Kulim Kht Bionavis SPR Real-time analysis of DNA

Real-time analysis of DNA hybridization with SAMP-SPR yields high specificity, improved signal-to-noise ratio, and a significantly cleaner sensorgram.

Background Experimental
Surface plasmon resonance (SPR) is a well-established technique for the monitoring of biomolecular interactions. SPR has been frequently used for the real-time analysis of hybridization of DNA and RNA oligonucleotides. One challenge in this context is that SPR is a non-specific detection method, i.e. any substance that adsorbs onto the sensor surface is detected. For example, SPR analysis does not indicate which strands in a mixture of oligos hybridize with an immobilized DNA strand on the chip surface. This is in sharp contrast to the high specificity of fluorescence- based analysis, where only the labelled oligo is detected.

Multi-Parametric SPR (MP-SPR) is a novel method utilizing the same physical principles as SPR, where not only the SPR peak minimum shift, but also other parameters from the optical signal are measured as a function of time.

In this Application Note we demonstrate how real-time SAMP-SPR analysis of DNA hybridization with MP-SPR using oligos labelled with Episentec™ dyes results in a specificity comparable to that of fluorescence analysis. Also the sensitivity sensitivity improved and disturbing signals are reduced, resulting in a better signal to noise ratio.

Hybridization of both unlabelled (native) and Episentec dye-labelled 25-mer DNA oligonucleotides (Episentec, www.episentec.com) were performed. Firstly, biotin-BSA conju- gate was spontaneously adsorbed onto a clean gold sensor chip followed by binding of avidin. A 25-mer DNA oligo with a spacer coupled to a biotin entity was then bound to the avidin. Subse- quently, a number of samples containing either native DNA, or DNA labelled with Episentec dye B10, were injected and hybrid- ized. Denaturation was performed with 25 mM sodium hydroxide. All experiments were performed using BioNavis Multi-Parametric SPR Navi™ 200. Enhanced sensorgrams were calculated in accordance with methods implemented in the EpiGrammer™ software.
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