The Identification Of Bacteria Through Matrix Assisted Laser Desorption/Ionization

2021 words - 8 pages

Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry, or MALDI-MS, is considered a laser-based soft ionization method. 7 MALDI allows the direct analysis of large, non-volatile and thermally unstable compounds in ionic gaseous states. 2,3,7 MALDI is modified form of laser desorption (LD) ionization. The difference between MALDI and LD is the addition of the matrix.2 A matrix is chosen based off of its ability to isolate the analyte from itself and its adsorption properties. Sample preparation consists of a solution of matrix, analyte and solvent. Matrix concentration must be in excess to ensure the isolation of analyte molecules. Solvents are important to help prevent the aggregation of large macromolecules.7 The solvent is then removed from the solution which results in a co-crystalline formation of the analyte and matrix molecules. This formation is considered homogenous due to the even dispersion of analyte molecules throughout the excess matrix molecules. A laser is used to ionize the co-crystalline sample produced. The wavelength is based off of the vibration excitation properties of the matrix molecules.2 Most commonly a wavelength within the range of ultraviolent, 10-400nm, and infrared, 400-700nm, is used.7 The laser is either shot directly, linear mode, or indirectly, reflection mode using a quartz mirror, on the surface of the co-crystalline sample.3,4 The laser is pulsed multiple times ranging from every 300 ps to 3ns. Each time the energy from the laser, which is directly related to the laser pulsation time, is transferred to the matrix molecules.7 This causes a vibrational excitation of the matrix molecules causing them to partially vaporize.2,7 The vaporization causes the formation of matrix ions. These charges from the ions are passed to the analyte molecules, thus producing intact analyte ions in the gaseous state.2,3,5,7 This factor of indirect ionization allows the direct analysis of the types of molecules referenced above.

Experiments

One initial paper talked about the use of MALDI-MS and rapid identification of bacteria with using whole cells. The basic procedure was the mixture of vortexed bacterium with matrix solutions of either 4-hydroxy-α-cyanocinnamic acid (4CHCA) or 3,5-dimethoxy-4-hydroxycinnamic acid (sinapinic acid), in a 1:9µL ratio. The matrices used in this paper are specific for carbohydrate and peptide/protein making them appropriate for whole cell analysis. In this paper a variation of MALDI-MS was used called MALDI time-of-flight mass spectroscopy or MALDI-TOF MS. MALDI-TOF MS compares the time it takes ions to move across the flight path to the mass of the ions. The cells were vertexed with the matrix solution and exposed to 337nm or ultraviolent radiation from a nitrogen laser. A linear mode was used with a flight path of 1.25m. An external calibration was used to help avoid any ion suppression caused by the internal standards of proteins used in the experiment. The use of this...

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