Versatile analysis of multiple macromolecular interactions by SPR imaging: application to p53 and DNA interaction

Abstract

The greatest challenge in the postgenomic era is the description of proteome interactions, such as protein-protein or protein-DNA interactions. Surface plasmon resonance (SPR) is an optical technique in which binding of an analyte to the surface changes the refractive index at the surface/solution interface. Molecular interactions are analysed in real time without a labeling step. Currently, the limit to SPR imaging is the small number of reactions that can be simultaneously analysed. Using a novel grafting technology and a new imaging system, we increased the throughput of SPR imaging. The interaction between p53 and DNA was chosen as a paradigm for validation of this assay. Using a tagged DNA methodology, we simultaneously targeted multiple DNA sequences on a single chip. The interaction between p53 and these DNA sequences was monitored by SPR imaging. Qualitative and quantitative analysis provides results similar to those obtained with conventional technologies.

Surface plasmon resonance imaging and versatile surface functionalization for real time comparisons of biochemical interactions

“…Thus five different 20 base long dsDNA sequences were chosen. The results, in press, will be very soon published [9]. Basically the main steps are the following.…”

“…Each initial target being linked to a unique zip code, the structure of the functionalized chip will reflect the initial distribution of the zip code probes. We used this technique for functionalization with double strand DNA (dsDNA) and subsequent interaction with a protein, p53 in this case [9].…”

Fano Resonances in Thin Metallic Grating for Refractive Index Sensing with High Figure of Merit

“…Especially in sensing applications, this phenomenon has been exploited in biosensor systems for real-time and label-free biochemical detection [4] with different experimental setup: angular interrogation technique [5], phase interrogation technique [6], and wavelength interrogation technique [7]. Up to now, these experimental approaches have been applied successfully for many biomolecular interaction analyses such as DNA-DNA [8,9], protein-DNA [10], and protein-protein [11] with good detection probability. However, these sensing approaches based on continuous metallic thin film have difficulty detecting biomolecules with small volume and very low concentration, because the effective change in the dielectric constant change at the metal/dielectric interface is too small [12].…”

Plasmonic DNA: Towards Genetic Diagnosis Chips

“…It allows quantification of the binding events, especially the quantity of bound material in quasiequilibrium states and the associated kinetic constants, or more generally the dynamics of the binding and unbinding reactions, allowing biochemical affinity measurements and comparisons. Interactions tested include DNA-DNA [9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24][25][26][27], DNA-protein [28][29][30][31][32][33][34], protein-protein [35,36], and applications such as health safety in food [37][38][39][40][41][42][43] or even microbial detection in space [44].…”