Formalin Fixation as Tissue Preprocessing for Multimodal Optical Spectroscopy Using the Example of Human Brain Tumour Cross Sections

Mona Stefanakis,Jörg W Bartsch,Barbara Boldrini,Susan Noell,Axel Pagenstecher,Karsten Rebner,Marcos Tatagiba,Marc Brecht,Jens Schittenhelm,Sabrina Hakelberg,Miriam C Bassler,Edwin Ostertag,Rainer Ritz,Anita Lorenz,Christopher Nimsky,Alexandra Wagner,Feride Severcan

doi:10.1155/2021/5598309

Abstract

Characterization of brain tumours requires neuropathological expertise and is generally performed by histological evaluation and molecular analysis. One emerging technique to assist pathologists in future tumour diagnostics is multimodal optical spectroscopy. In the current clinical routine, tissue preprocessing with formalin is widely established and suitable for spectroscopic investigations since degradation processes impede the measurement of native tissue. However, formalin fixation results in alterations of the tissue chemistry and morphology for example by protein cross-linking. As optical spectroscopy is sensitive to these variations, we evaluate the effects of formalin fixation on multimodal brain tumour data in this proof-of-concept study. Nonfixed and formalin-fixed cross sections of different common human brain tumours were subjected to analysis of chemical variations using ultraviolet and Fourier-transform infrared microspectroscopy. Morphological changes were assessed by elastic light scattering microspectroscopy in the visible wavelength range. Data were analysed with multivariate data analysis and compared with histopathology. Tissue type classifications deduced by optical spectroscopy are highly comparable and independent from the preparation and the fixation protocol. However, formalin fixation leads to slightly better classification models due to improved stability of the tissue. As a consequence, spectroscopic methods represent an appropriate additional contrast for chemical and morphological information in neuropathological diagnosis and should be investigated to a greater extent. Furthermore, they can be included in the clinical workflow even after formalin fixation.

Highlights

Characterization of brain tumours requires neuropathological expertise and is generally performed by histological evaluation and molecular analysis
As optical spectroscopy is sensitive to these variations, we evaluate the effects of formalin fixation on multimodal brain tumour data in this proof-of-concept study
In order to validate the diagnosis of the spectroscopically investigated tissue samples, a routine neuropathological diagnosis of the tissue sections was performed by two independent certified neuropathologists (AP and JS). e sample set I consisted of a World Health Organization (WHO)-grade I fibrous meningioma, a WHO grade II oligodendroglioma, a WHO grade III anaplastic ependymoma, and a WHO grade IV glioblastoma (Figures 1(a)–1(d))

Summary

Introduction

Characterization of brain tumours requires neuropathological expertise and is generally performed by histological evaluation and molecular analysis. Spectroscopic methods represent an appropriate additional contrast for chemical and morphological information in neuropathological diagnosis and should be investigated to a greater extent. They can be included in the clinical workflow even after formalin fixation. The process chain enables a simple integration of additional spectroscopic contrast methods, comparable to classical staining, to improve the selection [3, 4]. One of these additional contrast methods is optical spectroscopy [5, 6]. Grade IV indicates the most malignant tumours, such as glioblastoma multiforme [19,20,21,22]

Results

Discussion

Conclusion