2. What is Mass Spectrometry? Mass spectrometry could be considered as an analytical technique that involves the study in the gas phase of ionized molecules with the aim of one or more of the following: Molecular weight determination Structural characterization Gas phase reactivity study Qualitative and quantitative analysis of components in a mixture. Mass spectrometry consists basically of weighing ions in the gas phase. The instrument used could be considered as a sophisticated balance which determines with high precision the masses of individual atoms and molecules. Depending on the samples chemical and physical properties, different ionization techniques can be used. One of the main factor in choosing which ionization technique to be used is thermolability. For samples that are not themolabile and relatively volatile, ionization such as Electron Impact and/or Chemical Ionization can be effectively used. For samples that are thermolabile such as peptides, proteins and other samples of biological interest, soft ionization techniques are to be considered. Among the most used soft ionization techniques are Electrospray (ESI) and Matrix Assisted Laser Desorption (MALDI). The name given to a particular mass spec technique is usually pointing to the ionization method being used.

3. Ion source Mass analyzerDetector (EB=Electrostatic- Magnetic; ion trap; Q=Quadrupole; TOF=Time of Flight.) (EI=Electron Impact; CI=Chemical Ionization; SIMS=Secondary Ions Mass Spec; FAB=Fast Atom Bombardment; LDMS=Laser Desorption Mass Spec; PDMS=Plasma Desorption Mass Spec; TS=Thermospray; AS=Aerospray; ESMS=Electrospray Mass Spec). Mass Spectrometry The general operation of a mass spectrometer is: create gas-phase ions separate the ions in space or time based on their mass-to-charge ratio measure the quantity of ions of each mass-to-charge ratio The ion separation power of a mass spectrometer is described by the resolution, which is defined as: R = m / delta m, where m is the ion mass and delta m is the difference in mass between two resolvable peaks in a mass spectrum. E.g., a mass spectrometer with a resolution of 1000 can resolve an ion with a m/e of 100.0 from an ion with an m/e of 100.1.

8. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), first introduced in 1988 by Tanaka and independently by Hillenkamp and Karas, has become a widespread analytical tool for peptides, proteins and most other biomolecules (oligonucleotides, carbohydrates, natural products, and lipids). The efficient and directed energy transfer during a matrix-assisted laser-induced desorption event provides high ion yields of the intact analyte, and allows for the measurement of compounds with high accuracy and sub-picomole sensitivity. MALDI provides for the nondestructive vaporization and ionization of both large and small biomolecules. In MALDI analysis, the analyte is first co-crystallized with a large molar excess of a matrix compound, usually a UV-absorbing weak organic acid, after which pulse UV laser radiation of this analyte-matrix mixture results in the vaporization of the matrix which carries the analyte with it. The matrix therefore plays a key role by strongly absorbing the laser light energy and causing, indirectly, the analyte to vaporize. The matrix also serves as a proton donor and receptor, acting to ionize the analyte in both positive and negative ionization modes, respectively.

13. the kinetic energy of an ion leaving the ion source is: The ion velocity, v, is the length of the flight path, L, divided by the flight time, t: Time-of-Flight Mass Spectrometry (TOF-MS) A time-of-flight mass spectrometer uses the differences in transit time through a drift region to separate ions of different masses. It operates in a pulsed mode so ions must be produced or extracted in pulses. An electric field accelerates all ions into a field-free drift region with a kinetic energy of qV, where q is the ion charge and V is the applied voltage.

28. Protéine Peptides Séparation des peptides en fonction de leur rapport m/z Identification de la protéine Empreinte de masse m/z Intensité

31. Protein identification strategies Maldi-Tof MS - Small proteins Protein mixtures Unknown proteins