Tatiana N. Goryachkovskaya

Studying the non-thermal effects of terahertz radiation on E. coli/pKatG-GFP biosensor cells

Studies of the impact of terahertz radiation on living objects present a significant interest since its use for security systems is currently considered promising. We studied the non-thermal impact of terahertz radiation on E. coli/pKatG-gfp biosensor cells. The Novosibirsk free electron laser (NovoFEL), which currently has the worlds highest average and peak power, was used as the source of terahertz radiation. We demonstrated that exposure to terahertz radiation at the wavelengths of 130, 150, and 200 µm and a power of 1.4 W/cm(2) induces changes in green fluorescent protein (GFP) fluorescence values and thus induces the expression of GFP in E. coli/pKatG-gfp biosensor cells. Possible mechanisms of the E. coli response to non-thermal exposure to terahertz radiation are discussed.

Impact of Terahertz Radiation on Stress-Sensitive Genes of E.Coli Cell

Terahertz (THz) radiation is increasingly used in scientific studies, as well in practical applications, primarily in medicine and safety systems, therefore, exposure of living cells to THz-waves present significant interest. We studied the impact of THz radiation on E. coli biosensor cells containing plasmids with promoters of stress-sensitive genes controlling the expression of GFP. The impact of THz radiation was nonthermal, i.e., special care was taken to keep specimen temperature in the 35±2°C range during irradiation so that heat shock genes would not be induced. GFP level was measured by fluorometry. We found that THz radiation activates genes associated with oxidative stress response. Lower activation levels were found for copper homeostasis genes, while antibiotics resistance genes showed no response. Thus, THz radiation impact varies for different E. coli stress systems. We conclude that THz radiation significantly affects oxidative homeostasis, which is vital for E. coli cells.

Soft Ablation of Biological Objects Caused by Free-Electron Laser Submillimeter Radiation

In 1919, Aston [1] built the first mass spectrometer, which was used to determine the isotopic composition of many chemical elements. For example, the stable isotopes 13 C, 15 N, 17 O, and 2 H(D) have been discovered. Later, chemists mastered this method. At present, a mass spectrum can be recorded for any substance that can be transferred into a gas phase, the molecular weight being determined from the mother peak of the spectrum, and the molecular structure being reconstructed from the fragments of molecules [2]. The use of mass spectrometry in biology was prevented not so much by the large weights of biological molecules as by the impossibility to transfer them into a gas phase. This obstacle was overcome when the MALDI method [3] had been developed. In this method, a matrix containing protein is exposed to an intense laser impulse 10 –9 –10 –6 s in length. This causes ablation, i.e., knocks molecules and their fragments out of the surface of the substrate. Usually, standard UV or near-IR lasers with a high quantum energy are used, which causes substantial destruction of molecules of the sample analyzed.

Mild ablation of biological objects under the submillimeter radiation of the free electron laser

First line of FEL with smooth tuning of irradiation wavelength 100-200 microns and 400 W power is commissioned in Siberian Center for Photochemical Research. Irradiation was used for mild ablation of DNA, enzymes, and proteins. Transfer from surface occurs without molecular destruction. Particle size was determined using aerosol diffusion spectrometer.

A quantitative method for determination of PPDK concentration in miscanthus leaves

In this study, we used ELISA for quantification of PPDK in photosynthesizing leaves of miscanthus. We cloned a fragment of the gene encoding PPDK, purified the resulting protein by affinity chromatography, identified it using MALDI mass spectrometry, and obtained monoclonal antibodies by immunizing BALB/c mice. Selectivity of monoclonal antibodies was assessed by Western blot using the protein extracts of Soranovskii. The presence of PPDK was again verified by MALDI mass spectrometry. Therefore, we developed and tested the method for determining PPDK quantity in miscanthus using ELISA.

2.3THz radiation: Absence of genotoxicity/mutagenicity in Escherichia coli and Salmonella typhimurium.

The mutagenicity and genotoxicity in bacteria of 2.3THz radiation (THz) produced by a free-electron laser (NovoFEL) were evaluated; exposures were 5, 10, or 15min at average power 1.4W/cm(2). Two Ames mutagenicity test strains of Salmonella typhimurium, TA98 and TA102, were used. For the genotoxicity test, we measured SOS induction in Escherichia coli PQ37. No significant differences were found between exposed and control cells, indicating that THz radiation is neither mutagenic nor genotoxic under these conditions. Nevertheless, a small increase in total cell number of S. typhimurium after 15min exposure, and an increase in β-galactosidase and alkaline phosphatase activities in E.coli PQ37, were observed, indicating some effect of THz radiation on cell metabolism. We also examined the combined effect of 4-NQO (8μM; positive control) and THz exposure (5min) on genotoxicity in E.coli PQ37. Unexpectedly, THz radiation decreased 4-NQO genotoxicity.

Investigating passive immunodiffusion reactions in the channels of a microfluidic device

The passive immunodiffusion reaction in a microfluidic system is investigated via the interaction between fluorescein human albumin conjugate and monoclonal antibodies. The topology of a microfluidic device suitable for this reaction was designed, and the device was fabricated using LIGA technology. A system for transitioning from the macro- to the microlevel is developed and produced. The dissociation constants and specificity of interaction in complexes formed as a result of the reaction between fluorescein human albumin conjugates and monoclonal antibodies are determined. The results from our study reveal that systems based on combinations of passive immunodiffusion reactions and microfluidic technologies are applicable for detecting antigens and antibodies in the analyzed fluids.

FEL THz irradiation approach for the biochip production standardization

The terahertz emission was applied to the development of the technology for the biochip production standardization. The approach is based on the method of soft nondestructive ablation developed by authors, which means the transfer of target DNA to aerosol phase from solid substrate of a biochip under action of the terahertz emission.

Hypotensive and neurometabolic effects of intragastric Reishi (Ganoderma lucidum) administration in hypertensive ISIAH rat strain

BACKGROUND As the standard clinically used hypotensive medicines have many undesirable side effects, there is a need for new therapeutic agents, especially ones of a natural origin. PURPOSE One possible candidate is extract from the mushroom Reishi (Ganoderma lucidum), which is used in the treatment and prevention of many chronic diseases. STUDY DESIGN AND METHODS To study the effectiveness of Reishi, which grows in the Altai Mountains, as an antihypertensive agent, we intragastrically administered Reishi water extract to adult male hypertensive ISIAH (inherited stress-induced arterial hypertension) rats. RESULTS After seven weeks, Reishi therapy reduced blood pressure in experimental animals at a level comparable to that of losartan. Unlike losartan, intragastric Reishi introduction significantly increases cerebral blood flow and affects cerebral cortex metabolic patterns, shifting the balance of inhibitory and excitatory neurotransmitters toward excitation. CONCLUSION Changes in cerebral blood flow and ratios of neurometabolites suggests Reishi has a potential nootropic effect.

GC-based chemoprofile of lipophilic compounds in Altaian Ganoderma lucidum sample

The presented data contains information about component composition of lipophilic compounds in Ganoderma lucidum fungal body sample obtained using gas chromatography and subsequent mass spectrometry.