A Compact Aerosol Sensor and Spectroscopic Sorting with UV LEDs

We employ a linear array of UV LEDs whose individual elements may be fired in rapid sequence so as to continuously illuminate a particle during its time-of-flight.  A linear array of separate UV LEDs offers several advantages over a single element device; an increase in the total energy delivered to the particle, which is achieved both by extending the excitation time as well as enabling the use of higher injection currents, and a reduction in the background signal.

The array consists of 32 individually addressable elements, each with an optical aperture 200 um wide and 50 um high. The total array height is 3.2 mm and with 1:1 imaging optics the particle is tracked for this distance during its trajectory. In results presented later in this paper, both 340 nm and 275 nm LED arrays are used. The maximum CW output power before thermal rollover from a single 275 nm (340 nm) LED element is approximately 600 uW (1 mW). This occurs at a current injection level of 40 mA (100 mA) where the power is measured directly off the sapphire backside of the die with no integrating sphere. This represent state-of-the-art at both wavelengths, and the inclusion of a 275 nm LED array is the most recent addition to the system presented here.

1.   Compact spectroscopic optics: 


Schematic of the optical apparatus for acquiring single-particle spectra employing a UV LED array. (a) Side-view of the system illustrating the particle jetstream and placement of the aerodynamic deflector and (b) top-view showing the compact spectroscopic optics and simultaneous scatter channels.

A pair of simple lenses focuses the array to the particle jetstream and a bandpass excitation filter is used to remove the well-known long-wavelength tail associated with LEDs.  Scatter from the red LD focused onto the jetstream prior to the LED array, serves as the trigger to synchronize the subsequent operations. Scatter at the UV wavelength is collected at a right angle to the incident light and is detected by a simple photomultiplier tube (PMT) equipped with an appropriate bandpass emission filter. This serves to normalize the fluorescence signal. Simultaneously, the fluorescence spectrum is collected using a single lens, UV-grade transmission grating and a 32-anode PMT. Downstream from the UV LED array and fluorescence collection, we employ a solenoid-based aerodynamic deflector as a compact method to separate particles10. When energized by a current pulse an electromagnetic plunger opens the valve, causing a burst of nitrogen gas which lasts for approximately 250 usec. 

2.   Electronic Control

To achieve the required timing and control sequence, 3 electronics boards were custom built by Vtech Engineering Corporation. The central PhotoniQ-3G iQSP480 board is a commercial product whose primary function is to acquire and process the real-time fluorescence data from the 32-anode PMT and trigger the particle deflector based on a spectral algorithm applied to the fluorescence data.
 
Fluorescence collected by the 32-anode PMT is evaluated by a spectral algorithm running on the DSP (digital signal processor) in the iQSP480. Owing to the finite time that the particle is in the jetstream, signal collection and evaluation of the algorithm must be done in real time.Currently a set 8 spectral flags can be defined, each corresponding to a class of particle for instance, and for each particle the software records which, if any, of the flags matches the recorded spectrum. A trigger is generated to the deflector when any of a predefined subset of the spectral flags is recorded.

3.   Demonstration of Detection Capability
 

A piezoelectric generator (MicroDrop GmbH) is used to produce nominally 70 um diameter water droplets doped with varying amounts of tryptophan or NADH intended to simulate the fluorescence from a bacterial spore. Real time and average flourescence spectra aquired from single droplets and illuminated with a 275 nm LED array show the detection sensitivity of the system.
 


 

Typical data acquired using a 275 nm LED array and tryptophan-doped water droplets; (a) raw spectral data of 10 sequential droplets, and (b) average over 100 spectra acquired from a series of fluorophore concentrations.

4.   Demonstration of Sorting Capability

 To demonstrate the system’s capability to spectrally identify single particles among non-identical fluorescing particles and to physically sort a subset of particles from the background, dye-filled polystyrene latex spheres (PSL) are used as simulants of bio-fluorescence.  PSL spheres, as opposed to amino-acid mixtures such as tryptophan and NADH, are useful characterization tools for particle separation as they are present in discrete quantities (ie. 0, 1 or 2 PSL per drop) and can be counted on the substrate to verify experimental results. Two types of PSL are used, 1 um diameter blue fluorescing (Molecular Probes, F8815) and 2 um green fluorescing (Duke Scientific, XPR-801). Figure (a) shows fluorescence spectra from the mixture and indicates the particles which have been flagged as green.  Shown in (b) are the average fluorescence spectra of flagged events during a test of 5000 particles.

A solution of either blue or green doped PSL was separately passed through the system and a simple one-spectral-band algorithm was used to evaluate whether a particle was present or not. A flag to the deflector was generated upon the condition that the average charge collected was greater than a threshold level. In addition, a two-band algorithm where the same wavelength bands were used but where a flag was generated only upon the boolean condition (green = true) AND (blue = false) was used in order to eliminate the possibility of highly fluorescent blue particles, or clusters, from tripping the green flag.
The results of two band algorithm is shown by two epifluorescence images of the deflected and undeflected aggregates of spheres in the case where green PSL were selected for.

Among 5000 doped water droplets, 185 were flagged as having green fluorescence. Counting deflected particles yields nominally 180 green and 8 mistakenly removed blue ones. deflected blue particles indicate that the algorithm targeted to green particles yielded a flag for a blue one, or that clusters of multiple PSL were present in a single water drop. However, undeflected particles are exclusively blue, hence no ‘suspicious’ (green) particles were falsely categorized as ‘safe’ (blue), indicating that upon receipt of a trigger the deflector is extremely reliable in removing suspect particles from the jetstream.

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