Imaging of thin-layer chromatograms using matrix-assisted laser desorption/ionization mass spectrometry

Metastable decomposition of ions generated in matrix-assisted laser desorption/ionization (MALDI) mass spectrometers complicates analysis of biological samples that have labile bonds. Recently, several academic laboratories and manufacturers of commercial instruments have designed instruments that introduce a cooling gas into the ion source during the MALDI event and have shown that the resulting vibrational cooling stabilizes these labile bonds. In this study, we compared stabilization and detection of desorbed gangliosides on a commercial orthogonal time-of-flight (oTOF) instrument with results we reported previously that had been obtained on a home-built Fourier transform mass spectrometer. Decoupling of the desorption/ ionization from the detection steps resulted in an opportunity for desorbing thin-layer chromatography (TLC)-separated gangliosides directly from a TLC plate without compromising mass spectral accuracy and resolution of the ganglioside analysis, thus coupling TLC and oTOF mass spectrometry. The application of a declustering potential allowed control of the matrix cluster and matrix adduct formation, and, thus, enhanced the detection of the gangliosides. during matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) analyses of molecules having labile bonds. Although this phenomenon can be used for obtaining analyte structural information for small peptides, oligosaccharides, and some glycolipids [1-5] for larger or more fragile molecules, it can limit the mass accuracy and resolution and complicate interpretation of the spectra of heterogeneous samples. Thus, it is important to develop methods that allow control of the extent of metastable decay. In addition to the pressure of the background gas and extraction field strength in the MALDI technique, the major factors affecting metastable fragmentation are laser wavelength/pulse width and the choice of the MALDI matrix [6,7]. In addition, the presence of electrons may play a role in analyte fragmentation and may cause suppression of multiply charged ions in a spectrum [8,9].Control of the metastable fragmentation in MALDI by increasing the pressure in the ionization source and reduction of the ions internal energy was first introduced by Loboda et al. on orthogonal time-of-flight (oTOF) instruments [10] and later extended to Fourier transform mass spectrometry (FTMS) [11] and to the operation at atmospheric pressure on various types of instruments [12]. These methods have since been used for stabilization of glycolipids [13,14] and peptides and proteins [11,15]. The term vibrational cooling (VC) MALDI recently has been introduced to describe desorption in the pressure range where cooling of the excess bond energy is achieved [12]. Stabilization of the labile bonds under these conditions allows for control of the extent of fragmentation, which is particularly

mentioning

confidence: 98%

Ganglioside analysis by thin-layer chromatography matrix-assisted laser desorption/ionization orthogonal time-of-flight mass spectrometry

Ivleva

Sapp

O’Connor

et al. 2005

“…In the analysis of complex samples, such as biological tissue, MALDI is of particular interest because of its ability to desorb and ionize molecules of high molecular weight, such as proteins and peptides, providing excellent sensitivity while retaining considerable tolerance towards salts and other small molecules found at high concentration in tissue. It has been roughly 10 years since the first published applications in which MALDI-MS was used to create a chemical images of substrates [4,5]. The intervening decade has seen a considerable growth in techniques and instrumentation for MALDI-MS imaging, developed largely by Caprioli and coworkers [6 -12] with contributions to sampling techniques [13][14][15][16] and instrumentation [17,18] provided by others.…”

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confidence: 99%

MALDI-MS imaging of features smaller than the size of the laser beam

Jurchen

Rubakhin

Sweedler

2005

265

223

The feasibility of matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) imaging of features smaller than the laser beam size has been demonstrated. The method involves the complete ablation of the MALDI matrix coating the sample at each sample position and moving the sample target a distance less than the diameter of the laser beam before repeating the process. In the limit of complete sample ablation, acquiring signal from adjacent positions spaced by distances smaller than the sample probe enhances image resolution as the measured analyte signal only arises from the overlap of the laser beam size and the non-ablated sample surface. [3] has revolutionized the investigation of biological molecules by providing soft ionization methods linking biochemistry with the powerful analysis tools of mass spectrometry (MS). In the analysis of complex samples, such as biological tissue, MALDI is of particular interest because of its ability to desorb and ionize molecules of high molecular weight, such as proteins and peptides, providing excellent sensitivity while retaining considerable tolerance towards salts and other small molecules found at high concentration in tissue. It has been roughly 10 years since the first published applications in which MALDI-MS was used to create a chemical images of substrates [4,5]. The intervening decade has seen a considerable growth in techniques and instrumentation for MALDI-MS imaging, developed largely by Caprioli and coworkers [6 -12] with contributions to sampling techniques [13][14][15][16] and instrumentation [17,18] provided by others.It is generally accepted that the maximal spatial imaging or profiling resolution of microprobe imaging techniques is determined by a combination of the size of the microprobe and the precision of the sample or microprobe positioning device. MALDI mass spectrometers typically use lasers having relatively large beam sizes (about 100 m diameter) in the analysis of standard, dried-droplet preparations. Some effort has been put into decreasing the size of the laser beam sizes for MALDI-MS imaging and profiling of biological samples, particularly for samples containing a high proportion of peptidergic neurons or other secretory cells. Investigated biological samples of this nature include rat pituitary and rat pancreas [6], mouse brain and human brain tumor xenografts [8,14], rat brain and rat brain tumors [9,19], mouse epididymis [20], molluscan atrial gland [15], and molluscan peptidergic neurons [16].Ideally, the spatial resolution of MALDI-MS imaging of analyte-rich tissues would approach the size of a single mammalian cell, 5 to 20 m in diameter. Several strategies have been used or suggested to decrease the laser beam diameter in imaging applications, including the placement of a pinhole aperture between the outlet of the laser and the focusing optics of the mass spectrometer [6,13], decreasing the size of the fiber optic used to direct the laser into the MALDI source [9], and placing multiple lenses between the laser and the M...

“…Highly sensitive investigation of single cells, neurons, or tissue areas is a field of dramatically growing consideration [18 -24]. Due to instrumental and methodological limitations, MALDI imaging has so far been limited to a lateral resolution of approximately 30 m [18,[25][26][27][28][29][30], limiting its application to relatively large structures.…”

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confidence: 99%

Scanning microprobe matrix-assisted laser desorption ionization (SMALDI) mass spectrometry: Instrumentation for sub-micrometer resolved LDI and MALDI surface analysis

Spengler

Hubert

2002

270

256

A new instrument and method is described for laterally resolved mass spectrometric surface analysis. Fields of application are in both the life sciences and the material sciences. The instrument provides for imaging of the distribution of selected sample components from natural and artificial surfaces. Samples are either analyzed by laser desorption ionization (LDI) time-of-flight mass spectrometry or, after preparation with a suitable matrix, by matrixassisted laser desorption ionization (MALDI) mass spectrometry. Areas of 100 ϫ 100 m are scanned with minimal increments of 0.25 m, and between 10,000 and 160,000 mass spectra are acquired per image within 3 to 50 min (scan rate up to 50 pixels per s). The effective lateral resolution is in the range of 0.6 to 1.5 m depending on sample properties, preparation methods and laser wavelength. Optical investigation of the same sample area by UV confocal scanning laser microscopy was found to be very attractive in combination with scanning MALDI mass analysis because pixel-identical images can be created with both techniques providing for a strong increase in analytical information. This article describes the method and instrumentation, including first applicational examples in elemental analysis, imaging of pine tree roots, and investigation of MALDI sample morphology in biomolecular analysis. (J Am Soc Mass Spectrom 2002, 13, 735-748)