This work is devoted to formulation and development of a laser spectroscopic techniquefor rapid detection of biohazards, such as Bacillus anthracis spores. Coherent anti-StokesRaman scattering (CARS) is used as an underlying process for active retrieval ofspecies-specific characteristics of an analyte. Vibrational modes of constituent moleculesare Raman-excited by a pair of ultrashort, femtosecond laser pulses, and then probedthrough inelastic scattering of a third, time-delayed laser field.We first employ the already known time-resolved CARS technique. We apply itto the spectroscopy of easy-to-handle methanol-water mixtures, and then continuebuilding our expertise on solutions of dipicolinic acid (DPA) and its salts, which happento be marker molecules for bacterial spores. Various acquisition schemes are evaluated,and the preference is given to multi-channel frequency-resolved detection, when thewhole CARS spectrum is recorded as a function of the probe pulse delay. Wedemonstrate a simple detection algorithm that manages to differentiate DPA solutionfrom common interferents. We investigate experimentally the advantages anddisadvantages of near-resonant probing of the excited molecular coherence, and finallyobserve the indicative backscattered CARS signal from DPA and NaDPA powders. The possibility of selective Raman excitation via pulse shaping of the preparation pulses isalso demonstrated.The analysis of time-resolved CARS experiments on powders and B. subtilisspores, a harmless surrogate for B. anthracis, facilitates the formulation of a newapproach, where we take full advantage of the multi-channel frequency-resolvedacquisition and spectrally discriminate the Raman-resonant CARS signal from thebackground due to other instantaneous four-wave mixing (FWM) processes. Usingnarrowband probing, we decrease the magnitude of the nonresonant FWM, which isfurther suppressed by the timing of the laser pulses. The devised technique, referred to ashybrid CARS, leads to a single-shot detection of as few as 104 bacterial spores, bringingCARS spectroscopy to the forefront of potential candidates for real-time biohazarddetection. It also gives promise to many other applications of CARS, hindered so far bythe presence of the overwhelming nonresonant FWM background, mentioned above.
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