponce

Gregory H Bearman

Dmitri A Kossakovski

Kasthuri J Venkateswaran

James Patrick Kirby

Described herein are methods and apparatus for assays of bacterial spores. In particular, methods and apparatus for lateral flow immunoassay for bacterial spore detection and quantification, live/dead assay for bacterial spores, lifetime-gated measurements of bacterial spores and imaging bacterial spores using an active pixel sensor, and unattended monitoring of bacterial spores in the air are described.

Methods and apparatus for assays of bacterial spores

A method and an apparatus for detecting and quantifying bacterial spores on a surface. In accordance with the method: a matrix including lanthanide ions is provided on the surface containing the bacterial spores; functionalized aromatic molecules are released from the bacterial spores on the surface; a complex of the lanthanide ion and the aromatic molecule is formed on the surface; the complex of the lanthanide ion and the aromatic molecule is excited to generate a characteristic luminescence of the complex on the surface; and the bacterial spores exhibiting the luminescence of the complex on the surface are detected and quantified.

Method and apparatus for detecting and quantifying bacterial spores on a surface

A method and an apparatus for detecting and quantifying bacterial spores on a surface. In accordance with the method: bacterial spores are transferred from a place of origin to a test surface, the test surface comprises lanthanide ions. Aromatic molecules are released from the bacterial spores; a complex of the lanthanide ions and aromatic molecules is formed on the test surface, the complex is excited to generate a characteristic luminescence on the test surface; the luminescence on the test surface is detected and quantified.

A sample of unknown bacterial spores is added to a test strip. The sample of unknown bacterial spores is drawn to a first sample region on the test strip by capillary action. Species-specific antibodies are bound to the sample when the unknown bacterial spores match the species-specific antibodies, otherwise the sample is left unbound. DPA is released from the bacterial spores in the bound sample. The terbium ions are combined with the DPA to form a Tb-DPA complex. The combined terbium ions and DPA are excited to generate a luminescence characteristic of the combined terbium ions and DPA to detect the bacterial spores. A live/dead assay is performed by a release of the DPA for live spores and a release of DPA for all spores. The detection concentrations are compared to determine the fraction of live spores. Lifetime-gated measurements of bacterial spores to eliminate any fluorescence background from organic chromophores comprise labeling the bacterial spore contents with a long-lifetime lumophore and detecting the luminescence after a waiting period. Unattended monitoring of bacterial spores in the air comprises the steps of collecting bacterial spores carried in the air and repeatedly performing the Tb-DPA detection steps above. The invention is also apparatus for performing the various methods disclosed above.

Methods and apparatus and assays of bacterial spores

Optical time domain reflectometry caused by absorption of a volatile or analyte into the fiber optic cladding is used an optical nose. The fiber optics () are covered with a gas permeable film () which is patterned to leave millimeter wide gas permeable notches (-). The notches contain a sensing polymer that responds to different gases by expanding or contracting.

Method and apparatus for distributed sensing of volatiles using a long period fiber grating sensor with modulated plastic coating for environmental monitoring

An apparatus and method for automated monitoring of airborne bacterial spores. The apparatus is provided with an air sampler, a surface for capturing airborne spores, a thermal lysis unit to release DPA from bacterial spores, a source of lanthanide ions, and a spectrometer for excitation and detection of the characteristic fluorescence of the aromatic molecules in bacterial spores complexed with lanthanide ions. In accordance with the method: computer-programmed steps allow for automation of the apparatus for the monitoring of airborne bacterial spores.

Apparatus and method for automated monitoring of airborne bacterial spores

A lanthanide is combined with a medium to be tested for endospores. The dipicolinic acid released from the endospores binds the lanthanides, which have distinctive emission (i.e., luminescence) spectra, and are detected using photoluminescence. The concentration of spores is determined by preparing a calibration curve generated from photoluminescence spectra of lanthanide complex mixed with spores of a known concentration. A lanthanide complex is used as the analysis reagent, and is comprised of lanthanide ions bound to multidentate ligands that increase the dipicolinic acid binding constant through a cooperative binding effect with respect to lanthanide chloride. The resulting combined effect of increasing the binding constant and eliminating coordinated water and multiple equilibria increase the sensitivity of the endospore assay by an estimated three to four orders of magnitude over prior art of endospore detection based on lanthanide luminescence.

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Altadena, CA, United States of America

Adrian Ponce