A collaborative EDNAP exercise on SNaPshot™-based mtDNA control region typing.

A collaborative European DNA Profiling (EDNAP) Group exercise was undertaken to assess the performance of an earlier described SNaPshot™-based screening assay (denoted mini-mtSNaPshot) (Weiler et al., 2016) [1] that targets 18 single nucleotide polymorphism (SNP) positions in the mitochondrial (mt) DNA control region and allows for discrimination of major European mtDNA haplogroups. Besides the organising laboratory, 14 forensic genetics laboratories were involved in the analysis of 13 samples, which were centrally prepared and thoroughly tested prior to shipment. The samples had a variable complexity and comprised straightforward single-source samples, samples with dropout or altered peak sizing, a point heteroplasmy and two-component mixtures resulting in one to five bi-allelic calls. The overall success rate in obtaining useful results was high (97.6%) given that some of the participating laboratories had no previous experience with the typing technology and/or mtDNA analysis. The majority of the participants proceeded to haplotype inference to assess the feasibility of assigning a haplogroup and checking phylogenetic consistency when only 18 SNPs are typed. To mimic casework procedures, the participants compared the SNP typing data of all 13 samples to a set of eight mtDNA reference profiles that were described according to standard nomenclature (Parson et al., 2014) [2], and indicated whether these references matched each sample or not. Incorrect scorings were obtained for 2% of the comparisons and derived from a subset of the participants, indicating a need for training and guidelines regarding mini-mtSNaPshot data interpretation.

Structural and functional neuroimaging studies of the suicidal brain.

Suicidality is a major challenge for today's health care. Evidence suggests that there are differences in cognitive functioning of suicidal patients but the knowledge about the underlying neurobiology is limited. Brain imaging offers the advantage of a non-invasive in vivo direct estimation of detailed brain structure, regional brain functioning and estimation of molecular processes in the brain. We have reviewed the literature on neuroimaging studies of the suicidal brain. This article contains studies on structural imaging such as Computed Tomography (CT) and Magnetic Resonance Imaging (MRI) and functional imaging, consisting of Positron Emission Tomography (PET), Single Photon Emission Tomography (SPECT) and functional MRI (fMRI). We classified the results of the different imaging modalities in structural and functional imaging. Within our research, we found no significant differences in the suicidal brain demonstrated by Computed Tomography. Magnetic Resonance Imaging studies in subjects with a history of suicide attempt on the other hand deliver differing results, mostly pointing at a higher prevalence of white (especially deep white matter and periventricular) and grey matter hyperintensities in the frontal, temporal and/or parietal lobe and decreased volumes in the frontal and temporal lobe. There seems to be a trend towards findings of reduced grey matter volume in the frontal lobe. Overall, there is no consensus of opinion on structural imaging of the suicidal brain. Research on functional imaging is further divided into studies in resting state, studies in activation conditions and studies on brain neurotransmitters, transporters and receptors. A common finding in functional neuroimaging in resting conditions is a decreased perfusion in the prefrontal cortex of suicidal patients. During cognitive activation, perfusion deficits in the prefrontal cortex have been observed. After fenfluramine challenge, the prefrontal cortex metabolism seems to be inversely correlated to the lethality of previous suicide attempt. The few studies that examined the serotonin transporter in suicide found no significant differences in binding potential. In suicide attempters there seems to be a negative correlation between impulsivity and SERT binding. Our group found a reduced 5-HT(2A) binding in the frontal cortex in patients with a recent suicide attempt. The binding index was significantly lower in the deliberate self injury patients compared to the deliberate self poisoning patients. The few authors that examined DAT binding in suicide found no significant DAT differences between patients and controls. However they demonstrated significant negative correlations between DAT binding potential and mental energy among suicide attempters, but not in healthy control subjects. We did not find studies measuring the binding potential of the noradrenalin or gamma amino butyric acid transporter or receptor in suicidal subjects. Several reports have suggested abnormalities of GABA neurotransmission in depression. During our literature search, we have focused on neuroimaging studies in suicidal populations, but in the absence of evidence in the literature on this group or when further collateral evidence is appropriate, this overview expands to results in impulsive aggressive or in depressed subjects.

Influence of charge variation in the Streptomyces venezuelae alpha-amylase signal peptide on heterologous protein production by Streptomyces lividans.

With the aim of investigating determining factors for secretion of heterologous proteins by streptomycetes, we analysed the effect of charge variation in the Streptomyces venezuelae ATCC15068 alpha-amylase signal peptide on expression and secretion of mouse tumour necrosis factor alpha (mTNF) by Streptomyces lividans. To this end, the mTNF cDNA was fused to the wild-type alpha-amylase (aml) signal sequence and the fusion gene was expressed under the control of the S. venezuelae CBS762.70 subtilisin inhibitor gene (vsi) promoter, which has been shown to be very effective in initiating transcription. In addition, the number of positive charges in the N region of the alpha-amylase signal peptide was altered by in vitro mutagenesis. Secreted and intracellular mTNF levels were determined by sodium dodecyl sulphate/polyacrylamide gel electrophoresis and biological activity measurements. This revealed moderate amounts of secreted mTNF compared to the levels obtained in previous experiments using the vsi promoter in combination with the Vsi signal peptide. Levels of secreted mTNF could be increased sevenfold by introducing one extra positive charge in the N region of the signal peptide.

Preparation of monospecific polyclonal antibodies against Sambucus nigra lectin related protein, a glycosylated plant protein.

A simple, but highly efficient, method was developed for the purification of monospecific antibodies against the plant glycoprotein Sambucus nigra lectin related protein. In a first step, the antiserum is purified by affinity chromatography on a column with the immobilized antigen. To deplete the affinity-purified antiserum from aspecific cross-reacting antibodies directed against the glycan part of the glycoprotein, a second affinity chromatography on an unrelated plant glycoprotein, in casu the Robinia pseudoacacia agglutinin, is included.