Development of a Specific Monoclonal Antibody for the Quantification of Artemisinin in Artemisia annua and Rat Serum

Abstract

Artemisinin, extracted from Artemisia annua, and its derivatives are important frontline antimalarials. To produce specific antibodies for the detection and quantification of artemisinin, artemisinin was transformed to 9-hydroxyartemisinin by microbial fermentation, which was used to prepare a 9-succinate artemisinin hapten for conjugation with ovalbumin. A monoclonal antibody (mAb), designated as 3H7A10, was selected from hybridoma cell lines which showed high specificity to artemisinin. No competitive inhibition was observed with artesunate, dihydroartemisinin, and artemether for up to 20,000 ng mL−1. An indirect competitive enzyme-linked immunosorbent assay (icELISA) was developed, which showed a concentration causing 50% of inhibition (IC50) for artemisinin as 2.6 ng mL−1 and a working range of 0.6–11.5 ng mL−1. The icELISA was applied for the quantification of artemisinin in crude extracts of wild A. annua and the study of pharmacokinetics of artemisinin in rat serum after intraperitoneal injection. The results were highly correlated with those determined by HPLC-UV analysis (R2=0.9919). In comparison with reported anti-artemisinin mAbs which have broad cross reactivity with other artemisinin derivatives, the high specificity of 3H7A10 for artemisinin will enable development of methods for quantification of artemisinin in Artemisia plants and antimalarial drugs such as Arco®, and for pharmacokinetic studies.

The effects of mepiquat chloride on the lateral root initiation of cotton seedlings are associated with auxin and auxin-conjugate homeostasis

“…Monoclonal anti-GhIAR3a antibodies were generated in accordance with previously described procedures [43]. Briefly, Balb/c mice were immunized with GhIAR3a protein.…”

“…Sandwich ELISAs was performed as follows: Wells of microtiter plates were coated with rabbit serum and diluted in 0.05 M carbonate buffer at 37 °C for 3 h. Fifty microliters of standard-sample GhIAR3a or plant extract was then added to each well, followed by 50 μL of mAbs at 37 °C for 30 min. The subsequent procedure was performed as described in [43]. The data were calculated via OriginPro 9 (OriginLab) software.…”

Development of monoclonal antibody-based immunoassays for quantification and rapid assessment of dihydroartemisinin contents in antimalarial drugs

“…In this study, inspired by the transformation of artemisinin to DHA [16], we developed a mAb against DHA using a novel hapten 9-O-succinyldihydroartemisinin, which exhibited almost no cross-reactivity to artesunate and artemether. With this DHA antibody, an indirect competitive ELISA (icELISA) [13] and a colloidal gold-based lateral flow immunoassay were developed and evaluated for monitoring DHA content in ACTs.…”

Quality Testing of Artemisinin-Based Antimalarial Drugs in Myanmar

“…To facilitate ACA quality surveys in remote endemic settings, we developed antibody-based lateral-flow dipsticks as point-of-care devices for qualitative and semi-quantitative measurement of artemisinin derivatives (artemether, artesunate, and dihydroartemisinin) in ACAs. [18][19][20][21][22]…”