Bovine Brain Tissue Research Articles

Toward understanding the brain tissue behavior due to preconditioning: an experimental study and RVE approach.

Brain tissue under preconditioning, as a complex issue, refers to repeated loading-unloading cycles applied in mechanical testing protocols. In previous studies, only the mechanical behavior of the tissue under preconditioning was investigated; However, the link between macrostructural mechanical behavior and microstructural changes in brain tissue remains underexplored. This study aims to bridge this gap by investigating bovine brain tissue responses both before and after preconditioning. We employed a dual approach: experimental mechanical testing and computational modeling. Experimental tests were conducted to observe microstructural changes in mechanical behavior due to preconditioning, with a focus on axonal damage. Concurrently, we developed multiscale models using statistically representative volume elements (RVE) to simulate the tissue's microstructural response. These RVEs, featuring randomly distributed axonal fibers within the extracellular matrix, provide a realistic depiction of the white matter microstructure. Our findings show that preconditioning induces significant changes in the mechanical properties of brain tissue and affects axonal integrity. The RVE models successfully captured localized stresses and facilitated the microscopic analysis of axonal injury mechanisms. These results underscore the importance of considering both macro and micro scales in understanding brain tissue behavior under mechanical loading. This comprehensive approach offers valuable insights into mechanotransduction processes and improves the analysis of microstructural phenomena in brain tissue.

Replication of Akabane virus and related orthobunyaviruses in a fetal-bovine-brain-derived cell line.

Akabane virus (AKAV), Aino virus, Peaton virus, Sathuperi virus, and Shamonda virus are arthropod-borne viruses belonging to the order Elliovirales, family Peribunyaviridae, genus Orthobunyavirus. These viruses cause or may cause congenital malformations in ruminants, including hydranencephaly, poliomyelitis, and arthrogryposis, although their pathogenicity may vary among field cases. AKAV may cause relatively severe congenital lesions such as hydranencephaly in calves. Furthermore, strains of AKAV genogroups I and II exhibit different disease courses. Genogroup I strains predominantly cause postnatal viral encephalomyelitis, while genogroup II strains are primarily detected in cases of congenital malformation. However, the biological properties of AKAV and other orthobunyaviruses are insufficiently investigated in hosts in the field and in vitro. Here, we used an immortalized bovine brain cell line (FBBC-1) to investigate viral replication efficiency, cytopathogenicity, and host innate immune responses. AKAV genogroup II and Shamonda virus replicated to higher titers in FBBC-1 cells compared with the other viruses, and only AKAV caused cytopathic effects. These results may be associated with the severe congenital lesions in the brain caused by AKAV genogroup II. AKAV genogroup II strains replicated to higher titers in FBBC-1 cells than AKAV genogroup I strains, suggesting that genogroup II strains replicated more efficiently in fetal brain cells, accounting for the detection of the latter strains mainly in fetal infection cases. Therefore, FBBC-1 cells may serve as a valuable tool for investigating the virulence and tropism of the orthobunyaviruses for bovine neonatal brain tissues in vitro.

Isolation and Characterization of Neuroglobin and The Reducing Enzyme Metneuroglobin (Neuroglobin Fe3+) From Bovine Brain Tissue

Isolation and Characterization of Neuroglobin and The Reducing Enzyme Metneuroglobin (Neuroglobin Fe3+) From Bovine Brain Tissue

Complex Permittivity of Ex-Vivo Human, Bovine and Porcine Brain Tissues in the Microwave Frequency Range.

Accurate knowledge about the dielectric properties of biological tissues in the microwave frequency range may lead to advancement of biomedical applications based on microwave technology. However, the published data are very scarce, especially for human brain tissues. The aim of this work was to measure and report the complex permittivity of brain white matter, grey matter and cerebellum. Complex permittivity was measured on human, bovine and porcine brain tissues in the microwave frequency range from 0.5 to 18 GHz using an open-ended coaxial probe. The results present a valuable addition to the available data on the brain tissue complex permittivity. Some noticeable variations between the results lead to several conclusions. Complex permittivity variation within the same tissue type of the individual species was comparable to interspecies variation. The difference was prominent between human brains obtained from autopsies, while bovine brains obtained from healthy animals showed very similar complex permittivity. We hypothesize that the difference might have been caused by the basic pathologies of the patients, where the associated therapies could have affected the brain water content. We also examined the effect of excised tissue degradation on its complex permittivity over the course of three days, and the results suggest the gradual dehydration of the samples.

The bovine Prader-Willi/Angelman imprinted domain has four Sno-lncRNAs types.

Sno-lncRNAs are intron-derived long noncoding RNAs (lncRNAs) with snoRNA ends. Sno-lncRNAs were first discovered in the human Prader-Willi (PWS)/Angelman (AS) imprinted domain. Here, we report the identification and characterization of four sno-lncRNA types (sno-lncRNA1, sno-lncRNA2, sno-lncRNA3, and sno-lncRNA4) in the bovine PWS/AS imprinted domain. Reverse transcription-PCR first determined the cDNA sequences of the four bovine sno-lncRNAs. A gene structure analysis showed that sno-lncRNA1 lacks introns, but sno-lncRNA2 and sno-lncRNA3 have one and two introns respectively. The three sno-lncRNAs have similar snoRNA ends. Moreover, the three have similar snoRNAs at their 5' and 3' ends. The head-to-tail orientation has six sno-lncRNA copies arranged between bovine SNORD116-6 and SNORD116-12. Moreover, only a copy of sno-lncRNA4 was located between SNORD116-3 and SNORD116-4. The expression of the four sno-lncRNAs was analyzed in the bovine heart, liver, spleen, lung, kidney, muscle, fat, brain, and placenta tissues. The monoallelic expression of sno-lncRNA4 was determined in bovine tissues. The results showed that the four sno-lncRNAs are widely expressed in the nine tissues, although sno-lncRNA3 and sno-lncRNA4 were undetected in the placenta. Moreover, an informative single nucleotide polymorphism (rs448706424) revealed the allelic expression of sno-lncRNA4 in exon 2 of sno-lncRNA4. The bovine genome had six copies of sno-lncRNA1, sno-lncRNA2, and sno-lncRNA3, but their allelic expression was not identified.

The effect of large deformation on Poisson's ratio of brain white matter: An experimental study.

A more Accurate description of the mechanical behavior of brain tissue could improve the results of computational models. While most studies have assumed brain tissue as an incompressible material with constant Poisson's ratio of almost 0.5 and constructed their modeling approach according to this assumption, the relationship between this ratio and levels of applied strains has not yet been studied. Since the mechanical response of the tissue is highly sensitive to the value of Poisson's ratio, this study was designed to investigate the characteristics of the Poisson's ratio of brain tissue at different levels of applied strains. Samples were extracted from bovine brain tissue and tested under unconfined compression at strain values of 5%, 10%, and 30%. Using an image processing method, the axial and transverse strains were measured over a 60-s period to calculate the Poisson's ratio for each sample. The results of this study showed that the Poisson's ratio of brain tissue at strain levels of 5% and 10% was close to 0.5, and assuming brain tissue as an incompressible material is a valid assumption at these levels of strain. For samples under 30% compression, this ratio was higher than 0.5, which could suggest that under strains higher than the brain injury threshold (approximately 18%), tissue integrity was impaired. Based on these observations, it could be concluded that for strain levels higher than the injury threshold, brain tissue could not be assumed as an incompressible material, and new material models need to be proposed to predict the material behavior of the tissue. In addition, the results showed that brain tissue under unconfined compression uniformly stretched in the transverse direction, and the bulging in the samples is negligible.

Brain-specific monoallelic expression of bovine UBE3A is associated with genomic position.

Genomic imprinting is a rare epigenetic process in mammalian cells that leads to monoallelic expression of a gene with a parent-specific pattern. The UBE3A (ubiquitin protein ligase E3A) gene is imprinted with maternal allelic expression in the brain but biallelically expressed in all other tissues in humans. The silencing of the paternal UBE3A allele is thought to be caused by the paternally expressed antisense RNA transcript of UBE3A-ATS. The aberrant imprinted expression of the UBE3A is associated with several neurodevelopmental syndromes and psychological disorders. Cattle are a valuable model species in determining the genetic etiology of sporadic human disorder, and maternal expression of UEB3A has been revealed by next-generation sequencing study in the bovine conceptus. In this study, we investigated the allelic expression of UBE3A and UBE3A-ATS in adult bovine somatic tissues. To confirm the splicing pattern of bovine UBE3A, five 5' alternative transcripts (MT210534-MT210538) were first obtained from bovine brain tissue by RT-PCR. Based on 10 SNP genotypes, we found that the brain-specific monoallelic expression of bovine UBE3A did not occur along the entire locus, and there was a shift from biallelic expression to monoallelic expression in exon 14 of the UBE3A gene. However, the brain-specific monoallelic expression of bovine UBE3A-ATS occurred in the entire gene. These observations demonstrated that the monoallelic expression did not occur along the bovine UBE3A entire locus and was associated with the genomic position.

Dynamic mechanical characterization and viscoelastic modeling of bovine brain tissue

Brain tissue is vulnerable and sensitive, predisposed to potential damage under various conditions of mechanical loading. Although its material properties have been investigated extensively, the frequency-dependent viscoelastic characterization is currently limited. Computational models can provide a non-invasive method by which to analyze brain injuries and predict the mechanical response of the tissue. The brain injuries are expected to be induced by dynamic loading, mostly in compression and measurement of dynamic viscoelastic properties are essential to improve the accuracy and variety of finite element simulations on brain tissue. Thus, the aim of this study was to investigate the compressive frequency-dependent properties of brain tissue and present a mathematical model in the frequency domain to capture the tissue behavior based on experimental results. Bovine brain specimens, obtained from four locations of corona radiata, corpus callosum, basal ganglia and cortex, were tested under compression using dynamic mechanical analysis over a range of frequencies between 0.5 and 35 Hz to characterize the regional and directional response of the tissue. The compressive dynamic properties of bovine brain tissue were heterogenous for regions but not sensitive to orientation showing frequency dependent statistical results, with viscoelastic properties increasing with frequency. The mean storage and loss modulus were found to be 12.41 kPa and 5.54 kPa, respectively. The material parameters were obtained using the linear viscoelastic model in the frequency domain and the numeric simulation can capture the compressive mechanical behavior of bovine brain tissue across a range of frequencies. The frequency-dependent viscoelastic characterization of brain tissue will improve the fidelity of the computational models of the head and provide essential information to the prediction and analysis of brain injuries in clinical treatments.

Heat shock protein 90α increases superoxide generation from neuronal nitric oxide synthases

Neuronal nitric oxide synthase (nNOS) generates superoxide, particularly at sub-optimal l-arginine (l-Arg) substrate concentrations. Heat shock protein 90 (Hsp90) was reported to inhibit superoxide generation from nNOS protein. However, commercially available Hsp90 product from bovine brain tissues with unspecified Hsp90α and Hsp90β contents and an undefined Hsp90 protein oligomeric state was utilized. These two Hsp90s can have opposite effect on superoxide production by NOS. Importantly, emerging evidence indicates that nNOS splice variants are involved in different biological functions by functioning distinctly in redox signaling. In the present work, purified recombinant human Hsp90α, in its native dimeric state, was used in electron paramagnetic resonance (EPR) spin trapping experiments to study the effects of Hsp90α on superoxide generation from nNOS splice variants nNOSμ and nNOSα. Human Hsp90α was found to significantly increase superoxide generation from nNOSμ and nNOSα proteins under l-Arg-depleted conditions and Hsp90α influenced superoxide production by nNOSμ and nNOSα at varying degrees. Imidazole suppressed the spin adduct signal, indicating that superoxide was produced at the heme site of nNOS in the presence of Hsp90α, whereas l-Arg repletion diminished superoxide production by the nNOS-Hsp90α. Moreover, NADPH consumption rate values exhibited a similar trend/difference as a function of Hsp90α and l-Arg. Together, these EPR spin trapping and NADPH oxidation kinetics results demonstrated noticeable Hsp90α-induced increases in superoxide production by nNOS and a distinguishable effect of Hsp90α on nNOSμ and nNOSα proteins.

Archeological neuroimmunology: resurrection of a pathogenic immune response from a historical case sheds light on human autoimmune encephalomyelitis and multiple sclerosis

Aim of our study was to identify the target auto-antigen in the central nervous system recognized by the immune system of a unique patient, who died more than 60 years ago from a disease with pathological changes closely resembling multiple sclerosis (MS), following a misguided immunization with lyophilized calf brain tissue. Total mRNA was isolated from formaldehyde fixed and paraffin embedded archival brain tissue containing chronic active inflammatory demyelinating lesions with inflammatory infiltrates rich in B-lymphocytes and plasma cells. Analysis of the transcriptome by next generation sequencing and reconstruction of the dominant antibody by bioinformatic tools revealed the presence of one strongly expanded B-cell clone, producing an autoantibody against a conformational epitope of myelin oligodendrocytes glycoprotein (MOG), similar to that recognized by the well characterized monoclonal anti-MOG antibody 8-18C5. The reconstructed antibody induced demyelination after systemic or intrathecal injection into animals with T-cell mediated encephalomyelitis. Our study suggests that immunization with bovine brain tissue in humans may—in a small subset of patients—induce a disease with an intermediate clinical and pathological presentation between MS and MOG-antibody associated inflammatory demyelinating disease (MOGAD).

A constrained particle swarm optimization algorithm for hyperelastic and visco-hyperelastic characterization of soft biological tissues

A constrained particle swarm optimization algorithm (C-PSO) is introduced and modified for hyperelastic and visco-hyperelastic characterization of bovine brain tissue at three different strain rates. Using the elasticity compatibility and Drucker’s stability criterion, the constraints of the hyperelastic and visco-hyperelastic models are identified and implemented in the C-PSO algorithm and its performance is compared with the classic curve fitting algorithms including Levenberg-Marquardt and trust region reflective. The accuracy of the C-PSO was found to be superior for visco-hyperelastic characterization, as for some strain rates, the trust region reflective algorithm failed to provide a reasonable approximation.

Absorption of 5G Radiation in Brain Tissue as a Function of Frequency, Power and Time

The rapid release of 5G wireless communications networks has spurred renewed concerns regarding the interactions of higher radiofrequency (RF) radiation with living species. We examine RF exposure and absorption in ex vivo bovine brain tissue and a brain simulating gel at three frequencies: 1.9 GHz, 4 GHz and 39 GHz that are relevant to current (4G), and upcoming (5G) spectra. We introduce a highly sensitive thermal method for the assessment of radiation exposure, and derive experimentally, accurate relations between the temperature rise (ΔT), specific absorption rate (SAR) and the incident power density (F), and tabulate the coefficients, ΔT/ΔF and Δ(SAR)/ΔF, as a function of frequency, depth and time. This new method provides both ΔT and SAR applicable to the frequency range below and above 6 GHz as shown at 1.9, 4 and 39 GHz, and demonstrates the most sensitive experimental assessment of brain tissue exposure to millimeter-wave radiation to date, with a detection limit of 1 mW. We examine the beam penetration, absorption and thermal diffusion at representative 4G and 5G frequencies and show that the RF heating increases rapidly with frequency due to decreasing RF source wavelength and increasing power density with the same incident power and exposure time. We also show the temperature effects of continuous wave, rapid pulse sequences and single pulses with varying pulse duration, and we employ electromagnetic modeling to map the field distributions in the tissue. Finally, using this new methodology, we measure the thermal diffusivity of ex vivo bovine brain tissue experimentally.

Rate-dependent constitutive modeling of brain tissue.

In this paper, the dynamic behavior of bovine brain tissue, measured from a set of in vitro experiments, is investigated and represented through a nonlinear viscoelastic constitutive model. The brain samples were tested by employing unconfined compression tests at three different deformation rates of 10, 100, and 1000mm/s. The tissue exhibited a significant rate-dependent behavior with different compression speeds. Based onthe parallel rheological framework approach, a nonlinear viscoelastic model that captures the key aspects of the rate dependency in large-strain behavior was introduced. The proposed model was numerically calibrated to the tissue test data from three different deformation rates. The determined material parameters provided an excellent constitutive representation of tissue response in comparison with the test results. The obtained material parameters were employed in finite element simulations of tissue under compression loadings and successfully verified by the experimental results, thus demonstrating the computational compatibility of the proposed material model. The results of this paper provide groundwork in developing a characterization framework for large-strain and rate-dependent behavior of brain tissue at moderate to high strain rates which is of thehighest importance in biomechanical analysis of the traumatic brain injury.

A poro-hyper-viscoelastic rate-dependent constitutive modeling for the analysis of brain tissues

In this paper, the dynamic behavior of bovine brain tissue, measured from in-vitro unconfined compression tests, is examined and represented through a viscoelastic biphasic model. The experiments have been carried out under three compression speeds of 10, 100, and 1000 mm/s. The results exhibited significant rate-dependent behavior. The brain tissue is modeled as a biphasic continuum consisting of a compressible solid matrix, fully saturated with an incompressible interstitial fluid. The governing equations based on conservation of mass and momentum are used to describe the solid-fluid interactions. An inverse scheme is employed in which a finite element model runs iteratively to optimize constitutive constants. The obtained material parameters of the proposed biphasic model show relatively good agreement (R2 ≥ 0.96) with the experimental tissue mechanical responses at different rates. The model can successfully capture the key aspects of the rate-dependency for both solid and fluid phases under large strain deformation. This poro-hyper viscoelastic model can effectively estimate the global and local rate-dependent tissue deformations, the spatial variations in pore spaces, hydrostatic pressure as well as fluid diffusion through the tissue.

Anomalous water dynamics in brain: a combined diffusion magnetic resonance imaging and neutron scattering investigation.

Water diffusion is an optimal tool for investigating the architecture of brain tissue on which modern medical diagnostic imaging techniques rely. However, intrinsic tissue heterogeneity causes systematic deviations from pure free-water diffusion behaviour. To date, numerous theoretical and empirical approaches have been proposed to explain the non-Gaussian profile of this process. The aim of this work is to shed light on the physics piloting water diffusion in brain tissue at the micrometre-to-atomic scale. Combined diffusion magnetic resonance imaging and first pioneering neutron scattering experiments on bovine brain tissue have been performed in order to probe diffusion distances up to macromolecular separation. The coexistence of free-like and confined water populations in brain tissue extracted from a bovine right hemisphere has been revealed at the micrometre and atomic scale. The results are relevant for improving the modelling of the physics driving intra- and extracellular water diffusion in brain, with evident benefit for the diffusion magnetic resonance imaging technique, nowadays widely used to diagnose, at the micrometre scale, brain diseases such as ischemia and tumours.

Optimum recovery time for cyclic compression tests on bovine brain tissue

In conducting mechanical tests on the brain tissue, it is preferred to perform multiple tests on the same sample. In this study we investigated the behavior of the bovine brain tissue in repeated compression tests with six recovery periods (10, 60, 120, 180, 240 and 300 s). Compression tests were performed on cylindrical samples with an average diameter and height of 18.0 mm and 15.0 mm respectively. Two testing protocols were employed: first protocol comprised of experiments with 5, 25 and 125 mm/min loading speed up to 33% strain and the second protocol consisted of tests with 25 and 125 mm/min loading speed up to 17% strain. Each experiment was conducted in two cycles separated by a specific recovery period. Stress-strain data from the first and second cycles were compared using three criteria, namely Normalized root-mean-square error (NRMSE), coefficient of variation (R2) and effective height ratio (EHR). The analysis suggests that the optimum recovery period for the first and second protocols are 120 s and 180 s respectively. Moreover, differences between the first and second cycles of medium and high speed tests were found to be smaller compared to the low-speed experiments.

Easy Determination of Radiation Absorption in Brain Tissue from Mobile Phones Using Finite Element Method

Brain tissue plays a significant role in both cognitive and psychomotor behavior of humans. However, their interaction with radiation emanating from hand held mobile devices is still not fully understood. This research was aimed at investigating radiation absorption in brain tissue. Bovine brain tissues ranging from lesser than 1 year to greater than 10 years of age were bought from a specialty store (Sigma-Aldrich). The tissues were used within 72 h of extraction for ex vivo brain experiments. The brain tissue was stored at 6°C and then 16°C for 24 h in the MRI room to reach thermal equilibrium before any experiments were undertaken. The averages for the dielectric constant were measured from 1 - 4 GHz using open ended coaxial probe (OECP) (85,070E; Agilent Technologies). The results obtained for the dielectric properties were then used as raw data in the numerical computation and simulation of the radiation absorption by the brain tissues for both adolescent and adults bovine brain tissue using finite element method (FEM). The measured dielectric constants varied for the different brain tissue from 54.39 to 39.29. Analysis showed that adolescents tissue absorbed more radiation than adults from mobile phoneradiation which is due to the higher dielectric property of adolescent brain tissue. The results obtained can be applied to human brain tissue since bovine shares the same compositional properties with humans.

Development of Computed Tomography Head and Body Phantom for Organ Dosimetry

Introduction: Quality assurance in Computed tomography (CT) centers in developing countries are largely hindered by the unavailability of CT phantoms. The development of a local CT phantom for the measurement of organ radiation absorbed dose is therefore requisite. Material and Methods: Local CT phantoms were designed to meet the standard criteria of 32 cm diameter for body, 16 cm diameter for head, and 14 cm in length respectively. The outer plastic shell was made using poly (methyl methacrylate [PMMA]) sheet. The developed CT phantoms were validated against a standard phantom. Radiation absorbed dose was determined by scanning the setup with the same protocol used for the standard phantom. The local phantoms were then verified for organ radiation absorbed dose measurement using bovine tissues. The set up was CT-scanned, and Hounsfield units (HU) for bovine tissues were obtained. Results: There was no significant difference between the local and standard head phantoms (P=0.060). Similarly, no difference was noted between the local and standard body phantoms (P=0.795). The percentage difference in volume CT dose index (CTDIvol) between the body (local and standard) phantoms was higher than that for the head phantoms. There were no significant differences in HU between bovine and human brain, liver, kidney and lung tissues (P=0.938). Conclusion: The local phantoms showed good agreement with the standard ones. The developed phantoms can be used for CT organ radiation absorbed dose measurement in radiology departments in Nigeria.

Abstract 4111: Water content based Electrical Properties Tomography (wEPT) for modelling delivery of Tumor Treating Fields to the brain

Abstract Objective: The purpose of this study was to investigate the application of Water content based Electrical Properties Tomography for mapping electrical properties (EPs) of brain tissues in the frequency range of 100-1000 kHz. Background: TTFields are electric fields with frequencies of 100-500 kHz that disrupt mitosis. TTFields are approved for the treatment of glioblastoma multiforme. Determining the EPs of brain tissues is important for understanding how TTFields distribute within the head. The EPs of tissues are heterogeneous, especially in the region of the tumor. Therefore methods that map EPs within the brain with high spatial resolution are desired. Water content based EP tomography (wEPT) is a method that utilizes the ratio of two T1w MRI images with different relaxation times (TRs) to map EPs based on empirically derived relationships between T1, water content (WC) and EPs. wEPT has been applied to map EPs of healthy brain at 128 MHz using typical WC and EP values of healthy tissues reported in the literature to derive the empirical models. Here we adapted wEPT to map EPs in the 100-1000 kHz range utilizing in-house measurements of WC and EPs from healthy bovine and tumor-bearing rat brain tissue. Methods: The empirical model connecting MRI images, WC and EPs in the 100-1000 kHz range were created using 32 tissue samples derived from three 3 calf brains and 1 CSF sample of a pig. For each sample, T1w MRIs with TRs {700, 4000} ms were acquired and the image ratio (Ir) between the images was calculated. EPs of samples were measured using parallel plates, and WC was measured by the wet-dry weight method. Curve fitting yielded empirical models connecting Ir, WC and EP. Next, T1w MRIs of in-vivo tumor-bearing rat brains and 6 ex-vivo pieces of calf brain were acquired, and the empirical curves described above were used to map WC and EP within the rat brains and the pieces of calf brain. EPs and WC were measured on 6 small samples excised from each imaged brain. For each sample, measured values were compared to the median WC and EPs extracted from the corresponding Regions of Interest (ROIs) in the wEPT map. Results: Anatomical structures and the tumor were clearly visible in wEPT maps. WC estimated using wEPT agreed well with measurements on excised samples. There was a clear connection between EPs estimated with wEPT and the measured values. However, in some samples large differences between wEPT-derived EP values and measurements were found. In particular, the differences between tumor and healthy tissues conductivity estimated using wEPT was significantly higher than the measured difference in conductivities within the corresponding excised samples. Conclusion: wEPT maps WC in healthy and tumor brain tissues and provides information on local electrical properties at frequencies of 100-1000 kHz. Further investigation is needed to clarify the relationship between WC and EP within this frequency range. Citation Format: Catherine Tempel-Brami, Cornelia Wenger, Hadas S. Hershkovich, Moshe Giladi, Ze'ev Bomzon. Water content based Electrical Properties Tomography (wEPT) for modelling delivery of Tumor Treating Fields to the brain [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4111.

Detection of Bovine Central Nervous System Tissues in Rendered Animal By-Products by One-Step Real-Time Reverse Transcription PCR Assay

Contamination of rendered animal byproducts with central nervous system tissues (CNST) from animals with bovine spongiform encephalopathy is considered one of the vehicles of disease transmission. Removal from the animal feed chain of CNST originated from cattle of a specified age category, species-labeling of rendered meat products, and testing of rendered products for bovine CNST are tasks associated with the epidemiological control of bovine spongiform encephalopathy. A single-step TaqMan real-time reverse transcriptase (RRT) PCR assay was developed and evaluated for specific detection of bovine glial fibrillary acidic protein (GFAP) mRNA, a biomarker of bovine CNST, in rendered animal by-products. An internal amplification control, mammalian b -actin mRNA, was coamplified in the duplex RRT-PCR assay to monitor amplification efficiency, normalize amplification signals, and avoid false-negative results. The functionality of the GFAP mRNA RRT-PCR was assessed through analysis of laboratory-generated binary mixtures of bovine central nervous system (CNS) and muscle tissues treated under various thermal settings imitating industrial conditions. The assay was able to detect as low as 0.05 % (wt/wt) bovine brain tissue in binary mixtures heat treated at 110 to 130°C for 20 to 60 min. Further evaluation of the GFAP mRNA RRT-PCR assay involved samples of industrial rendered products of various species origin and composition obtained from commercial sources and rendering plants. Low amounts of bovine GFAP mRNA were detected in several bovine-rendered products, which was in agreement with declared species composition. An accurate estimation of CNS tissue content in industrial-rendered products was complicated due to a wide range of temperature and time settings in rendering protocols. Nevertheless, the GFAP mRNA RRT-PCR assay may be considered for bovine CNS tissue detection in rendered products in combination with other available tools (for example, animal age verification) in inspection programs.