Biomedical subjects

Loïc Dayon

Publications and source records attributed to Loïc Dayon.

5 recordsLinked to original sources

PubMed2006

Multitrack electrospray chips.

Multitrack electrospray chips (MTEC) were fabricated by UV-photoablation of polyethylene terephthalate (PET) substrates. They are composed of an array of up to six microchannels that are successively used as electrospray ionization (ESI) emitters for mass spectrometry (MS). There is no requirement for alignment of the different spraying microchannels with the mass spectrometer orifice. The MTEC is thus fixed in front of the mass spectrometer and the successive MS analyses are performed without moving the chip. The sequential electrospraying by successive application of an identical high voltage in each off-axis microchannel was evaluated for the fast screening of peptides and proteins. The counting of cysteines in peptides through chemical modification and the relative quantification of a peptide in two samples are presented herein as two original strategies based on this new analytical tool.

Journal Article↗

PubMed2006

Probing cysteine reactivity in proteins by mass spectrometric EC-tagging.

The on-line electrochemical tagging (EC-tagging) of cysteine residues in proteins during mass spectrometry is studied to probe the cysteine environment. Benzoquinone probes electrogenerated at a microspray electrode react with the thiol functions of the proteins within a microchannel and the products are analyzed by mass spectrometry. The fundamentals of the technique are discussed, with a focus on the kinetic aspects. The EC-tagging efficiency of the cysteine residues in proteins is used to probe their environment. Experiments with unmodified proteins and their chemically reduced forms highlight the strong effect of the cysteine site reactivity on the tagging efficiencies. This study highlights relevant parameters for such on-line electrochemical derivatization/MS detection strategies.

Animals↗

PubMed2005

On-line counting of cysteine residues in peptides during electrospray ionization by electrogenerated tags and their application to protein identification.

The electrochemically induced mass spectrometric tagging of cysteines by substituted hydroquinones was studied for peptides in a classical electrospray solvent (i.e., MeOH/H2O/AcOH 50/49/1). The tagging efficiency was tested with different hydroquinone compounds on an undecapeptide containing one cysteine residue. 2-carboxymethylhydroquinone was the most reactive probe and revealed to be suitable for cysteine quantification in peptides containing up to three cysteine residues, even in the case of two consecutive cysteines in the sequence. We demonstrate the compatibility of the on-line electrochemical tagging method for the cysteine content analysis of peptides coming from gel-free protein digestion procedures. The identification of bovine serum albumin and human alpha-lactalbumin digest samples in a peptide mapping strategy was greatly improved by the application of the electrotagging technique as post-column treatment. Indeed, the determination of cysteine content in the tryptic peptides provided powerful information in order to enhance the identification score as well as the discrimination against other protein candidates. The tagging method was then applied to the determination of four proteins in a model mixture.

Animals↗

PubMed2004

Generation of mass tags by the inherent electrochemistry of electrospray for protein mass spectrometry.

We present herein a review of our work on the on-line electrochemical generation of mass tags toward cysteine residues in peptides and proteins. Taking advantage of the inherent electrochemical nature of electrospray generated from a microfabricated microspray emitter, selective probes for cysteine were developed and tested for on-line nonquantitative mass tagging of peptides and proteins. The nonquantitative aspect of the covalent tagging thus allows direct counting of free cysteines in the mass spectrum of a biomolecule through additional adduct peaks. Several substituted hydroquinones were investigated in terms of electrochemical properties, and their usefulness for on-line mass tagging during microspray experiments were assessed with L-cysteine, peptides, and intact proteins. Complementarily, numerical simulations were performed to properly understand the respective roles of mass transport, kinetics of electrochemical-chemical reactions, and design of the microspray emitter in the mass tagging overall efficiency. Finally, the on-line electrochemical tagging of cysteine residues was applied to the analysis of tryptic peptides of purified model proteins for protein identification through peptide mass fingerprinting.

Cysteine↗

PubMed2003

Microfluidic systems in proteomics.

We present the state-of-the-art in miniaturized sample preparation, immunoassays, one-dimensional and multidimensional analyte separations, and coupling of microdevices with electrospray ionization-mass spectrometry. Hyphenation of these different techniques and their relevance to proteomics will be discussed. In particular, we will show that analytical performances of microfluidic analytical systems are already close to fulfill the requirements for proteomics, and that miniaturization results at the same time in a dramatic increase in analysis throughput. Throughout this review, some examples of analytical operations that cannot be achieved without microfluidics will be emphasized. Finally, conditions for the spreading of microanalytical systems in routine proteomic labs will be discussed.

Electrophoresis↗