The quenching of electrochemiluminescence upon oligonucleotide hybridization

• Published in: Luminescence (Chichester, England) England), 2004-09, Vol.19 (5), p.287-295
• Main Authors: Spehar, Anna-Maria, Koster, Sander, Kulmala, Sakari, Verpoorte, Elisabeth, de Rooij, Nico, Koudelka-Hep, Milena
• Format: Article
• Language: English
• Subjects: Carbocyanines - chemistry; electrochemiluminescence; Electrochemistry; Genome; hybridization; labelling; Luminescent Measurements; Nucleic Acid Hybridization - methods; oligonucleotide; Oligonucleotide Array Sequence Analysis - methods; Oligonucleotides - chemistry; Organometallic Compounds - chemistry; quenching; Sensitivity and Specificity
• ISSN: 1522-7235; 1522-7243
• DOI: 10.1002/bio.786

Many genomic assays rely on a distance‐dependent interaction between luminescent labels, such as luminescence quenching or resonance energy transfer. We studied the interaction between electrochemically excited Ru(bpy)32+ and Cy5 in a hybridization assay on a chip. The 3′ end of an oligonucleotide was labelled with Ru(bpy)32+ and the 5′ end of a complementary strand with Cy5. Upon the hybridization, the electrochemiluminescence (ECL) of Ru(bpy)32+ was efficiently quenched by Cy5 with a sensitivity down to 30 nmol/L of the Cy5‐labelled complementary strand. The quenching efficiency is calculated to be 78%. A similar phenomenon was observed in a comparative study using laser‐excitation of Ru(bpy)32+. The hybridization with the non‐labelled complementary or labelled non‐complementary strand did not change the intensity of the ECL signal. Resonance energy transfer, electron transfer and static quenching mechanisms are discussed. Our results suggest that static quenching and/or electron transfer are the most likely quenching mechanisms. Copyright © 2004 John Wiley & Sons, Ltd.