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Enzymatic assay for total plasma Cys

Nature Protocols volume 3, pages 1778–1781 (2008)Cite this article

Abstract

Patients with vascular disease and end-stage renal disease have significantly higher concentrations of plasma total Cys (tCys) than do healthy individuals. Described here is a nonradioactive, precise, rapid and sensitive enzymatic colorimetric assay for tCys in plasma samples and is homogeneous in that it avoids separation methods. The tCys assay uses only the recombinant enzymes methionine α,γ-lyase (rMETase) and S-adenosylhomocysteine hydrolase (rSAHH) cloned from Pseudomonas putida and Trichomonas vaginalis, respectively. The rSAHH traps homocysteine and the rMETase thereby produces H2S exclusively from Cys. The reaction product, H2S, is measured colorimetrically following its reaction with N,N-dibutylphenylenediamine (DBPDA). The procedure takes 3–4 h to complete.

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Figure 1
Figure 2: Dilution linearity.
Figure 3: 96-well microplate layout.
Figure 4: Comparison of the enzymatic and HPLC tCys assays.

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  1. AntiCancer, Inc., 7917 Ostrow Street, San Diego, 92111, CA, USA

    Qinghong Han & Robert M Hoffman

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  1. Qinghong Han
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  2. Robert M Hoffman
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Correspondence to Robert M Hoffman.

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Han, Q., Hoffman, R. Enzymatic assay for total plasma Cys. Nat Protoc 3, 1778–1781 (2008). https://doi.org/10.1038/nprot.2008.178

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