Rapid and label-free classification of pathogens based on light scattering,reduced power spectral features and support vector machine

The rapid identification of pathogens is crucial in controlling the food quality and safety. The proposed system for the rapid and label-free identification of pathogens is based on the principle of laser scattering from the bacterial microbes. The clinical prototype consists of three parts: the laser beam, photodetectors, and the data acquisition system. The bacterial testing sample was mixed with 10 mL distilled water and placed inside the machine chamber. When the bacterial microbes pass by the laser beam, the scattering of light occurs due to variation in size, shape, and morphology. Due to this reason, different types of pathogens show their unique light scattering patterns. The photo-detectors were arranged at the surroundings of the sample at different angles to collect the scattered light. The photodetectors convert the scattered light intensity into a voltage waveform. The waveform features were acquired by using the power spectral characteristics, and the dimensionality of extracted features was reduced by applying minimal-redundancy-maximal-relevance criterion (mRMR). A support vector machine (SVM) classifier was developed by training the selected power spectral features for the classification of three different bacterial microbes. The resulting average identification accuracies of E. faecalis, E. coli and S. aureus were 99%, 87%, and 94%, respectively. The overall experimental results yield a higher accuracy of 93.6%, indicating that the proposed device has the potential for label-free identification of pathogens with simplicity, rapidity, and cost-effectiveness.     

On-chip classification of micro-particles using laser light scattering and machine learning

The rapid detection of microparticles exhibits a broad range of applications in the field of science and technology. The proposed method differentiates and identifies the 2 μm and 5 μm sized particles using a laser light scattering. The detection method is based on measuring forward light scattering from the particles and then classifying the acquired data using support vector machines. The device is composed of a microfluidic chip linked with photosensors and a laser device using optical fiber....     

An optical biosensor for rapid and label-free detection of cells

We report a broadly applicable optical method for rapid and label-free detection of as few as 45 cells. In this method, bacterial cells are detected by measuring the amount of laser light transmitted through a small glass well functionalized with antibodies which specifically recognize and capture the cells. The described approach is simple, rapid, economical, and promising for portable and high-throughput detection of a wide variety of pathogenic and infectious cells.     

Feasibility study on identification of green, black and Oolong teas using near-infrared reflectance spectroscopy based on support vector machine (SVM)

Near-infrared (NIR) spectroscopy has been successfully utilized for the rapid identification of green, black and Oolong teas. The spectral features of each category are reasonably differentiated in the NIR region, and the spectral differences provided enough qualitative spectral information for identification. Support vector machine as a pattern recognition was applied to attain the differentiation of the three tea categories in this study. The top five latent variables are extracted by principal component analysis as the input of SVM classifiers. The identification results of the three tea categories were achieved by the RBF SVM classifiers and the polynomial SVM classifiers in different parameters. The best identification accuracies were up to 90%, 100% and 93.33%, respectively, when training, while, 90%, 100% and 95% when test. It was obtained using the RBF SVM classifier with sigma=0.5. The overall results ensure that NIR spectroscopy combined with SVM discrimination method can be efficiently utilized for rapid and simple identification of the different tea categories.     

Rapid detection of Staphylococcus aureus by a combination of monoclonal antibody-coated latex and capillary electrophoresis

The rapid detection of pathogenic bacteria is extremely important in biotechnology and clinical diagnosis. CE has been utilized in the field of bacterial analysis for many years, but to some extent, simultaneous separation and identification of certain microbes from complex samples by CE coupled with UV detector is still a challenge. In this paper, we propose a new strategy for rapid separation and identification of Staphylococcus aureus (S. aureus) in bacterial mixtures by means of specific mAb-coated latex coupled with CZE. An appropriate set of conditions that selectively isolated S. aureus from the microorganisms Escherichia coli, Pseudomonas aeruginosa, and Klebsiella pneumoniae were established. S. aureus could be differentiated from the others by unique peaks in the electropherograms. The validity was also confirmed by LIF with antibodies specific to both the latex and the microbial cells. The LOD is as low as 9.0 x 10(5) colony forming unit/mL. We have also utilized this technology to identify S. aureus in a stool sample coming from a healthy volunteer spiked successfully with S. aureus. This CZE-UV technique can be applied to rapid diagnosis of enteritis caused by S. aureus or other bacterial control-related fields needing rapid identification of target pathogens from microbial mixtures. In theory, this method is suitable for the detection of any bacterium as long as corresponding bacterium-specific antibody-coated latex is available.     

Spontaneous resonance hyper-Raman scattering of intense laser fields

The features of multicomponent spectral structure of the spontaneous hyper-Raman (SHR) scattered light from a four-level system irradiated simultaneously by three near-resonant laser fields are investigated.     

Measuring masses of single bacterial whole cells with a quadrupole ion trap

A novel method has been developed to precisely measure the masses of single bacterial whole cells using a quadrupole ion trap as an electrodynamic balance. The bacterial cells were introduced into the ion trap by matrix-assisted laser desorption/ionization, confined in space by audio frequency ac fields, and detected by elastic light scattering. Mass measurement accuracy approaching 0.1% was achieved for Escherichia coli K-12 with a mass distribution of +/-3% from 60 repetitive measurements of the particles and their clusters. This is the first high-precision mass measurement reported for any intact microorganisms with masses greater than 1 x 1010 Da. The method opens new avenues for high-precision mass measurement of single microbial particles and offers an alternative approach for rapid identification of microorganisms by mass spectrometry.     

Scattered light interference from a single metal nanoparticle and its mirror image

The spatial distribution of surface plasmon scattering from a single nanoparticle changes dramatically near a metal surface as a result of interference from the direct scattered light and indirect scattered light from the mirror reflection. The unique interference patterns have been reproduced by simulations based on Huygens-Fresnel wave propagation theory. The large spectral width of the surface plasmon scattering enables a vertical distance measurement with 10 nm resolution through this nonintrusive far field interferometry.     

Raman, fluorescence, and time-resolved light scattering as optical diagnostic techniques to separate diseased and normal biomedical media.

Studies of Raman scattering, fluorescence and time-resolved light scattering were conducted on cancer and normal biomedical media. Fourier transform Raman spectroscopic measurements were performed on human normal, benign and cancerous tissues from gynecological (GYN) tracts. A comparison of the intensity differences between various Raman modes as well as the number of Raman lines, enables one to distinguish normal GYN tissues from diseased tissues. Fluorescence spectroscopic measurements on human breast tissues show that the ratio of fluorescence intensity at 340 nm to that at 440 nm can be used to distinguish between cancerous and non-cancerous tissues. Separate studies on normal and cancerous breast cell lines show spectral differences. The measurements of back-scattered ultrafast laser pulses from human breast tissues show differences in the scattered pulse profiles for different tissues. These studies show that various optical techniques have the potential to be used in medical diagnostic applications.     

Separation and detection of individual submicron particles by capillary electrophoresis with laser-light-scattering detection

Separation and detection of individual submicron polystyrene spheres using capillary electrophoresis with laser-light-scattering detection has been demonstrated. Electrophoretically separated particles were passed through a focused laser beam and light scattered from individual particles was collected at 90 degrees. Each diameter of polystyrene spheres injected (from 110 to 992 nm) resulted in the observation of a well-defined migration window containing multiple peaks, each arising from the light scattered by an individual particle. The migration time window for individual particles of a particular size corresponded well to the migration time of a peak from a population of particles of the same size detected using a UV absorbance detector. The electrophoretic mobility and scattered light intensity were determined for each particle detected. The average scattered light intensity for each particle size was consistent with Mie scattering theory. Particles as small as 110 nm in diameter were detected individually using this method, but particles with a diameter of 57 nm could not be individually detected. The number of single particle scattering events was counted and compared to the theoretical number of particles injected electrokinetically, and the detection efficiency determined ranged from 38 to 57% for polystyrene spheres of different sizes. The laser-light-scattering detection method was directly compared to laser-induced fluorescence detection using fluorescent polystyrene microspheres. The number of particles detected individually by each method was in agreement.     

Calibration of Polarization-Sensitive and Dual-Angle Laser Light Scattering Methods Using Standard Latex Particles

A polarization-sensitive laser light scattering (PSLLS) method and a dual-angle laser light scattering (DALLS) method have been studied for in situ measurement of submicrometer hydrosol and aerosol particles. By using standard monodisperse polystyrene latex particles suspended in water and air as test particles, calibration of systems built based on the above methods have been performed. The effects of light scattered by agglomerated aerosol particles (multiplets) were corrected by considering the fraction of multiplets as determined with an aerosol measurement technique using a differential mobility analyzer. The change in the measured intensities of scattered light with particle diameter was then determined by calculations based on Mie theory. It was shown that the PSLLS system can determine particle diameters as small as approximately 60 nm for the test hydrosol particles and approximately 100 nm for test aerosol particles, respectively. The DALLS system can determine smaller diameters than the PSLLS system for test particles with no light absorption. The change in scattered light intensities with particle diameter was also investigated by theoretical calculations with various refractive indexes and scattering angles. The PSLLS and DALLS systems promise to become routine measurement tools for absorbing and nonabsorbing particles, respectively. Copyright 2001 Academic Press.     

Rapid Detection of Foot-and-Mouth Disease Virus with Optical Microchip Sensors

Rapid identification of infectious disease pathogens such as foot-and-mouth disease virus (FMDV) during new outbreaks of disease is of fundamental importance in disease control. SpectroSens^TM optical microchip sensors demonstrating rapid, label-free detection of FMDV are presented; these contain multiple high-precision planar Bragg gratings and function as low-cost, robust refractive-index sensors. Sensor selectivity to FMDV is imparted by functionalising the top-surface of specific sensing channels with anti-FMDV monoclonal antibodies (mAbs). Selective binding of cognate antigens within the test sample to surface-immobilised FMDV mAbs results in localised changes in refractive index within specific sensing channels; these antibody-antigen interactions manifest as increases in wavelength of light reflected from the multi-channel sensor chip (light is coupled into and out of the chip via optical fibres). Selective identification of FMDV within minutes of sample introduction has been demonstrated by referenced measurement of changes in sensor reflected wavelength from anti-FMDV channels against sensor controls; simplified ‘snap-shot’ assay data are displayed in the form of a simple yes/no readout using a robust, hand-portable device, with further semi-quantitative information available to the ‘super-user’. The characteristics of the SpectroSens^TM multiplexed detection platform highlight its potential for in-field detection of foot-and-mouth disease and prospective expansion into diagnoses of other infectious veterinary diseases.     

Label-free detection of a bacterial pathogen using an immobilized siderophore, deferoxamine

Pathogenic bacteria obtain the iron necessary for survival by releasing an iron chelator, termed a siderophore, and retrieving the iron-siderophore complex via a cell surface siderophore receptor. We have exploited the high affinity of Yersinia enterocolitica for its siderophore, deferoxamine, to develop a rapid method for capture and identification of Yersinia. In this methodology, a deferoxamine-bovine serum albumin conjugate is printed onto a gold-plated chip in a parallel line pattern. After flowing a suspension of Yersinia across the siderophore-derivatized chip, any Yersinia that binds to the chip is detected by dark-field microscopy analysis of the scattered light, followed by Fourier transform analysis of the scattering pattern. Since peak intensities are found to correlate with pathogen concentration, pathogen titers as low as 10(3) cfu/ml can be readily detected. Moreover, immobilized deferoxamine can distinguish Y. enterocolitica, which binds ferrioxamine (deferoxamine-Fe), from Staphylococcus aureus, Mycobacterium smegmatis and Pseudomonas aeruginosa, which don't. Because human pathogens cannot easily mutate their iron retrieval systems without loss of viability, we suggest that few if any mutant Yersinia will emerge that can avoid detection. Together with previous results demonstrating selective capture of Pseudomonas aeruginosa by its immobilized siderophore (pyoverdin), these data suggest that pathogen-specific siderophores may constitute effective and immutable capture ligands for rapid detection and identification of their cognate pathogens.     

Multiple light scattering as a probe of foams and emulsions

Light propagating in foams or emulsions is strongly scattered by the gas–liquid or liquid–liquid interfaces. This feature makes it generally impossible to directly observe the structure and dynamics deep within the bulk of such materials. However, multiple light scattering can be used as the basis of non-invasive experimental techniques that probe the average bubble size, droplet size or the dispersed volume fraction. If the sample is illuminated with a laser, the transmitted or backscattered light forms a speckled interference pattern whose temporal fluctuations reveal the dynamics of internal structural changes. Such changes can be due to coarsening, flocculation, or applied strain. We briefly recall the fundamental principles of multiple light scattering and present an overview of the experimental techniques that have been developed in recent years.     

Constructing a blue light photodetector on inorganic/organic p-n heterojunction nanowire arrays

We described a new structure photodetector, which is constructed by p-n heterojunction nanowire arrays of PANI (polyaniline)/CdS. The nanowire arrays exhibit excellent rectifying features and a diode nature and show a sensitive spectral response to blue light under 420 nm. The rectification ratio plots of different illumination intensities show straight line behavior, implying that the quantitative detection of illumination intensity can be achieved. The p-n heterojunction nanowire array is a great candidate for applications in high-sensitivity and high-speed blue light photodetectors.     

Effect of the Wavelength of the Laser Beam on the Response of an Evaporative Light Scattering Detector

Abstract

The effects of the wavelength of the laser beam on the response of an evaporative light scattering detector (ELSD) are discussed. Data characterizing the response of the detector and its dependence on the sample size have been collected for six solutes, using a pulsed dye laser as light source. The experimental results suggest that there is little influence of the wavelength on the intensity of the scattered light. On the other hand, the noise decreases in proportion to the wavelength of the incident light beam. Thus, the detection limit (at constant value of the signal to noise ratio) decreases with decreasing wavelengths. The performance of the ELSD improves when a short wavelength is used.     

Fingerprinting bacterial strains using ion mobility spectrometry

Ion mobility spectrometry (IMS) is currently in widespread use for the detection and identification of narcotic and explosive compounds without prior sample clean-up or concentration steps. IMS analysis is rapid, less than a minute, and sensitive, with detection limits in the nanogram to picogram range, depending on the target analyte. Our studies indicate that this technique has potential for detection of specific components of bacterial cells and for identification and differentiation of bacterial strains and species within a minute, and with no specialized test kits or reagents required. When microgram quantities of whole bacterial cells are thermally desorbed, complex positive or negative ion patterns (plasmagrams) are obtained. These plasmagrams differ reproducibly for different strains and species and for different conditions of growth, and can be used for the classification and differentiation of specific strains and species of bacteria, including pathogens. Methods for improved ion peak detection, most notably sequential sample desorption at stepped increases in temperature (programmed temperature ramping), are described.     

Gelatin and the Tyndall Effect: A Colorful and Tasty Demonstration

This demonstration is a safe and nontoxic example of the Tyndall effect that utilizes common, even edible, materials. A glass of gelatin and a laser pointer are used to illustrate the Tyndall effect. Laser light is scattered when it is shone through a colloidal gelatin dispersion. At least two glasses of gelatin are aligned so that students can see scattering through the first glass, then a lower overall intensity of scattered light through the second glass. Variations of the demonstration are presented, such as shining the laser light through a glass of salt water to show the effect as the salt dissolves.     

Light scattering of polyaniline films responding to electrochemical switching

When a laser with 543, 668 or 790±50 nm was used to irradiate the reduced polyaniline film on an indium tin oxide electrode in hydrochloric acid, the light was scattered in all directions. The intensity of the scattered light decreased with increase in the detection angle up to the right angle. When the electrode potential was scanned between the insulating and the conducting domain, the intensity varied sigmoidally with hysteresis. The intensity increased with a decrease in the absorbance. The scattering of the light can be ascribed to multiple reflection, luminescence, or Rayleigh scattering. The spectra of the scattered light was identical with that of the incident light, suggesting the absence of luminescence. The intensity increased with an increase in thickness of the film, indicating a negligible contribution of multiple reflection. Since the volume of the oxidized film is larger than that of the reduced one, the film synthesized in the oxidized state is deformed by the electrode reduction. Then, the film density becomes locally inhomogeneous and this may give rise to the Rayleigh scattering. The potential-variation of the light scattering occurred at a more negative potential than that of the absorbance at 310 nm and of the current did.     

Towards single-microorganism detection using surface-enhanced Raman spectroscopy

The identification and discrimination of microorganisms is important not only for clinical reasons but also for pharmaceutical clean room production and food-processing technology. Vibrational spectroscopy such as IR, Raman, and surface-enhanced Raman scattering (SERS) can provide a rapid ‘fingerprint’ on the chemical structure of molecules and is used to obtain a ‘fingerprint’ from microorganisms as well. Because of the requirement that a single bacterium cell and noble metal nanoparticles must be in close contact and the lack of a significant physical support to hold nanoparticles around the single bacterium cell, the acquisition of SERS spectra for a single bacterium using colloidal nanoparticles could be a challenging task. The feasibility of SERS for identification down to a single bacterium is investigated. A Gram-negative bacterium, Escherichia coli, is chosen as a model for the investigation. Because the adsorption of silver nanoparticles onto the bacterial cell is an exclusive way for locating nanoparticles close to the bacterium cell, the absorption characteristics of silver nanoparticles with different surface charges are investigated. It is demonstrated that the citrate-reduced colloidal silver solution generates more reproducible SERS spectra. It is found that E. coli cells aggregate upon mixing with silver colloidal solution, and this may provide an additional benefit in locating the bacterial cell under a light microscope. It is also found that a laser wavelength in the UV region could be a better choice for the study due to the shallow penetration depth. It is finally shown that it is possible to obtain SERS spectra from a single cell down to a few bacterial cells, depending on the aggregation properties of bacterial cells for identification and discrimination.