MALDI

Question 1

MALDI Acrynm

Answer 1

• matrix assisted laser desorption ionization
• soft ionization method useful for macromolecules of >200kDa

Question 2

What is MALDI

Answer 2

• soft ionization technique which leaves molecules intact
• can be used for liquid or solid substrates
• can be interfaced with liquid chromotography and electrophoresis

Question 3

MALDI Mechanism

Answer 3

• sample is mixed with a matrix and allowed to dry which results its crystal formation
• laser flash ionizes the matrix molecules which in turn transfer protons to the analyte molecules

Question 4

MALDI Specifications

Answer 4

• pulsed laser of wavelength 193-430nm
• pulse width 0.5-10ns
• energy per pulse 1X10^-7microjoules
• mean power of pulse 10²W-5X10³
• diameter of pulse 10-200micrometers
• energy per laser pulse and unit area (fluence) 3X10^-5 to 60X10^-6 microjoules per meter squared
• laser penetration depth 50-200nm

Question 5

MALDI Matricies

Answer 5

• absorb a large amount of energy from the laser thus increasing its temperature by 100-600C
• the matrix then relays the energy to the sample in controlled amounts allowing the analyte macromolecules to desorb without decomposition
• matrix also provides a source of protons for the analyte

Question 6

MALDI Matrix Solution

Answer 6

• the mixing and the solvent must be compatable with the analyte so that uniform distribution occurs
• a plate needs to be spotted for analysis but it can be automated with a fraction collector
• 1:10,000 ratio of analyte to matrix molecules so that the analyte imolecules are completely isolated from one another
• coupling of the matrix with the laser causes rapid vibrational excitation and localized disintigration
• this causes clusters of matrix and analyte to be ejected from the surface thereby leaving the analyte in the gas phase and the matrix as violatile ions

Question 7

MALDI Analyte Ionization

Answer 7

• photoexcited matrix molecules transfer protons via a secondary ion-molecule charge-transfer reaction
• since catiions can be produced desalting is an important step
• peptides and proteins typically have higher proton affinities than standard matrix materials

Question 8

MALDI Spectrum

Answer 8

• charge one molecular ion
• high background at (m/z)<1000
• suitable for high throughput screening

Question 9

Ambient Ionization

Answer 9

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