Enzyme kinetics on a molecular level with optical microscopy

Volker Uhl; Goetz Pilarczyk; Karl-Otto Greulich

doi:10.1117/12.297961

29 December 1997 Enzyme kinetics on a molecular level with optical microscopy

Volker Uhl, Goetz Pilarczyk, Karl-Otto Greulich

Author Affiliations +

Proceedings Volume 3197, Optical Biopsies and Microscopic Techniques II; (1997) https://doi.org/10.1117/12.297961
Event: BiOS Europe '97, 1997, San Remo, Italy

Abstract

First results with enzyme catalyzed reactions show that examination of reactions of single molecules with classical (non-scanning) fluorescence microscopy is possible. Two picoliters of enzyme solution (lactate dehydrogenase, LDH-1) are injected into an area with an excess of substrate (Lactate, NA⁺). From the dilution of the enzyme solution one can calculate that only few enzyme molecules (in the order of 10²) are injected. After a minute discrete zones with increasing fluorescence intensity from the reaction's product NADH can be observed in a microscope. The variation of the fluorescence intensity and the size of these zones is used to monitor the reaction kinetics and to estimate the number of individual enzyme molecules which catalyzed the reaction in every zone. Three reaction zones are observed which may be caused by 1, 1 and 2 LDH-1 molecules up to 5, 5 and 10 molecules.

Citation Download Citation

Volker Uhl, Goetz Pilarczyk, and Karl-Otto Greulich "Enzyme kinetics on a molecular level with optical microscopy", Proc. SPIE 3197, Optical Biopsies and Microscopic Techniques II, (29 December 1997); https://doi.org/10.1117/12.297961

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