Joshua L. Santarpia

Fluorescence of bioaerosols: mathematical model including primary fluorescing and absorbing molecules in bacteria

This paper describes a mathematical model of fluorescent biological particles composed of bacteria, viruses, or proteins. The fluorescent and/or light absorbing molecules included in the model are amino acids (tryptophan, etc.); nucleic acids (DNA, RNA, etc.); coenzymes (nicotinamide adenine dinucleotides, flavins, and vitamins B₆ and K and variants of these); and dipicolinates. The concentrations, absorptivities, and fluorescence quantum yields are estimated from the literature, often with large uncertainties. The bioparticles in the model are spherical and homogeneous. Calculated fluorescence cross sections for particles excited at 266, 280, and 355 nm are compared with measured values from the literature for several bacteria, bacterial spores and albumins. The calculated 266- and 280-nm excited fluorescence is within a factor of 3.2 of the measurements for the vegetative cells and proteins, but overestimates the fluorescence of spores by a factor of 10 or more. This is the first reported modeling of the fluorescence of bioaerosols in which the primary fluorophores and absorbing molecules are included.

Changes in fluorescence spectra of bioaerosols exposed to ozone in a laboratory reaction chamber to simulate atmospheric aging

A laboratory system for exposing aerosol particles to ozone and rapidly measuring the subsequent changes in their single-particle fluorescence is reported. The system consists of a rotating drum chamber and a single-particle fluorescence spectrometer (SPFS) utilizing excitation at 263 nm. Measurements made with this system show preliminary results on the ultra-violet laser-induced-fluorescence (UV-LIF) spectra of single aerosolized particles of Yersinia rohdei, and of MS2 (bacteriophage) exposed to ozone. When bioparticles are exposed in the chamber the fluorescence emission peak around 330 nm: i) decreases in intensity relative to that of the 400-550 nm band; and ii) shifts slightly toward shorter-wavelengths (consistent with further drying of the particles). In these experiments, changes were observed at exposures below the US Environmental Protection Agency (EPA) limits for ozone.

Trapping of individual airborne absorbing particles using a counterflow nozzle and photophoretic trap for continuous sampling and analysis

We describe an integrated opto-aerodynamic system and demonstrate that it enables us to trap absorbing airborne micron-size particles from air, hold them and then release them, and to repeat this sequence many times as would be appropriate for continuous sampling of particles from air. The key parts of the system are a conical photophoretic optical trap and a counter-flow coaxial-double-nozzle that concentrates and then slows particles for trapping. This technology should be useful for on-line applications that require monitoring (by single particle analyses) of a series of successively arriving particles (e.g., from the atmosphere or pharmaceutical or other production facilities) where the total sampling time may last from minutes to days, but where each particle must be held for a short time for measurements (e.g., Raman scattering).

Improved Method for the Evaluation of Real-Time Biological Aerosol Detection Technologies

There is a growing need to evaluate bioaerosol sensors under relevant operational conditions. New methods are needed that can mimic the temporal fluctuations of ambient aerosol backgrounds and present biological aerosol challenges in a way that simulates a plausible biological agent attack. The Dynamic Concentration Aerosol Generator was developed to address this need. The authors developed a series of aerosol challenges consisting of Bacillus thuringiensis kurstaki (Btk) spores in the presence of background aerosols using a newly developed ramp testing method. Using ramping style tests, 5-min Btk releases were overlaid on top of a background aerosol that fluctuated at varying rates. Background aerosol compositions for different tests were designed to simulate the types of aerosol in the ambient environment. Background aerosol concentration was varied between 7.0 × 103 and 1.5 × 104 particles per liter of air (ppL). Aerosol number concentrations of Btk for the challenges were approximately 2.5 × 103 ppL and the culturable fraction of the collected Btk aerosol was estimated to be 1.25 × 103 colony forming-units (cfu)/L-air. Results of these experiments demonstrate a novel technique for dynamic aerosol generation that can be used to test biological aerosol sensors under controlled conditions designed to reproduce observed fluctuations in the ambient aerosol.

Spectrally-resolved fluorescence cross sections of aerosolized biological live agents and simulants using five excitation wavelengths in a BSL-3 laboratory

A system for measuring spectrally-resolved fluorescence cross sections of single bioaerosol particles has been developed and employed in a biological safety level 3 (BSL-3) facility at Edgewood Chemical and Biological Center (ECBC). It is used to aerosolize the slurry or solution of live agents and surrogates into dried micron-size particles, and to measure the fluorescence spectra and sizes of the particles one at a time. Spectrally-resolved fluorescence cross sections were measured for (1) bacterial spores: Bacillus anthracis Ames (BaA), B. atrophaeus var. globigii (BG) (formerly known as Bacillus globigii), B. thuringiensis israelensis (Bti), B. thuringiensis kurstaki (Btk), B. anthracis Sterne (BaS); (2) vegetative bacteria: Escherichia coli (E. coli), Pantoea agglomerans (Eh) (formerly known as Erwinia herbicola), Yersinia rohdei (Yr), Yersinia pestis CO92 (Yp); and (3) virus preparations: Venezuelan equine encephalitis TC83 (VEE) and the bacteriophage MS2. The excitation wavelengths were 266 nm, 273 nm, 280 nm, 365 nm and 405 nm.

Relationship Between Biologically Fluorescent Aerosol and Local Meteorological Conditions

Time-resolved characterization of biological aerosol is important both for understanding environmental processes that affect biological aerosols and for determining realistic test conditions for the evaluation of bioaerosol detection systems. Very little work has been done to develop an understanding of the temporal fluctuations in bioaerosol concentration. During an experiment from 1–10 November 2008 ambient biological aerosol and meteorological data were collected. A FLIR/ICx/S3I Instantaneous Bioaerosol Analysis and Collection sensor was used to count both the biological and nonbiological aerosol in two size bins. The data indicate that the ambient relative humidity affects the optically observable concentration of biological aerosol with higher relative humidity generally associated with higher biological aerosol concentrations. The short timescale over which these correlations exist implies an aerosol process, rather than a change in aerosol source.

Measurement of back-scattering patterns from single laser trapped aerosol particles in air

We demonstrate a method for measuring elastic back-scattering patterns from single laser trapped micron-sized particles, spanning the scattering angle range of θ=167.7°-180° and φ=0°-360° in spherical coordinates. We calibrated the apparatus by capturing light-scattering patterns of 10 μm diameter borosilicate glass microspheres and comparing their scattered intensities with Lorenz-Mie theory. Back-scattering patterns are also presented from a single trapped Johnson grass spore, two attached Johnson grass spores, and a cluster of Johnson grass spores. The method has potential use in characterizing airborne aerosol particles, and may be used to provide back-scattering data for lidar applications.

Position-resolved Raman spectra from a laser-trapped single airborne chemical droplet

It could be very useful to detect and monitor the molecules and molecular reactions located at different positions within a microsized particle as they respond to various micro-local environments. In this Letter, a particular optical trap using two focusing counterpropagating hollow beams was able to stably trap both absorbing and nonabsorbing particles in air for lengthy observation. A technique that can measure the Raman spectra from different submicrometer positions of a laser-trapped single airborne particle was developed. Spontaneous and stimulated Raman scattering spectra originating from different positions of a diethyl phthalate droplet were recorded, and the strong Raman scattering signals are the result of cavity-enhanced effects and the localized strong light illumination.

Raman scattering and red fluorescence in the photochemical transformation of dry tryptophan particles.

Tryptophan is a fluorescent amino acid common in proteins. Its absorption is largest for wavelengths λ ≲ 290 nm and its fluorescence emissions peak around 300-350 nm, depending upon the local environment. Here we report the observation of red fluorescence near 600 nm emerging from 488-nm continuous-wave (CW) laser photoexcitation of dry tryptophan (Trp) particles. With an excitation intensity below 0.5 kW/cm2, dry Trp particles yield distinctive Raman scattering peaks in the presence of relatively weak and spectrally broad emissions with λ ∼500-700 nm, allowing estimation of particle temperature at low excitation intensities. When the photoexcitation intensity is increased to 1 kW/cm2 or more for a few minutes, fluorescence intensity dramatically increases by more than two orders of magnitude. The fluorescence continues to increase in intensity and gradually shift to the red when photoexcitation intensity and the duration of exposure are increased. The resulting products absorb at visible wavelengths and generate red fluorescence with λ ∼ 650-800 nm with 633-nm CW laser excitation. We attribute the emergence of orange and red fluorescence in the Trp products to a photochemical transformation that is instigated by weak optical transitions to triplet states in Trp with 488-nm excitation and which may be expedited by a photothermal effect.

Opto-aerodynamic focusing of aerosol particles

ABSTRACT We describe a new method for focusing and concentrating a stream of moving micron-sized aerosol particles in air. The focusing and concentrating process is carried out by the combined drag force and optical force that is generated by a double-layer co-axial nozzle and a focused doughnut-shaped hollow laser beam, respectively. This method should supply a new tool for aerosol science and related research. Copyright