Label-free and sensitive detection assay for terminal deoxynucleotidyl transferase via polyadenosine-coralyne fluorescence enhancement strategy

Yuanyuan Wang, Xu Sun, Jianxiong Zeng, Minggang Deng, Nan Li, Qiutong Chen, Hua Zhu, Fenyong Liu, Xiwen Xing

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

Terminal deoxynucleotidyl transferase (TdT) is a unique template-free polymerase that randomly adds multiple deoxyribonucleoside triphosphates (dNTPs) to the 3′-OH terminus of ssDNA. This characteristic makes TdT a versatile enzymatic tool in many fields. Moreover, aberrant TdT expression is a well-recognized biomarker of several leukemic diseases and is related to carcinogenesis. In this study, we developed a facile, rapid, label-free, and convenient assay for TdT detection. TdT-generated poly A tails formed a fluorescent enhancement complex in the presence of coralyne. To achieve a better signal-to-noise ratio, we used potassium thiocyanate (KSCN), instead of other halogen anions (KCl, KBr, KI, NaI) as the quenching agent of dissociate coralyne. Our results demonstrate that this assay is extremely facile, rapid, and label-free; at levels as low as 0.025 U/mL, TdT was distinctly detected within 55 min. And the determination of TdT activity in RBL-2H3 and Reh cells lysates exhibited a good sensing performance, demonstrating its potential applications in biochemical research and clinical diagnosis.

Original languageEnglish (US)
Pages (from-to)85-89
Number of pages5
JournalAnalytical Biochemistry
Volume567
DOIs
StatePublished - Feb 15 2019

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

Keywords

  • Coralyne
  • Fluorescence enhancement
  • Polyadenosine
  • Terminal deoxynucleotidyl transferase

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