Glial-fibrillary-acidic-protein (GFAP) biomarker detection in serum-matrix: Functionalization strategies and detection by an ultra-high-frequency surface-acoustic-wave (UHF-SAW) lab-on-chip.

Glial-fibrillary-acidic-protein (GFAP) has recently drawn significant attention from the clinical environment as a promising biomarker. The pathologies which can be linked to the presence of GFAP in blood severely affect the human central nervous system. These pathologies are glioblastoma multiforme (GBM), traumatic brain injuries (TBIs), multiple sclerosis (MS), intracerebral hemorrhage (ICH), and neuromyelitis optica (NMO). Here, we develop three different detection strategies for GFAP, among the most popular in the biosensing field and never examined side by side within the experimental frame. We compare their capability of detecting GFAP in a clean-buffer and serum-matrix by using gold-coated quartz-crystal-microbalance (QCM) sensors. All the three detection strategies are based on antibodies, and each of them focuses on a key aspect of the biosensing process. The first is based on a polyethylene glycol (PEG) chain for antifouling, the second on a protein-G linker for controlling antibody-orientation, and the third on antibody-splitting and direct surface immobilization for high-surface coverage. Then, we select the best-performing protocol and validate its detection performance with an ultra-high-frequency (UHF) surface-acoustic-wave (SAW) based lab-on-chip (LoC). GFAP successful detection is demonstrated in a clean-buffer and serum-matrix at a concentration of 35 pM. This GFAP level is compatible with clinical diagnostics. This result suggests the use of our technology for the realization of a point-of-care biosensing platform for the detection of multiple brain-pathology biomarkers.

Ultra-High Mass Resolution MALDI Imaging Mass Spectrometry of Proteins and Metabolites in a Mouse Model of Glioblastoma.

MALDI mass spectrometry imaging is able to simultaneously determine the spatial distribution of hundreds of molecules directly from tissue sections, without labeling and without prior knowledge. Ultra-high mass resolution measurements based on Fourier-transform mass spectrometry have been utilized to resolve isobaric lipids, metabolites and tryptic peptides. Here we demonstrate the potential of 15T MALDI-FTICR MSI for molecular pathology in a mouse model of high-grade glioma. The high mass accuracy and resolving power of high field FTICR MSI enabled tumor specific proteoforms, and tumor-specific proteins with overlapping and isobaric isotopic distributions to be clearly resolved. The protein ions detected by MALDI MSI were assigned to proteins identified by region-specific microproteomics (0.8 mm2 regions isolated using laser capture microdissection) on the basis of exact mass and isotopic distribution. These label free quantitative experiments also confirmed the protein expression changes observed by MALDI MSI and revealed changes in key metabolic proteins, which were supported by in-situ metabolite MALDI MSI.

Time evolution of interhemispheric coupling in a model of focal neocortical epilepsy in mice.

Epilepsy is characterized by substantial network rearrangements leading to spontaneous seizures and little is known on how an epileptogenic focus impacts on neural activity in the contralateral hemisphere. Here, we used a model of unilateral epilepsy induced by injection of the synaptic blocker tetanus neurotoxin (TeNT) in the mouse primary visual cortex (V1). Local field potential (LFP) signals were simultaneously recorded from both hemispheres of each mouse in acute phase (peak of toxin action) and chronic condition (completion of TeNT effects). To characterize the neural electrical activities the corresponding LFP signals were analyzed with several methods of time series analysis. For the epileptic mice, the spectral analysis showed that TeNT determines a power redistribution among the different neurophysiological bands in both acute and chronic phases. Using linear and nonlinear interdependence measures in both time and frequency domains, it was found in the acute phase that TeNT injection promotes a reduction of the interhemispheric coupling for high frequencies (12-30 Hz) and small time lag (<20 ms), whereas an increase of the coupling is present for low frequencies (0.5-4 Hz) and long time lag (>40 ms). On the other hand, the chronic period is characterized by a partial or complete recovery of the interhemispheric interdependence level. Granger causality test and symbolic transfer entropy indicate a greater driving influence of the TeNT-injected side on activity in the contralateral hemisphere in the chronic phase. Lastly, based on experimental observations, we built a computational model of LFPs to investigate the role of the ipsilateral inhibition and exicitatory interhemispheric connections in the dampening of the interhemispheric coupling. The time evolution of the interhemispheric coupling in such a relevant model of epilepsy has been addressed here.

Association of anthracycline derivatives with DNA: a fluorescence study.

The DNA affinity for 26 anthracycline derivatives was studied by the quenching fluorescence technique. The stereochemical requirements for DNA intercalation are discussed. The relationship between the DNA affinity and bioactivity is also pointed out.

Self-association of doxorubicin and related compounds in aqueous solution.

The dimerization of doxorubicin, daunorubicin, and their 4-demethoxy, 4'-epi, and 4'-deoxy analogues was studied spectrophotometrically. Self-association was found to be influenced by buffer composition and ionic strength. Kd values were 1.3 X 10(4) and 2.3 X 10(4) M-1 for doxorubicin and daunorubicin, respectively, and ranged from 3.8 X 10(3) to 6.1 X 10(3) M-1 for the 4-demethoxy analogues. For 4'-epi- and 4'-deoxydoxorubicin, tetramerization has also been considered. On this basis, values of 2.0 X 10(4) and 2.2 X 10(4) M-1 were found, respectively, for the formation constant of the dimerization process. Stability of the dimeric species appears to be strongly influenced by substitution of the chromophore moiety.