Monitoring ATP release from individual cells with a biosensor

Romanov, Roman A.; Хохлов, А. А.; Bystrova, M. F.; Rogachevskaja, Olga A.; Yatzenko, Y. E.; Kolesnikov, Stanislav S.

doi:10.1134/s1990747807030075

Cited by 3 publications

(9 citation statements)

References 13 publications

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“…2A). Such properties indicate that ion channels are the main structures responsible for transportation of ATP from the taste cells [2,8,20]. The absence of the dependence of ATP secretion on the calcium level was an additional confirmation of the above conclusion.…”

Section: Resultssupporting

confidence: 68%

“…It was found that, in response to depolarization of nearly all type-II cells, the level of Са 2+ in the cytoplasm of sensor cells increased, which was indicative of depolarizationinduced secretion of ATP by the taste cells. It should be noted that ATP secretion by cells of types I and III was not recorded; ATP secretion appeared to be a phenomenon strictly specific for type-II cells (C, D) [2,20].…”

Section: Resultsmentioning

confidence: 91%

“…This probably explains the fact that classical synaptic structures in the type-II cells are absent. Therefore, we conclude that ATP functions as a neurotransmitter transferring taste information on bitter and sweet and also on the taste of a few amino acids from type-II taste cells to afferent terminals of the gustatory nerves [2,8,10,20,21].…”

Section: Resultsmentioning

confidence: 99%

“…Our experiments showed that the studied COS-1 cells are more effective as ATP sensors. Their selectivity with respect to ATP exceeded modified cells of lines НЕК-293 and СНО, and they are easy to use [20].…”

Section: Resultsmentioning

confidence: 99%

“…For examination of the release of ATP from single isolated cells, application of a conventional luciferin-luciferase technique is impossible, since a significant number of cells is necessary in this case. An alternative approach is the use of ATP-sensitive cells, i.e., ATP biosensors [19,20]. The use of sensor cells makes it possible to record the probable release of ATP from single taste cells under conditions of simultaneous recording of the activity of both a sensor cell and a taste cell (Fig.…”

Section: Resultsmentioning

confidence: 99%

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Cell-to-Cell Communication in the Taste Bud: ATP and Acetylcholine as Primary Mediators

et al. 2012

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It was shown that physiological processes in taste buds (peripheral sensory gustatory organs in vertebrates) are realized with the involvement of several signal systems. In these structures, a number of "classical" neurotransmitters, including glutamate, serotonin, GABA, ATP, noradrenaline, and others, as well as receptors to these agents, were identified. The physiological roles of the above systems (separate ones and all as a whole) remain, however, far from final elucidation. We studied purinergic and cholinergic systems in the taste buds. Based on the data obtained in behavioral experiments using knockout animals, which indicated that ATP is an afferent neurotransmitter, we found stimulation-induced secretion of ATP by type-II cells. The release of ATP does not require the entry of external calcium and is mediated by ion channels permeable for ATP. The obtained data allowed us to explain the fact that classical synaptic structures are absent in the type-II cells. The type-I cells coat other elements including type-II cells; they provide formation of compartments in the intercellular space of the taste buds (this limits ATP diffusion). We showed that taste cells of just type I mostly generate calcium signals in response to the action of ATP and acetylcholine. These cell responses are generated with the involvement of metabotropic purine receptors (isoforms P2Y1, P2Y2, and P2Y4) and muscarinic receptors (isoforms M1, M3, and M5), respectively. Functioning of these receptors is combined with a phosphoinositide cascade, mobilization of intracellular Са 2+ , and subsequent activation of calciumactivated Cl -channels. It seems probable that purinergic and cholinergic signal systems in type-I cells are elements of negative feedback in the taste buds, which promote the process of adaptation to the action of gustatory stimuli.

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Section: Resultssupporting

confidence: 68%

Section: Resultsmentioning

confidence: 91%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

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Cell-to-Cell Communication in the Taste Bud: ATP and Acetylcholine as Primary Mediators

et al. 2012

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Highly active engineered-enzyme oriented monolayers: formation, characterization and sensing applications

Ulman

Ioffe

Patolsky

et al. 2011

Journal of Nanobiotechnology

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BackgroundThe interest in introducing ecologically-clean, and efficient enzymes into modern industry has been growing steadily. However, difficulties associated with controlling their orientation, and maintaining their selectivity and reactivity is still a significant obstacle. We have developed precise immobilization of biomolecules, while retaining their native functionality, and report a new, fast, easy, and reliable procedure of protein immobilization, with the use of Adenylate kinase as a model system.MethodsSelf-assembled monolayers of hexane-1,6-dithiol were formed on gold surfaces. The monolayers were characterized by contact-angle measurements, Elman-reagent reaction, QCM, and XPS. A specifically designed, mutated Adenylate kinase, where cysteine was inserted at the 75 residue, and the cysteine at residue 77 was replaced by serine, was used for attachment to the SAM surface via spontaneously formed disulfide (S-S) bonds. QCM, and XPS were used for characterization of the immobilized protein layer. Curve fitting in XPS measurements used a Gaussian-Lorentzian function.Results and DiscussionWater contact angle (65-70°), as well as all characterization techniques used, confirmed the formation of self-assembled monolayer with surface SH groups. X-ray photoelectron spectroscopy showed clearly the two types of sulfur atom, one attached to the gold (triolate) and the other (SH/S-S) at the ω-position for the hexane-1,6-dithiol SAMs. The formation of a protein monolayer was confirmed using XPS, and QCM, where the QCM-determined amount of protein on the surface was in agreement with a model that considered the surface area of a single protein molecule. Enzymatic activity tests of the immobilized protein confirmed that there is no change in enzymatic functionality, and reveal activity ~100 times that expected for the same amount of protein in solution.ConclusionsTo the best of our knowledge, immobilization of a protein by the method presented here, with the resulting high enzymatic activity, has never been reported. There are many potential applications for selective localization of active proteins at patterned surfaces, for example, bioMEMS (MEMS - Micro-Electro-Mechanical Systems. Due to the success of the method, presented here, it was decided to continue a research project of a biosensor by transferring it to a high aspect ratio platform - nanotubes.

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Enhanced Luminescent Detection of Circulating Tumor Cells by a 3D Printed Immunomagnetic Concentrator

et al. 2021

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Circulating tumor cells (CTCs) are an indicator of metastatic progression and relapse. Since non-CTC cells such as red blood cells outnumber CTCs in the blood, the separation and enrichment of CTCs is key to improving their detection sensitivity. The ATP luminescence assay can measure intracellular ATP to detect cells quickly but has not yet been used for CTC detection in the blood because extracellular ATP in the blood, derived from non-CTCs, interferes with the measurement. Herein, we report on the improvement of the ATP luminescence assay for the detection of CTCs by separating and concentrating CTCs in the blood using a 3D printed immunomagnetic concentrator (3DPIC). Because of its high-aspect-ratio structure and resistance to high flow rates, 3DPIC allows cancer cells in 10 mL to be concentrated 100 times within minutes. This enables the ATP luminescence assay to detect as low as 10 cells in blood, thereby being about 10 times more sensitive than when commercial kits are used for CTC concentration. This is the first time that the ATP luminescence assay was used for the detection of cancer cells in blood. These results demonstrate the feasibility of 3DPIC as a concentrator to improve the detection limit of the ATP luminescence assay for the detection of CTCs.

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Monitoring ATP release from individual cells with a biosensor

Cited by 3 publications

References 13 publications

Cell-to-Cell Communication in the Taste Bud: ATP and Acetylcholine as Primary Mediators

Cell-to-Cell Communication in the Taste Bud: ATP and Acetylcholine as Primary Mediators

Highly active engineered-enzyme oriented monolayers: formation, characterization and sensing applications

Enhanced Luminescent Detection of Circulating Tumor Cells by a 3D Printed Immunomagnetic Concentrator

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