Zachary Coker

Design and development of a simple UV fluorescence multi-spectral imaging system

Healthcare access in low-resource settings is compromised by the availability of affordable and accurate diagnostic equipment. The four primary poverty-related diseases – AIDS, pneumonia, malaria, and tuberculosis - account for approximately 400 million annual deaths worldwide as of 2016 estimates. Current diagnostic procedures for these diseases are prolonged and can become unreliable under various conditions. We present the development of a simple low-cost UV fluorescence multi-spectral imaging system geared towards low resource settings for a variety of biological and in-vitro applications. Fluorescence microscopy serves as a useful diagnostic indicator and imaging tool. The addition of a multi-spectral imaging modality allows for the detection of fluorophores within specific wavelength bands, as well as the distinction between fluorophores possessing overlapping spectra. The developed instrument has the potential for a very diverse range of diagnostic applications in basic biomedical science and biomedical diagnostics and imaging. Performance assessment of the microscope will be validated with a variety of samples ranging from organic compounds to biological samples.

BISTRO measurement also means better measurement (Conference Presentation)

Brillouin microspectroscopy is an emerging technique in optical elastography. It allows measuring local high-frequency viscoelastic modulus in cells and tissues in a matter of seconds or hundreds of milliseconds. BISTRO (Brillouin Imaging and Sensing via Time-Resolved Optical) measurements relies on impulsive stimulated Brillouin scattering to increase the signal strength and, hence, the speed of imaging, which can be as high as 1,000,000 pixels per second. However, there are several other untapped advantages of BISTRO measurements, and a particular intriguing one is the accuracy of Brillouin shift and linewidth assessments, which are substantially improved through BISTRO assessment. This allows high quality measurements not only to be done fast and more accurately, but also allows highly reproducible measurements over an extended period of time.

Developing upconversion nanoparticle-based smart substrates for remote temperature sensing

Recent developments in understanding of nanomaterial behaviors and synthesis have led to their application across a wide range of commercial and scientific applications. Recent investigations span from applications in nanomedicine and the development of novel drug delivery systems to nanoelectronics and biosensors. In this study, we propose the application of a newly engineered temperature sensitive water-based bio-compatible core/shell up-conversion nanoparticle (UCNP) in the development of a smart substrate for remote temperature sensing. We developed this smart substrate by dispersing functionalized nanoparticles into a polymer solution and then spin-coating the solution onto one side of a microscope slide to form a thin film substrate layer of evenly dispersed nanoparticles. By using spin-coating to deposit the particle solution we both create a uniform surface for the substrate while simultaneously avoid undesired particle agglomeration. Through this investigation, we have determined the sensitivity and capabilities of this smart substrate and conclude that further development can lead to a greater range of applications for this type smart substrate and use in remote temperature sensing in conjunction with other microscopy and spectroscopy investigations.

Dual Raman-Brillouin spectroscopic investigation of plant stress response and development

Raman and Brillouin spectroscopy are powerful tools for non-invasive and non-destructive investigations of material chemical and mechanical properties. In this study, we use a newly developed custom-built dual Raman-Brillouin microspectroscopy instrument to build on previous works studying in-vivo stress response of live plants using only a Raman spectroscopy system. This dual Raman-Brillouin spectroscopy system is capable of fast simultaneous spectra acquisition from single-point locations. Shifts and changes in a samples Brillouin spectrum indicate a change in the physical characteristics of the sample, namely mechano-elasticity; in measuring this change, we can establish a relationship between the mechanical properties of a sample and known stress response agents, such as reactive oxygen species and other chemical constituents as indicated by peaks in the Raman spectra of the same acquisition point. Simultaneous application of these spectroscopic techniques offers great promise for future development and applications in agricultural and biological studies and can help to improve our understanding of mechanochemical changes of plants and other biological samples in response to environmental and chemically induced stresses at microscopic or cellular level.

Using Brillouin microspectroscopy to characterize adipocytes’ response to lipid droplet accumulation

Obesity and overweight are accompanied by an enlargement of adipocytes, which is commonly related to the increasing number or size of lipid droplets within the cells. Some studies have shown that the accumulation of lipid droplets within adipocytes results in their increased stiffness. Recently, Brillouin microspectroscopy has been introduced as a nondestructive method of imaging the elasticity of cells. Unlike other imaging modalities, it is capable of assessing the elastic properties on both tissue- and cell levels. In this study, Brillouin spectroscopy was used to measure the elasticity changes in response to accumulation of lipid droplets within adipocyte during adipogenesis. The cell line used in the study is 3T3-L1, with chemically-induced differentiation from pre-adipocytes to mature adipocytes. The Brillouin shift measurements of the cells before and after differentiation indicate that the stiffness of adipocytes increases due to accumulation of lipid droplets. The results are in agreement with previous atomic force microscopy (AFM) nanoindentation studies. Brillouin microspectroscopy is a technique suitable for measuring the changes of elasticity of adipocytes in response to lipid droplet accumulation.

Toward investigating changes in cell mechanoelastic properties in response to nanosecond pulsed electric fields

Nanosecond electric pulses (nsEPs) are known to cause a variety of effects on mammalian cells, ranging from destabilization of cell membranes to changes in cytoskeleton and elastic moduli. Measurement of a cells mechanoelastic properties have previously been limited to only invasive and destructive techniques such as atomic force microscopy or application of optical tweezers. However, due to recent advances, Brillouin spectroscopy has now become viable as a non-contact, non-invasive method for measuring these properties in cells and other materials. Here, we present progress toward applying Brillouin spectroscopy using a unique microscopy system for measuring changes in CHO-K1 cells when exposed to nsEPs of 600ns pulse duration with intensity of 50kV/cm. Successful measurement of mechanoelastic changes in these cells will demonstrate Brillouin spectroscopy as a viable method for measuring changes in elastic properties of other cells and living organisms.

Investigation of burn effect on skin using simultaneous Raman-Brillouin spectroscopy, and fluorescence microspectroscopy

Burns are thermal injuries that can completely damage or at least compromise the protective function of skin, and affect the ability of tissues to manage moisture. Burn-damaged tissues exhibit lower elasticity than healthy tissues, due to significantly reduced water concentrations and plasma retention. Current methods for determining burn intensity are limited to visual inspection, and potential hospital x-ray examination. We present a unique confocal microscope capable of measuring Raman and Brillouin spectra simultaneously, with concurrent fluorescence investigation from a single spatial location, and demonstrate application by investigating and characterizing the properties of burn-afflicted tissue on chicken skin model. Raman and Brillouin scattering offer complementary information about a materials chemical and mechanical structure, while fluorescence can serve as a useful diagnostic indicator and imaging tool. The developed instrument has the potential for very diverse analytical applications in basic biomedical science and biomedical diagnostics and imaging.

What is next for Brillouin microscopy in biology and medicine

Brillouin microscopy is an emerging technique in biomedical imaging capable of non-invasive assessing viscoelastic properties on a microscopic scale. In this report, we outline the latest developments in Brillouin spectroscopy instrumentation and applications in an attempt to anticipate the future impact areas of this new imaging modality.

Brillouin micro-elastography of laser-processed materials

3D laser processing is becoming a mainstream of micro- and nano- fabrication. However, very little is known how physical, chemical and/or structural changes induced by laser irradiation affect local mechanical properties. In this report, we, for the first time, utilize Brillouin microspectroscopy to assess and image the variation of viscoelastic properties of materials induced by laser processing.