1. MALDI-TOF
By Miss Adnya A. Desai
Dept. of Microbiology

2. MATRIX ASSISTED LASER DESORPTION
IONIZATION-TIME OF FLIGHT
• Maldi is soft ionization technique used in mass
spectrometry, allowing the analysis of
biomolecules such as DNA, proteins, peptides &
sugar or polymers such as dendrimers and
macromolecules
• It is three steps method.
I. The sample is mixed with a suitable matrix &
applied to a metal plate.
II. A pulsed laser irradiate a sample triggering
desorption of matrix material.
III. Ionization of analyte molecules

3. HISTORY OF MS
• MALDI- Term was coined in 1985 by Franz
Hillenkamp, Michael Karas
• They found that amino acid alanine could be
ionized easily if it was mixed with amino acid
tryptophan & irradiated with pulsed 266nm
laser.
• Here, tryptophan absorbed the laser energy &
helped to ionize the non absorbing alanine.

4. • To absorbs biomolecules more clearly Kochi
Tanaka & his co-workers used ultra fine metal
plus liquid matrix method that combined 30nm
cobalt particles in glycerol with 337nm N2
laser for ionization.
• Using this laser matrix combination Tanaka was
able to ionize biomolecule as large as 34,472 Da
protein Carboxypeptidase-A. For this invention he
received one quarter of the 2002 Noble Prize in
Chemistry.

5. MASS ANALYZER
High Vacuum System
Inlet Detector
Data
System
Mass
Analyzer
Ion
Source
Time of flight
(TOF)
Quadraupole
Ion Trap
Magnetic Sector
FTMS
Turbo
molecular
pumps
HPLC
Flow
injection
Sample plate
Microchannel
Plate
Electron
Multiplier
Hybrid with
photomultiplier
PC
Sun SPARK
Station
DEC Station

6. Essential Features of all MS
• Production of ions in the gas phase
• Acceleration of the ions to a specific velocity
in an electric field
• Separation of ions in a mass analyzer
• Detection of each species of particular m/z
ratio

7. MALDI TOF
Camera
Laser
Sample
plate
Timed ion
selector Reflector
Pumping Pumping
Linear
detector
Extraction
grids
Reflector
detector

8. Matrix
• Matrix consists of crystallized molecule of which the
most 3 commonly used are 3,5dimthoxy-4-hydroxy
cinnamic acid (sinapinic acid), alpha-cyano-4
hydroxycinnamic acid (CHCA/alpha-cyano/alpha-matrix),
2,5-dihydroxybenzoic acid (DHB).
• A solution of one of these molecules is made, often
in a mixture of highly purified water & an organic
solvent ACN i. e. acetonitril.
• Trifluoroacetic acid may also be added .
• A good example of matrix solution would be 20ug/ml
sinapinic acid in ACN:water:TFA (50:50:0.1).

9. Structure of Matrix

10. Laser
 Numerous gas and solid state lasers have been developed
for use in MALDI.
 Most MALDI devices use a pulsed UV laser
 N2 source at 337 nm
 Neodymium-yttrium aluminum garnet (Nd:YAG)
 emits at 355 nm and gives a longer pulse time
 IR lasers are also used
 The most common IR laser is the erbium doped-yttrium
aluminum garnet (Er:YAG)
 Emits at 2.94 micrometer
 it is “softer” than the UV, which is useful for certain biomolecules
 matrices available for IR absorption are limited

11. Ionization Mechanism
• The mechanism remains
uncertain
• It may involve absorption of light
by the matrix
• Transfer of this energy to the
analyte
– which then ionizes into the
gas phase as a result of the
relatively large amount of
energy absorbed.
– To accelerate the resulting
ions into a flight-tube in the
mass spectrometer they are
subjected to a high electrical
field [2].

13. Time Mass Detectors
• The typical detector used
with MALDI is the time of
flight mass detector
(TO-FMS)
• TOF is a method where the
ions are accelerated by an
electric field, resulting in
ions of the same strength to
have the same kinetic
energy .
• Time it takes for each ion to
traverse the flight tube and
arrive at the detector is
based on its mass-to-charge
ratio; therefore the heavier
ions have shorter arrival
times compared to lighter
ions

14. detector
Time-of-flight (TOF) Mass Analyzer
Source Drift region (flight tube)
• Ions are formed in pulses.
• The drift region is field free.
• Measures the time for ions to reach the detector.
• Small ions reach the detector before large ones.

15. MALDI Advantages
• Gentle Ionization technique
• High molecular weight analyte can be ionized
• Molecule need not be volatile
• Sub-picomole sensitivity easy to obtain
• Wide array of matrices
see reference 8

16. TOF Advantages
• All ions detected at once
• High mass accuracy and resolving power
possible
• Reasonable performance for cost
– <5 ppm mass accuracy and >20,000 resolving
power commercially available
• High mass, low charge ions not a problem
– Theoretically unlimited mass range
Reference 8

17. APPLICATIONS
Chemistry
Pharmaceutical
analysis
Clinical
analysis
MICROBIOLOGY
Taxonomy

18. APPLICATIONS
Proteomics Pharmaceutical Analysis
• To identify, verify, and
quantitate: metabolites,
recombinant proteins,
proteins isolated from
natural source, peptides &
their amino acid sequences
• Bioavailability studies
• Drug metabolism
studies,
pharmacokinetics
• Characterization of
potential drugs
• Drug degradation
product analysis
• Screening of drug
candidates
• Identifying drug targets

19. APPLICATIONS
Microbiology
• It is used for the
identification of
microorganisms.
• Species diagnosis by
this procedure is much
faster, more accurate &
cheaper than other
procedures based on
biochemical tests.
Forensic analysis
Environmental
analysis
• Pesticides on foods
• Soil and groundwater
contamination

20. Questions
• Who coined the term MALDI?
• Name the different types of lasers are used in
maldi-tof?

21. REFERENCES
• www.google.com
• www.slideshare.com