1. Introduction
Tissue microarrays (TMAs) play an important role in translational and clinical studies [1]. By concentrating hundreds of small tissue cores typically of 0.6 mm in diameter onto a single paraffin block, dozens of biomarkers can be studied on a large number of patients while at the same time sparing costs, resources and tissues [2,3,4]. TMAs are used to study protein markers by immunohistochemistry and to investigate DNA aberrations, mRNA and miRNA expression using various in situ hybridization techniques [5,6,7]. Together with clinically annotated and appropriately powered sample cohorts, TMAs are a powerful tool not only for biomarker screening but also for prognostic and predictive modeling of disease outcome. In addition, TMAs allow that the same experimental conditions be applied to hundreds of samples simultaneously [8].
Recent advances in TMA technology rely on digital pathology and automated tissue microarraying. One approach is to use a slide scanner to digitally visualize stained tissue sections, which can then be annotated using a TMA tool of various sizes (Figure 1). These annotated slides are matched to their corresponding donor blocks, which are precisely punched out at the desired locations and transferred into recipient blocks, automatically. At our institute, this approach of combining modern TMA technology with histopathological expertise and biostatistics is referred to as next-generation TMA (ngTMA) [9].
Figure 1  (A) Digital annotation of a scanned slide after double-immunohistochemistry staining with pan-cytokeratin and caldesmon; (B) Hematoxylin and Eosin staining of a tissue microarray (TMA) constructed from annotated regions of histological interest.   Clearly, TMA instrumentation could also be advantageous for applications other than construction of TMAs. In fact, punches taken from donor blocks can be placed directly into tubes and therefore used for molecular analysis using PCR-based approaches.
However, the question of possible cross-contamination across tissue samples using the same punching device, especially one embedded in an automated tissue microarrayer, has frequently been posed but never addressed. This may be of particular concern if nucleic acids are amplified by PCR. Therefore, in this study, we determine the level of contamination transferred between samples after automated punching using a sensitive assay for mycobacteria and a pyrosequencing assay for KRAS mutation analysis.