Sensors that are able to provide reagent-free, continuous monitoring for potential bio-aerosol hazards are required in many environments. In general, increasing the number of optical and spectroscopic properties of individual airborne particles that can be measured increases the level of detection confidence and reduces the risk of false-positive detection. This paper describes the development of relatively low-cost multi-parameter prototype sensors that can monitor and classify the ambient aerosol by simultaneously recording both a 2x2 fluorescence excitation-emission matrix and multi-angle spatial elastic scattering data from individual airborne particles. The former can indicate the possible presence of specific biological fluorophores in the particle whilst the latter provides an assessment of particle size and shape.
The following paper describes the core technology of a real time optical biological agent detection system which has been developed from Biral's existing and proven ASAS<sup>TM</sup> family of biological agent triggers and sensors. The sensor for the system has additional capabilities based on the introduction of a fluorescence sensor providing greater discrimination than previously available. The new sensor will be incorporated in the Integrated Sensor Management System (ISMS) which utilises complex real-time data tracking algorithms that simultaneously monitor the different aerosol characteristics. The system is capable of tracking and adjusting the sensor alert levels to take into account the constantly changing aerosol environment and thus significantly reduces the risk of false alarms. The use of two measurement systems, ASAS<sup>TM</sup> and fluorescence in one sensor is a unique combination and a major advancement in the field of airborne biological agent detection and warning.
Laser diodes and light-emitting diodes capable of continuous sub-300 nm radiation emission will ultimately represent optimal excitation sources for compact and fieldable bio-aerosol monitors. However, until such devices are routinely available and whilst solid-state UV lasers remain relatively expensive, other low-cost sources of UV can offer advantages. This paper describes one such prototype that employs compact xenon discharge UV sources to excite intrinsic fluorescence from individual particles within an ambient aerosol sample. The prototype monitor samples ambient air via a laminar sheathed-flow arrangement such that particles within the sample flow column are rendered in single file as they intersect the beam from a continuous-wave 660nm diode laser. Each individual particle produces a scattered light signal from which an estimate of particle size (down to ~1 um) may be derived. This same signal also initiates the sequential firing (~10 us apart) of two xenon sources which irradiate the particle with UV pulses centred upon ~280 nm and ~370 nm wavelength, optimal for excitation of bio-fluorophores tryptophan and NADH respectively. For each excitation wavelength, fluorescence is detected across two bands embracing the peak emissions of the same bio-fluorophores. Thus, for each particle, a 2-dimensional fluorescence excitation-emission matrix is recorded together with an estimate of particle size. Current measurement rates are up to ~125 particles/s (limited by the xenon recharge time), corresponding to all particles for concentrations up to ~2 x 10<sup>4</sup> particles/l. Developments to increase this to ~500 particles/s are in hand. Analysis of results from aerosols of E.coli, BG spores, and a variety of non-biological materials are given.
Developments in real time optical biological agent detection and sensing are presented which describe start of the art advances in the detection and warning of these pathogens. The following paper describes the basic operating principles of the current BIRAL ASAS (Aerosol Size and Shape) system which measures the optically determined particle properties, on a particle by particle basis, and uses the information to describe the size and shape characteristics of the aerosol. Furthermore, recent development of the existing technology to also encompass fluorescence detection is described, which significantly increases the detection ability of the ASAS aerosol suite. This operational improvement is a major advancement in the field of airborne biological agent detection and allows
for near generic detection and warning. Applications of this device include all aspects of bio-aerosol monitoring, including the use as a biological agent detector and generic identifier, use as a general bio-agent monitor and also for use as a hazardous environment monitor. Such a device would be particularly useful in the fields of Armed Forces protection and National Defence either as a point detector or as a "plug and play" biosensor detector in a network.
We describe the construction of a bio-aerosol monitor designed to capture and record intrinsic fluorescence spectra from individual aerosol particles carried in a sample airflow and to simultaneously capture data relating to the spatial distribution of elastically scattered light from each particle. The spectral fluorescence data recorded by this PFAS (<i>Particle Fluorescence and Shape</i>) monitor contains information relating to the particle material content and specifically to possible biological fluorophores. The spatial scattering data from PFAS yields information relating to particle size and shape. The combination of these data can provide a means of aiding the discrimination of bio-aerosols from background or interferent aerosol particles which may have similar fluorescence properties but exhibit shapes and/or sizes not normally associated with biological particles. The radiation used both to excite particle fluorescence and generate the necessary spatially scattered light flux is provided by a novel compact UV fiber laser operating at 266nm wavelength. Particles drawn from the ambient environment traverse the laser beam in single file. Intrinsic particle fluorescence in the range 300-570nm is collected via an ellipsoidal concentrator into a concave grating spectrometer, the spectral data being recorded using a 16-anode linear array photomultiplier detector. Simultaneously, the spatial radiation pattern scattered by the particle over 5°-30° scattering angle and 360° of azimuth is recorded using a custom designed 31-pixel radial hybrid photodiode array. Data from up to ~5,000 particles per second may be acquired for analysis, usually performed by artificial neural network classification.
We describe a low-cost prototype bio-aerosol fluorescence sensor designed for unattended deployment in medium to large area networks. The sensor uses two compact xenon flash units to excite fluorescence in an aerosol sample volume drawn continuously from the ambient environment. In operation, the xenons are pulsed alternately at 300ms intervals whilst absorption filters restrict their radiation output to UV bands ~260-290nm and ~340-380nm respectively, optimal for exciting the biological fluorophores tryptophan and NADH. Fluorescence from all particles instantaneously present within a sensing volume is measured using two miniature photomultiplier detectors optically filtered to detect radiation in the bands ~320-600nm and ~410-600nm. The second of these bands covers the principal emission from NADH, whilst the difference between the first and second detector channels yields fluorescence in the 320-410nm band, covering much of the tryptophan emission. Whilst each sensor is clearly limited in specificity, the low sensor cost (<$5k) offers potential for the deployment in large networks that would be prohibitively expensive using particle fluorescence sensors based on currently available UV lasers. Preliminary details are also given of a variant of the sensor, currently under development, in which xenon illumination is used to acquire single particle fluorescence data at rates of up to 200 particles per second.
Several workers have reported the development of systems which allow the measurement of intrinsic fluorescence from particles irradiated with ultra-violet radiation. The fluorescence data are frequently recorded in conjunction with other parameters such as particle size, measured either as a function of optical scatter or as an aerodynamic size. The motivation for this work has been principally the detection of bioaerosols within an ambient environment. Previous work by the authors has shown that an analysis of the scattering profile of a particle, i.e.: the spatial distribution of light scattered by the particle carried in a sample air-stream, can provide an effective means of particle characterization and classification in terms of both size and shape parameters. Current work is aimed at the simultaneous recording of both spatial scattering and fluorescence data from individual particles with a view to substantially enhanced discrimination of biological aerosols. A prototype instrument has recently been completed which employs a cw 266 nm laser source to produce both elastic (spatial scattering) and inelastic (fluorescence) signals from individual airborne particles. The instrument incorporates a custom designed high-gain multi- pixel hybrid photodiode (HPD) to record the spatial scattering data and a single photomultiplier to record total fluorescence from the illuminated particle. Recorded data are processed to allow the classification of airborne particles on the basis of size, shape, and fluorescence for both biological and non- biological aerosols.