Developing formalin-based fixative agents for post-mortem brain MRI at 9.4T


Purpose: Ultra-high field Magnetic Resonance Imaging of whole post-mortem brains is complementary to histology and pathology studies and can be useful for tissue characterization, sectioning, 3D-reconstruction of sectioned tissue data, and for diagnosis and investigation of neurodegenerative diseases. To preserve the tissue, the cerebrospinal fluid must be replaced by a fixative agent such as formalin.
Here we show how such agents can be tailored to achieve a) dielectric properties that ensure a homogeneous B1-field, b) a magnetic susceptibility matching the tissue to improve B0 homogeneity, and c) a singulet 1H-NMR spectra to prevent chemical-shift-artefacts.

Method: Dielectric properties of formalin-based agents were assessed (100MHz-4.5GHz), and four candidate fixatives with/without polyvinylpyrrolidone (PVP) and different salt concentrations were formulated. B1-field and MR-properties (T1, R2*, R2, R2 and magnetic susceptibility (QSM)) were observed in white and gray matter of pig brain samples during 0.5-35days of immersion fixation. The kinetics was fitted using exponential functions. The immersion time required to reach maximum R2*-values at different tissue-depths was used to estimate the Medawar-coefficient for fixative penetration. The impact of replacing the fixatives with Fluoroinert and phosphate-buffered-saline (PBS) as embedding media was also evaluated.

Results: The dielectric properties of formalin were non-linearly modified by increasing amounts of additives. With 5%PVP and 0.04%NaCl, the dielectric properties and B1-field reflected in-vivo-conditions (Figure 1). The highest B1 values were found in white matter with PVP and varied significantly with tissue depth and embedding media, but not with immersion-time (Figure 2). MR-properties depended on PVP yielding lower T1, higher R2* a more paramagnetic QSM values, and a lower Medawar-coefficient(0.9mm/(hour)0.5 ; without PVP: 1.5). Regardless of fixative, switching to PBS as embedder caused a paramagnetic shift in QSM and decreased R2* that progressed during 1month of storage, while no differences were found with Fluorinert.

Conclusion: In vivo-like B1-fields can be achieved in formalin-fixatives using PVP and a low salt-concentration, yielding lower T1, higher R2* and more paramagnetic QSM than without additives. The kinetics of R2* allowed estimation of fixative tissue penetration (Figure 3).

Azadeh Nazemorroaya, Ali Aghaeifar, Thomas Shiozawa, Bernhard Hirt, Hildegard Schulz, Klaus Scheffler, and Gisela E. Hagberg
Developing formalin-based fixative agents for post-mortem brain MRI at 9.4T
Magnetic Resonance in Medicine. 21 Dec 2021.
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