Development of indirect competitive ELISA for quantification of mitragynine in Kratom (Mitragyna speciosa (Roxb.) Korth.)

Supattra Limsuwanchote, Juraithip Wungsintaweekul, Niwat Keawpradub, Waraporn Putalun, Satoshi Morimoto, Hiroyuki Tanaka

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Monoclonal antibody (MAb) against mitragynine (MG), an analgesic alkaloid from Kratom leaves (Mitragyna speciosa), was produced. MG was coupled to carrier proteins employing either 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS), a zero-length cross linker or a 5-carbon length glutaraldehyde cross linker. To confirm the immunogenicity, the hapten numbers were determined using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Preparation of the MAb was accomplished by the electrofusion method. Hybridoma 1A6 that was constructed from the fusion between splenocytes of EDC/NHS conjugate immunized mice and SP2/0-Ag14 myeloma cells was selected, cloned twice and expanded. The cross-reactivities (CRs) of this MAb 1A6 with a series of indole alkaloids were 30.54%, 24.83% and 8.63% for speciogynine, paynantheine and mitraciliatine, respectively. Using this MAb, an indirect competitive enzyme-linked immunosorbent assay (icELISA) was developed with a measurement range of 32.92-250μg/mL. Quantitative analysis of the MG contents in plant samples by icELISA correlated well with the standard high performance liquid chromatography method (R2=0.994). The MAb against mitragynine provided a tool for detection of MG in Kratom preparations.

Original languageEnglish
Pages (from-to)70-77
Number of pages8
JournalForensic Science International
Volume244
DOIs
Publication statusPublished - Jan 1 2015

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Mitragyna
Enzyme-Linked Immunosorbent Assay
Monoclonal Antibodies
Ethyldimethylaminopropyl Carbodiimide
Indole Alkaloids
Carbodiimides
Haptens
Hybridomas
Glutaral
Alkaloids
Analgesics
Mass Spectrometry
Carrier Proteins
Lasers
Carbon
High Pressure Liquid Chromatography
mitragynine

All Science Journal Classification (ASJC) codes

  • Pathology and Forensic Medicine

Cite this

Development of indirect competitive ELISA for quantification of mitragynine in Kratom (Mitragyna speciosa (Roxb.) Korth.). / Limsuwanchote, Supattra; Wungsintaweekul, Juraithip; Keawpradub, Niwat; Putalun, Waraporn; Morimoto, Satoshi; Tanaka, Hiroyuki.

In: Forensic Science International, Vol. 244, 01.01.2015, p. 70-77.

Research output: Contribution to journalArticle

Limsuwanchote, Supattra ; Wungsintaweekul, Juraithip ; Keawpradub, Niwat ; Putalun, Waraporn ; Morimoto, Satoshi ; Tanaka, Hiroyuki. / Development of indirect competitive ELISA for quantification of mitragynine in Kratom (Mitragyna speciosa (Roxb.) Korth.). In: Forensic Science International. 2015 ; Vol. 244. pp. 70-77.
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