Native chromatographic sample preparation of serum, plasma and cerebrospinal fluid does not comprise a risk for proteolytic biomarker loss.

We recently developed a native multidimensional chromatographic method for serum and plasma fractionation for proteomic biomarker search. This method has several advantages:parallelization and automation, high reproducibility and proteome coverage, flexible dynamic range with respect to molecular weight and sample amount, optional enzymatic and immunological analytics additional to mass spectrometry, retaining metabolites, and information on complex formation, modification, and fragmentation of constituents. Nevertheless, native conditions have the probable risk of proteome alteration and biomarker loss by intrinsic proteinases. Hence, we tried to quantify here intrinsic proteolytic activity in native samples and fractions from serum, plasma and cerebrospinal fluid, as well as the effectiveness of intrinsic anti-proteinases during sample handling and preparation under our fractionation conditions. Therefore, we used several quantitative measures: (1) total proportion of intrinsic protein and peptide fractions, (2) azocasein hydrolysis and (3) mass spectrometric protein coverage and peptide numbers. To 1: In all non-fractionated specimens, neither decrease of protein concentration or molecular weight nor increase of peptide concentration was found after variable clotting or pre-incubation time. To 2: No azocasein hydrolysis was seen in these samples when prepared within a few hours at room temperature. Trypsin, when added in concentrations not higher than 0.85 µg/mL (0.04 µM), even was completely inhibited. Moreover, in native 1-D fractions no proteinase activity could be observed. To 3: Mass spectrometry confirmed that neither protein coverage nor peptide numbers differ significantly in 1-D or 2-D fractions after variable incubation time. These results suggest that intrinsic, native proteinase inhibitors potentially protect the proteomes considered, enabling "top-down" proteomic approaches under native conditions with serum, plasma and cerebrospinal fluid.

Positron emission tomography imaging in human immunodeficiency virus-1-associated neurocognitive disorders: an interesting case and a review of the positron emission tomography literature.

Human immunodeficiency virus (HIV)-1-associated neurocognitive disorder can manifest with a variety of neurologic, cognitive, and behavioral impairments. We report a case of a 49-year-old non-HIV risk woman with an occult HIV infection who posed a diagnostic challenge as she suffered from a HIV-1-associated neurocognitive disorder with predominant motor symptoms mimicking upper motor neuron disease. Functional imaging using F-18 fluorodeoxyglucose positron emission tomography provided evidence of involvement of several cerebral regions which exhibited a distinct pattern of relative cerebral hypermetabolism (subcortical, brainstem, and cerebellar regions) and hypometabolism (sensorimotor cortex, mesiofrontal, and mesiotemporal areas) and functionally corresponded to the clinical symptoms. The results of the positron emission tomography scan are discussed in comparison with the current positron emission tomography literature and future perspectives are illustrated.