MALDI-TOF (Matrix Assisted Laser Desorption Ionization Time Of Flight

TOPIC: MALDI-TOF(Matrix Assisted
Laser Desorption/Ionization Time
Of Flight)

INTRODUCTION:
 Proteome - complete set of proteins encoded in a genome.
Protein identification requires their separation by methods:
1. IEF
2. SDS- gel electrophoresis
3. Mass spectroscopy (MS)
MASS SPECTROSCOPY:
 Separation of charged molecules/ molecules according to
their mass to charge.
 Determines relative molecular mass (M).
 High resolution, precision, & sensitivity.
 Requires charged gaseous molecules for its analysis.

COMPONENTS OF MASS SPECTROSCOPY:
Ionization source:
1. Matrix-assisted laser desorption/ ionization - time of
flight(MALDI).
2. Electrospray (ES) Analyser m/z: TOF, Magnet.

HISTORY OF MALDI-TOF:
The history of MALDI-TOF mass spectrometry began in the 1960s
with the development of laser desorption techniques.
However, it wasn’t until 1985 that Franz Hillenkamp, Michael
Karas, and their colleagues coined the term “matrix-assisted laser
desorption ionization” (MALDI).
The breakthrough came in 1987 when Koichi Tanaka used a
combination of ultraviolet laser and “ultra-fine metal plus liquid
matrix method” to ionize biomolecules as large as the 34,472 Da
protein carboxypeptidase-A.
 This innovation led to the development of MALDI-TOF
instruments, which have since become a crucial tool in various
fields, including proteomics, genomics, and metabolomics.

MALDI-TOF:
Introduction:
 MALDI is the abbreviation for "Matrix Assisted Laser
Desorption/lonization.“
 MALDI is appropriate to analyze biomolecules like
peptides, lipids, saccharides, or other organic
macromolecules.
 TOF stands for "Time of Flight". This generates
characteristic mass spectral fingerprints which is
compared with large library of mass spectra.
 A versatile analytical technique to detect and
characterize mixtures of organic molecules.

MALDI-TOF (Matrix Assisted Laser Desorption Ionization Time Of Flight

PRINCIPLE:
 Matrix absorbs the ultraviolet light (nitrogen laser light,
wavelength 337 nm) and converts it to heat energy.
 The analyte is embedded on the Target Plate.
 After a laser pulse, the irradiated spot is rapidly heated
and becomes vibrationally excited.
 Matrix molecules energetically ablated from the surface
of the sample.
 During ablation, the analyte molecules are usually
ionized by being protonated or deprotonated with the
nearby matrix molecules.

PARTS OF MALDI TOF1:
1) Sample plate
2) Matrix
3) Laser
4) Variable voltage grid
5) Vacuum system
6) Flight tube
7) Time mass detectors
8) Camera
9) Clock

Matrix:
Matrix consists of crystallised molecules of which the three
most commonly used are
 3,5 dimethoxy-4-hydroxy cinnamic acid (sinapinic acid)
 a-cyano-4-hydroxycinnamic acid (alpha-cyano or
alpha-matrix)
 2,5 dihydroxy benzoic acid (DHB)

Laser:
 MALDI techniques typically employ the use of UV lasers
such as nitrogen lasers (337 nm) and frequency-tripled
and quadrupled Nd:YAG lasers (355 nm and 266 nm
respectively).
 Although not as common, infrared lasers are used due
to their softer mode of ionization.
 IR-MALDI also has the advantage of greater material
removal (useful for biological samples), less low- mass
interferences, and compatibility with other matrix-free
laser desorption mass spectrometry methods.

TOF ANALYSER:
 The mass analyser used for MALDI is TOF analyser.
 It allows accurate mass determination for high
molecular weight species.
 It makes TOF ideal for biomolecules.
 The ions desorbed by the laser pulse for accelerated in
an electric field to a kinetic energy of several KeV.

Flight tube length (2m) and flight time
(nanoseconds)
At the end of the tube, ions hit the detector.
And drift time is measured electronically.
Lighter ions are accelerated more than the heavier ions and reach
the detector first. TOF analyser.
Kinetic energy of the drifting ions is defined as Ekin =1/2 .m.v² = z. E.
V
The velocity can be defined as V= L/t Substituting the velocity in eq.
 Ekin= (1/2)m(L/t)² = zeV

HOW DOES MALDI-TOF WORKS?
 Step 1: Sample Preparation:
• The sample is mixed with a matrix material, typically a crystalline substance.
• The matrix helps to absorb the laser energy and transfer it to the sample
molecules.
• The sample-matrix mixture is then applied to a target plate.
 Step 2: Laser Desorption/Ionization:
• A high-powered laser is fired at the sample-matrix mixture on the target
plate.
• The laser energy is absorbed by the matrix, causing it to vaporize and
create a plasma.
• The plasma contains ionized sample molecules, which are then ejected into
the gas phase.

 Step 3: Ion Separation and Detection:
• The ionized sample molecules are then separated according to
their mass-to-charge ratio using a time-of-flight (TOF) analyzer.
• The TOF analyzer measures the time it takes for each ion to
travel through a field-free region.
• The ions are then detected by a detector, which records the
arrival time and intensity of each ion.
• The resulting mass spectrum is then analyzed to identify the
sample molecules.

Applications of Maldi:
 Used for the analysis of proteins and peptides changes
in structure can also be identified.
 Vey low amount of sample sample preparation is easy
and spectra is simple.
 Very fast separation method.

ADVANTAGES OF MALDI-TOF:
 High sensitivity and specificity.
 Fast analysis time.
 Low sample requirement.
 Ability to analyze complex mixtures.
LIMITATION OF MALDI-TOF:
 Limited dynamic range
 Matrix effects
 Ion suppression
 Instrumentation cost and maintenance

What's its future?
o Will help revolutionize the medical world and will help
lead to treatments for many diseases.
o Will be useful for DNA sequencing, thus can be useful for
forensic investigations.

MALDI-TOF (Matrix Assisted Laser Desorption Ionization Time Of Flight

More Related Content

What's hot (20)

Similar to MALDI-TOF (Matrix Assisted Laser Desorption Ionization Time Of Flight (20)

Recently uploaded (20)

MALDI-TOF (Matrix Assisted Laser Desorption Ionization Time Of Flight