Abstract
Since the birth of proteomics science in the 1990, the number of applications and of sample preparation methods has grown exponentially, making a huge contribution to the knowledge in life science disciplines. Continuous improvements in the sample treatment strategies unlock and reveal the fine details of disease mechanisms, drug potency, and toxicity as well as enable new disciplines to be investigated such as forensic science.This chapter will cover the most recent developments in sample preparation strategies for tissue proteomics in three areas, namely, cancer, toxicology, and forensics, thus also demonstrating breath of application within the domain of health and well-being, pharmaceuticals, and secure societies.In particular, in the area of cancer (human tumor biomarkers), the most efficient and multi-informative proteomic strategies will be covered in relation to the subsequent application of matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) and liquid extraction surface analysis (LESA), due to their ability to provide molecular localization of tumor biomarkers albeit with different spatial resolution.With respect to toxicology, methodologies applied in toxicoproteomics will be illustrated with examples from its use in two important areas: the study of drug-induced liver injury (DILI) and studies of effects of chemical and environmental insults on skin, i.e., the effects of irritants, sensitizers, and ionizing radiation. Within this chapter, mainly tissue proteomics sample preparation methods for LC-MS/MS analysis will be discussed as (i) the use of LC-MS/MS is majorly represented in the research efforts of the bioanalytical community in this area and (ii) LC-MS/MS still is the gold standard for quantification studies.Finally, the use of proteomics will also be discussed in forensic science with respect to the information that can be recovered from blood and fingerprint evidence which are commonly encountered at the scene of the crime. The application of proteomic strategies for the analysis of blood and fingerprints is novel and proteomic preparation methods will be reported in relation to the subsequent use of mass spectrometry without any hyphenation. While generally yielding more information, hyphenated methods are often more laborious and time-consuming; since forensic investigations need quick turnaround, without compromising validity of the information, the prospect to develop methods for the application of quick forensic mass spectrometry techniques such as MALDI-MS (in imaging or profiling mode) is of great interest.
Highlights
Since the birth of proteomics science in the '90, the number of applications and of sample preparation methods has grown exponentially, making a huge contribution to the knowledge in lifescience disciplines
The methodologies and considerations are applicable to the techniques of: Matrix Assisted Laser Desorption Ionisation Mass Spectrometry Profiling and Imaging (MALDI-MSP and MALDI-Mass spectrometry imaging (MSI) respectively) and Liquid Extraction Surface Analysis (LESA)
The groups reported various differences within the proteomic signatures observed from this multicentre study with four proteins identified by the Munich group to be significantly associated with the tumour stromal region and three of these signals were evident in the Leiden data set
Summary
The preparation and pre-treatment of tissues for the study of proteomics in cancer is both unique and challenging considering the information desired by the clinician or research scientist. A two centre study performed in 2014 held in Leiden, Netherlands and in Munich, Germany did one such comparison Both groups embarked on a comparative set of experiments and produced the output entitled; Multicenter Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Imaging (MALDI MSI) Identifies Proteomic Differences in Breast-Cancer-Associated Stroma [6]. Formalin Fixed Paraffin Embedded - Preparing FFPE tissues for mass spectrometry profiling and/or imaging presents many challenges, some of which include; detachment of tissue from the microscope slide, interference from remaining paraffin wax, delocalisation and loss of target species of interest These factors affect reproducibility and increase the requirement for biological replicates in addition to technical repeats. On a larger scale, the tissue treatments before mass spectrometry experimental work are comparable only differing by the trypsin and matrix deposition instrumentation being the Image Prep (Bruker Daltonik, GmbH, Bremen, Germany) in this study
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