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Title 

Monoclonal antibody-based screening assay for factor inhibiting hypoxia-inducible factor inhibitors

Authors 

S H LeeJeong Hee MoonEun-Ah ChoSeong Eon RyuMyung Kyu Lee

Publisher 

SAGE Publications (UK and US)

Issue Date 

2008

Citation 

Journal of Biomolecular Screening, vol. 13, no. 6, pp. 494-503

Keywords 

ELISAfactor-inhibiting hypoxia-inducible factorhypoxia-inducible factor-1αinhibitor screeningmonoclonal antibodylinked immunosorbent assayhigh throughput screeningimmunoassaybiological assayimmunosorbent assay

Abstract 

The factor-inhibiting hypoxia-inducible factor (FIH) hydroxylates the asparagine 803 (Asn803) residue of the hypoxia-inducible factor 1α (HIF-1α), and the modification abrogates the transcriptional activity of HIF-1α. Because FIH is more active on HIF-1α than prolyl hydroxylase domain proteins under hypoxic conditions, its inhibitors have potential to be developed as anti-ischemic drugs targeting normal cells stressed by hypoxia. In this study, the authors developed the first monoclonal antibody, SHN-HIF1α, specifically to Asn803 hydroxylated HIF-1α and a sensitive assay system for FIH inhibitors using the monoclonal antibody (Mab). SHN-HIF1α showed 740 times higher affinity to the Asn803 hydroxylated HIF-1α peptide than the unmodified one. An enzyme-linked immunosorbent assay-based system using SHN-HIF1α displayed at least 30 times more sensitivity than previous methods for screening FIH inhibitors and was easily applicable to develop a high-throughput screening system. SHN-HIF1α also showed an Asn803 hydroxylation-dependent specificity to HIF-1α in cells. Taken together, the results suggest that it may be used to analyze the in vivo and in vitro activities of FIH inhibitors.

ISSN 

1087-0571

Link 

http://dx.doi.org/10.1177/1087057108318800

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2017-04-19


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