Raman Spectroscopy of Gases and Liquids

Raman scattering
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Robert N. Compton and Michael A. Duncan

Suppliers Products News White papers Add your company. Method and apparatus for monitoring and controlling characteristics of process effluents. In the case of gases, information about rotational energy can also be gleaned. An unexpected error occurred. In one embodiment, the invention comprises a system for optimizing basic absorbent solution regeneration in a gas processing plant.

Development of this sensor can follow an iterative path that leads to the creation of a new clinical early detection tool. By using commercial, hand-held Raman devices, microfluidics, and multiplexing it is possible to perform infrastructure-free, field-based diagnostics for constellations of disease markers. Endoscopic sensors can be made with nanofabrication methods to place the sensor on the end of an optical fiber.

Micron sized optical fibers allow sensor placement inside 30 gauge needles for in vivo analysis. These schemes extend the power of SERS detection beyond the lab or clinic and into the world. Early disease detection is critical to saving lives and utilizing the least invasive or irreversibly damaging treatment procedures.

Can Raman spectra be obtained from solids, liquids and gases?

For example, finding life-threatening cancers at the pre-cancer or carcinoma in situ phase can allow for endoscopic excision of the neoplasm before its progression requires treatment with radiation and chemotherapy. Similarly, early detection and treatment of respiratory bacterial infections can reduce permanent damage to the airway or lungs and reduce the duration of infectiousness to help prevent disease transmission. In addition to improving patient health, early detection can reduce health-care costs for both hospitals and patients through simpler treatments and preventative care.

Around the world more than , people die per year as a result of this disease. However, if detected early and treated with life-style changes or pre-cancer targeted excision, the prognosis improves dramatically.

Raman effect

To that end, there have been a great number of studies to determine various correlated pre-cancerous markers, but no method to detect these marker constellations simultaneously in a clinical setting. Around the world, approximately 45 million people contract the disease and approximately , die each year, with the highest mortality rate in infants less than 4 months old.

Internationally, it is the leading causes of vaccine preventable death. Even amongst the well-vaccinated American populace, the rate of infection is on the rise - represented by focal outbreaks in California in , Washington and Vermont in - with the highest rate of infections since the s. Early treatment can cut mortality, shorten the infectious period, and abate respiratory tract damage. However, due to a partially vaccinated populace and the typical delay in seeking treatment, current methods of detection, including qPCR and culturing, can be ineffective for early detection.

Therefore a new, direct oral or serological detection method is needed for early diagnosis. For example, the bacteria B. Unfortunately, the lack of animal models and the inability to track phenotypic specific secretions in situ has left scientists in the dark as to what environmental conditions propel the bacteria from phase to phase - even to the point that there is no consensus on what causes the "whoop" or the cough in whooping cough.

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The devices described in the present disclosure can enable a method to track the bacteria's phenotypic state in real-time, in situ and in vitro, to obtain a greater understanding of the possible in vivo bacterial growth cycle to inform methods for early detection and eradication.

In Raman spectroscopy, laser light is scattered off a molecule and returns at a different wavelength, having donated or accepted energy from a vibrational mode. The wavelength differences between the incident and collected light gives unique information about the molecule's chemical bonding, conformation and atomic species; by collecting a spectrum of these shifts, it is possible to essentially find a "chemical fingerprint. However, as described above, by creating nano structured metallic surfaces it is possible to amplify this effect in a technique known as surface enhanced Raman spectroscopy SERS.

When combined with aptamer sensitization of the gold bulbs, it is possible to measure sub-clinical concentrations of proteins, for example measuring pM to fM concentrations of thrombin as illustrated in Fig. The measurements represented in Fig. Instead, when the thrombin is captured by the aptamer, it is possible to see in the SERS spectrum, unique chemical structures only found in the thrombin that was not present before binding. Furthermore, the measurements in Fig.

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This stems from the fact that the signal intensity of both of these spectroscopic techniques can be heavily dependent on particular optical excitation and collection conditions, and SERS can also be dependent on the location- specific enhancement factor. Therefore, particularly for SERS, it can be difficult to have an "apples to apples" comparison when comparing concentration measurements on different chips or even different locations on a single chip.

In the present disclosure, a method is described to perform "relative peak quantification," that takes advantage of the fabrication methods of the present disclosure to normalize chip to chip or optical variability. In some embodiments, these peaks were 1 the thrombin binding aptamer TBA and 2 thrombin. At each thrombin concentration the same thrombin specific peak intensity can be compared to the TBA specific peak intensity and this ratio can be plotted with respect to thrombin concentration.

It is thus possible to find that the thrombin:TBA peak intensity ratio scales linearly with thrombin concentration. Therefore, when analyzing a sample with an unknown concentration of thrombin, the concentration can be deduced by finding the relative SERS peak ratio and locating where it sat on the concentration curve.

For example, data collected at pM concentrations on three separate chips is shown in Fig. This quantification method is based on two unique features. The first feature is the ability to measure both the "yardstick" and "target" peaks in a single measurement.

By measuring both the TBA and thrombin peaks in a simultaneous measurement, any optical measurement imperfection misalignment, defocusing etc. By taking a 5- 10s measurement any differential contributions based on probe gyration can also be averaged out. Since the substrates used for the measurement of Fig. Therefore, each additional bound thrombin located in a field maximum contributed equally to an increase in the relative intensity of its unique peak. Some peaks are shifted and some peaks are new thrombin specific peaks. The graph on the left illustrates the intensity ratio of thrombin, thrombin binding aptamer TBA vs thrombin concentration on 3 separate chips.

Raman Spectroscopy on Liquids and Solids with the tecRaman Probe

The graph on the right illustrates optical denaturing and recovery of TBA. Guanine peaks are blue-shifted due to reduced steric hindrance, while conformation dependent peaks disappear during denaturing and reappear during TBA recovery. The process, an exemplary embodiment of which is illustrated in Fig. To prevent non-specific binding any uncomplexed TAA can be passivated with acetic anhydride Photolysis of the DECM-CG can be performed with light at wavelengths between about nm, conveniently capturing two common photolithographic exposure lines nm and nm.

A photo-lithographic mask can be used to expose a certain area of functionalized nano-bulbs releasing the DECM-CG, re-exposing the amine in that region A molecular probe can be attached to the amine group followed by acetic anhydride passivation of any left-over amine groups. Then a new area will be uncaged by the photo-mask, functionalized and passivated, and so on, iteratively. At each functionalization step, completely different binding agents, such as antibodies, aptamers , cDNA, etc.

While this repetitive process may seem time intensive to perform on a single-chip, using the parallelism of wafer scale lithography thousands of chips could be sensitized simultaneously. For example, the TAA linker may be replaced with any suitable linker comprising an amine group, and the DECM-CG can be replaced with any suitable photo-uncageable linker.

Additionally, any appropriate passivating agent may be used instead of the acetic anhydride. Furthermore, the above steps of creating a multiplex sensor need not be performed on a chip as illustrated, for example, in Fig. Any suitable substrate may be used to attach the amine-comprising linker and then the photo- uncageable linker to the amine linker. For example, a silicon or glass surface may be used in the method of fabricating a multiplex sensor as described above. One of the oral cancer markers is Interleukin-8 IL In some embodiments, a first step comprises functionalizing the gold nano-bulbs on several chips with an IL-8 antibody AB.

After functionalization, microscope-coupled SERS can be performed within a fluidic environment containing appropriate buffers to determine the unique spectral features of the antibody. For each concentration, the relative intensity of IL-8 vs. AB specific peaks can be considered to perform relative peak concentration quantification as described in the present disclosure. In another step, the detection specificity can be tested against other commonly coincident proteins due to oral cancer, such as IL-6 and IL For functionalization, thiolated cDNA oligos can be commercially obtained; however, they are typically provided as two oligos linked by a disulfide bond.

Methods to cleave this bond into two thiols, without re-oxidation, can be carried out through the use of biochemical tools such as reductants and de-salting columns to prepare DNA for functionalization. As with the protein measurements at several sub- to supra- clinical concentrations, measurements can be performed to determine the relative-peak quantification curve. To uniquely identify mRNA, one possibility is to determine the presence of Uracil and ribose sugars in the Raman spectrum.

In this step the multiplexed functionalization method of Fig. After sensitization the chip can be placed in a buffered fluidic environment where SERS measurements can be taken of the molecular probes on each half of the nano-bulb array and compared to previously collected spectra to check for cross contamination during functionalization.

Subsequently, IL-8 protein and mRNA can be introduced in a single mixed sample with the analytes at various concentrations that simulate baseline, pre-cancerous, and symptomatic oral cancer samples. SERS can be performed on both halves of the nano-bulbs and the relative peak intensity versus concentration results can be compared to those previously collected to check for any confounding effects of the photolytic functionalization.

In these embodiments, an alternative probe can be used.

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For example, a recently created IL-8 specific aptamer could be used as a probe. To prevent this, post-functionalization passivation of any empty bulb areas can be considered, with a mixture of short and long-chain alkane-thiols. This method can create a carpet to fill in the gaps between any probes, such as cDNA probes, and stop physisorption of nucleic acids such as mRNA directly onto the gold.