The overall goal of the following experiment is to map the spatial distribution of chemicals in a biological tissue using an ambient mass spectrometry technique. This is achieved by using the pneumatically assisted spray of DESI to disor and ionize the molecules present in the tissue. As a second step, the DESI ion source is scanned across the sample area in a controlled fashion.
Next, the mass spectral information is correlated to the spatial motion of the sample stage in order to plot the ion's intensity as a function of X and Y position. Ultimately creating a chemical image. The results show that there are distinct distributions of lipids in wrapped brain tissues between the gray and white matter based on the mass spectrometry images of selected lipid ions.
Though this method can provide insights into the chemical distributions in biological tissues. It can also be used for a variety of other systems, including thin layer chromatography plates, mineral surface reactions, and marine algae. Prior to starting this procedure, mount the tissue on a microscope glass slide using ACEDONITE trial with 1%acetic acid as the DESI solvent turn on the syringe pump with a flow rate of five microliters per minute.
Once the solvent appears dripping out of the DESI source tip, turn on the nitrogen nebulizing gas with a pressure of 160 PS.I then turn on the high voltage power supply connected to the ion source, applying 3, 600 volts for optimal transfer of ions. Adjust the stage height so that the MS interface capillary hovers over the sample surface. The capillary should be less than one millimeter from the surface and have a collection angle of about 15 degrees.
Once the slide has been mounted on the motion stage, position the tip three millimeters from the sample surface and five millimeters from the MS capillary inlet with the probe at an angle of 55 degrees with respect to the sample surface. Next, align the DESI probe in the X dimension with respect to the MS capillary inlet so that they are directly in line. Using the extra tissue section.
Set the Y separation between the source tip and the capillary inlet to about four millimeters and the Z separation between the tip and the sample surface to about 1.5 millimeters. Following this, adjust the distance that the inner capillary extrudes from the outer capillary of the source for maximum sensitivity and minimal impact spot size. The glare of light in the room on the microscope slide can be used to better see the impact spot to improve the sensitivity during the ionization process.
Heat the transfer capillary using a rope heater or heating block to 100 degrees Celsius program. The lab view VI control software for desired imaging conditions based on DESI impact plume size and optimal sensitivity. Use a stage scan speed of 160 micrometers per second and align spacing between rows of 200 micrometers.
Give the directory path and the file name for the position and the time files to be recorded during images within the lab view vi. In preparation for the mass spectral data acquisition, the software automatically calculates the total time required for imaging input the total time in minutes into the mass spectrometer prior to starting the data acquisition. Next position the spray impact spot in the top left of the area to be imaged.
Begin the acquisition of the MS data and the stage motion simultaneously. Upon conclusion of the data acquisition, return the mass spectrometer to standby mode, turn off the high voltage of the DESI source, turn off the nitrogen gas and the syringe pump using the data manager within mass center. Convert the acquired data to TED Data and then export it in CDF format.
Finally, upload the raw CDF mass spectral data and the two text files for position and time to the Omnis SPECT website, or use bio map to visualize data processed in Firefly. Shown here is a representative spectrum obtained in the positive mode from an untreated rat brain section. In the positive mode, the mass spectrum is dominated by phosphatidylcholine due to their high iron and ionization efficiencies, which are attributed to the positively charged quaternary ammonium group.
In addition, the total ion image of the tissue section shows an abundant signal across the entire brain section. The spatial distribution of example lipids shows how the relative abundance of different phosphatidylcholine species varies between the gray and white matter of the brain. For example, the species potass phosphatidylcholine 34 to one with a mass charge ratio of 7 98 0.5364 shows increased intensity in the cerebellar cortex or gray matter.
In contrast, the species potass phosphatidylcholine 36 to one with a masto charge ratio of 8 26 0.5558 shows increased intensity in the cerebellar peduncle or white matter. The composite image obtained for the two ions highlights the contrast in lipid distribution across the tissue section. When performing this type of experiment, it's important to carefully optimize the DSI source sample stage and inlet geometry.
These variables are critical for successful dsi imaging.
