To begin, place the slide containing the matrix deposited seed slices in the mass spectrometer. Load the sample image, and use correction pen marks to set teaching points on the referenced software. Then set the 99%data reduction factor, save the FID file for the posterior data calibration, and save the method.
Delimitate the area to be analyzed using the add polygon measurement region tool from the mass spectrometer software. Set the raster width to 100 micrometers. Edit the measurement region parameters indicating the saved method and save the imaging run.
Then start the imaging acquisition. Use matrix cluster and known contaminants to create a mass list in the software in the calibrant tab. Then in the calibration tab, open the created mass list.
Right click to open a dialogue box and choose the set lock masses option. Select Gaussian window mode with 0.5 Gaussian broadening and 3.5 line broadening. Leave online calibration unchecked.
Set mode to single, threshold to 1000, and mass tolerance to five PPM. Calibrate the data with the process and save a 2D serial dataset tool. After calibration, open the dot MIS file to a compatible software and alter the normalization from no norm to RMS or TIC.
Click on the edit tab and select automatic mass filtering. Fill start mass and end mass with the minimum and maximum number of Dalton at your interest threshold, and click on the okay icon. Select the highlighted peak of interest at the filtered list, generated with the number corresponding to the mass value of interest in Dalton.
Change the percentage to better view the interested area. Click on the intensity scale bar and the color map icons. Click on the image area and drag slide the mouse to position the area of interest.
Change the transparency percentage to produce a merged signal image with the scan section image as the background. If the analytes to be mapped are known, plot each M by Z value for each analyte and save the generated images and the spectral average plot. The mass spectrum of Euterpe precatoria and Euterpe edulis seeds tissue obtained by MALDI IMS in positive mode is shown here.
This MALDI IMS analysis of E.precatoria seeds exhibited peaks representing adducts of hexoses oligomers without adding salt to the matrix. Hexoses dimers, trimers, tetramers, pentamers, hexamers, and up to 14 unit oligomers were identified. The same results were also obtained for E.edulis seed tissue.
The box plots for both samples indicate the peak intensity of each hexose oligomer, found in the seed endosperm, demonstrating their distributions, and a slightly higher content of a high degree of polymerization of oligomers.
