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Vol 32(2016) N 5 p. 69-76; DOI 10.21519/0234-2758-2016-32-5-69-76
K.A. Lukyanenko1, I.A. Denisov1*, A.S. Yakimov1, E.N. Esimbekova1,2, K.I. Belousov3, A.S. Bukatin1, I.V. Kukhtevich1, V.V. Sorokin1, A.A. Evstrapov3,4, P.I. Belobrov1,2

Analytical Enzymatic Reactions in Microfluidic Systems

1The Siberian Federal University, Krasnoyarsk Russia
2The Institute for Biophysics, Russ. Acad. Sci., Siberian Branch, Krasnoyarsk Russia
3The ITMO University, St.-Petersburg Russia
4The Institute for Analytical Instruments, St.-Petersburg Russia

*d.ivan.krsk@gmail.com

References

An approach to transfer enzyme reactions into the functional elements of the microfluidic chip has been suggested. It was developed and evaluated using the bienzyme bioluminescent reaction with the NAD(P)H:FMN-oxidoreductase and luciferase enzymes. The assessment of the catalytic activity of these enzymes is the basis of the enzymatic bioassay of various liquids. It was found out that all components of the reaction should be placed in one reaction chamber of microfluidic chip to increase the reproducibility of measurements. The use of starch gel as a carrier for immobilization and gelatin scaffold allowed to preserve the enzyme activity during the room-temperature sealing of the chips. To improve the efficiency of the analysis, it isnecessary to use the active mixing of a sample in the reaction chamber of the chip. The results of the study permitto suggest a prototype of a microfluidic chip based on enzyme bioluminescent reaction for the bioassay with the detection limit of copper sulfate of 3 мM, which is in consent with the traditional lux-biosensors based on living cells. Moreover, the analysis time was reduced up to 1 min, and the procedure could be performed by staff without special laboratory skills.

bioluminescence, enzymatic bioassay, luciferase, microfluidics, microfluidic system.



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