1. Heavy chain single-domain antibodies to detect native human soluble epoxide hydrolase
- Author
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Cui, Yongliang, Li, Dongyang, Morisseau, Christophe, Dong, Jie-Xian, Yang, Jun, Wan, Debin, Rossotti, Martín A, Gee, Shirley J, González-Sapienza, Gualberto G, and Hammock, Bruce D
- Subjects
Biochemistry and Cell Biology ,Biological Sciences ,2.1 Biological and endogenous factors ,Amino Acid Sequence ,Animals ,Camelids ,New World ,Cross Reactions ,Enzyme-Linked Immunosorbent Assay ,Epoxide Hydrolases ,Humans ,Molecular Sequence Data ,Sequence Homology ,Amino Acid ,Single-Domain Antibodies ,Soluble epoxide hydrolase ,sEH ,ELISA ,Magnetic beads ,VHH ,Chemical Sciences ,Engineering ,Analytical Chemistry ,Biological sciences ,Chemical sciences - Abstract
The soluble epoxide hydrolase (sEH) is a potential pharmacological target for treating hypertension, vascular inflammation, pain, cancer, and other diseases. However, there is not a simple, inexpensive, and reliable method to estimate levels of active sEH in tissues. Toward developing such an assay, a polyclonal variable domain of heavy chain antibody (VHH) sandwich immunoassay was developed. Ten VHHs, which are highly selective for native human sEH, were isolated from a phage-displayed library. The ten VHHs have no significant cross-reactivity with human microsomal epoxide hydrolase, rat and mouse sEH, and denatured human sEH. There is a high correlation between protein levels of the sEH determined by the enzyme-linked immunosorbent assay (ELISA) and the catalytic activity of the enzyme in S9 fractions of human tissues (liver, kidney, and lung). The VHH-based ELISA appears to be a new reliable method for monitoring the sEH and may be useful as a diagnostic tool for diseases influenced by sEH. This study also demonstrates the broad utility of VHH in biochemical and pharmacological research.
- Published
- 2015