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Colloquia with Prof. Guido Verbeck, University of North Texas

"New Development in Mass Spectrometry: From Single Cell Analysis to Preparative Devices"

Nanomanipulation-Coupled to Mass Spectrometry
The advent of small volume analysis with mass spectrometry has opened
the door to investigation of micron and submicron analytes, and bringing the sampling
directly to the interest. The nanomanipulator is a multistage bioworkstation
consisting of a four-positioner system that has been directly and indirectly coupled with
nanospray mass spectrometry. This coupling allows for new significant applications
developments in the areas of single organelle analysis and
combinatorial library sequencing. The option to employ multiple end-effectors simultaneously
is ideal for cellular probing. Single organelle analysis has been demonstrated with
cotton seed samples. Single lipid bodies were stained with BODIPY and extracted from
within the cells into a suitable spray solvent. Analysis of an extracted lipid body
was subsequently completed by nanospray mass spectrometry, and the results
showed the distribution of triglycerides present. Recent work has also proven the
nanomanipulator to be a rapid and efficient tool for one-bead-one-compound,
also known as split-mix, combinatorial peptide library analysis. In this
experiment, a peptide sequence coated on a 130 um resin bead was extracted using a
suitable spray solvent. The resulting extract was then collected and analyzed, and
tandem mass spectral data allowed the elucidation of the peptide sequences.
The translational resolution of the nanomanipulator, which is beyond the optical
limit in both coarse and fine modes, allows for fine precision and control of the tools
required for probing in these types of experiments. We also highlight the suitability
of LPME –MS (Liquid Phase Microextraction-Mass Spectrometry) by extracting
and analyzing molecules from three different types of samples—1) visualization of an
extracted fluorescent dye, 2) non-polar polymer additives in vertebrate serum,
and 3) controlled acid-catalyzed hydrolysis of triacylglycerols within industrial refined
cottonseed oil demonstrating the ability to perform chemical reactions within
nanospray capillaries. Coupling of the nanomanipulator to nanospray mass spectrometry
has been extremely beneficial due to the small (300 nL) volume and sample mass (300 attograms) required.

All of these developments are leading to a tool that would allow the
researcher to perform cradle-to-grave analysis without leaving the microscope.
This greatly reduces prep and gives the researcher better control over the experiment.
We have started to use this instrument to tackle puzzling questions about metabolic disease
and cancer cell heterogeneity.

Preparative Mass Spectrometry
The soft landing of ionized nanoclusters, metal-ligand complexes,
polymers, and biopolymers on a hard surface is a challenging task, as most soft landed
molecules either contain too much kinetic energy, usually in the range of 0.1 - 50 KeV.
This amount of energy is not acceptable for the landing of nanoclusters as the
cluster would fragment upon impact. The design and fabrication of a novel
soft-landing system using front-end chemistry and ion optics at pressures spanning
from 1 – 100 torr rather than usual UHV conditions, and an actual softer land of
around 0.01-1.0 eV, would deposit the clusters intact on a surface allowing further
characterization of deposited materials on surfaces.

Two instruments have been developed to soft-land species with less than
1eV. First is the laser ablation-coupled drift tube. The chamber was done with the
idea of using an inert gas, with helium as the buffer gas. This allows us to reduce the
clusters KE from 40 eV to about 1 eV by producing thermal collisions between the gas
and our sample. The clusters formed were then discriminated by cross-section and
transported through the drift cell with a potential from -100 to -500 V to
our collecting surface. The isolated sample was then removed for characterization
using Raman spectroscopy and electrochemistry. The second instrument is a rectilinear
ion trap with integrated ultra fast pulse valve. This instrument has some novel
electronics to allow complete shut-down of the applied RF in 2 cycles. This coupled to
the high pressure acquisition makes it an ideal mass filter for preparative mass
spectrometry. Though the ion current is an order of magnitude less than the drift tube, the
resolution is 2 orders of magnitude greater, making the two instruments complimentary.
Both instruments will be shown to be the work horses of our novel preparative combing method.

Mon Apr 15, 2013 4:15pm – 5:30pm Central Time
5211SC (map)