**2.1. Sample preparation**

**1. Introduction**

trometer work independently from each other [8].

ablated material even if the concentration remains constant [5].

**-** sum of different selected isotopes ("extracted ion current", EIC),

**-** pixel per pixel normalization versus overall factor normalization,

**-** different homogeneously distributed isotopes as reference,

**-** masking of background data.

In this chapter, different normalization methods were tested and compared:

**-** sum of all measured isotopes (more or less reflecting "total ion current", TIC),

The final generation of element distribution maps from the different normalization methods was done with the aid of Excel Laser Ablation Imaging (ELAI) representing a software tool that is based on Microsoft Excel 2007 Visual Basic for Applications. This software is freely

Laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) is today a wellestablished technique for direct analysis of solid samples [1, 2]. It allows the uncomplicated elemental analysis of many different kinds of sample materials with inexpensive preparation methods. It can be used for the precise determination of major, minor, and trace element distributions or isotope ratios and is therefore already applied in many research fields [3, 4]. The basic principle of LA-ICP-MS is the multielement analysis of a mostly solid sample. The sample is placed in an ablation chamber purged with a constant argon gas flow and ablated line-by-line with a laser beam focused on the sample surface. The amount of ablated material and thus the composition of the gas phase are highly depending on the sample and laser properties [5, 6]. The aerosol consisting of the ablated material and argon is transported to the plasma torch for evaporation, atomization, and ionization [7]. After refocusing the gas stream in the sampler/skimmer interface, the generated ions are separated in the quadrupole mass spectrometer according to their mass-to-charge (*m/z*) ratio. The acquired signal-ion intensities are then measured and recorded as a continuous list of data points without any trigger signal indicating the line shift. This is necessary because the laser ablation unit and the mass spec-

400 Applications of Laser Ablation - Thin Film Deposition, Nanomaterial Synthesis and Surface Modification

In LA-ICP-MS measurements, the ion intensities depend on many parameters. Since it is not possible to maintain perfectly constant conditions per sample and throughout a measurement run, the obtained data have to be normalized allowing the comparison of absolute element concentrations [9, 10]. There are several ways for normalization that depend on the sample material and the experimental setup. Common normalization methods [10] are either based on one isotope, the total ion current (TIC), or on selected isotopes that should represent the ion current attributable to the sample matrix termed extracted ion current (EIC). The use of internal standards as reference is desirable, but challenging with respect to the choice of a matching material and to its application or introduction [10]. Normalization can either be performed using a separate reference for each data point or a normalization factor constant throughout a measurement run or sample. In addition, for the calibration of the laser ablation process, matrix-matched standards are essential to compensate changes in the amount of Mice organs (brain and liver) were cryo-cut into 30 μm slices with a standard research cryostat (CM3050 S, Leica Biosystems, Wetzlar, Germany) at −18°C box temperature and −16°C object temperature. In brief, the frozen organ was fixed on a sample holder with Tissue-Tek® O.C.T. compound (Sakura Finetek Europe B.V., The Netherlands). Slices were thaw-mounted onto StarFrost® adhesive microscope slides (Waldemar Knittel Glasbearbeitungs GmbH, Braunschweig, Germany), dried, and stored at room temperature until used for LA-ICP-MS measurement.
