Extracellular RNAs present in biofluids have emerged as potential biomarkers for disease. Where most studies focus on blood-derived fluids, other biofluids may be more informative. We present an atlas of messenger, circular, and small RNA transcriptomes of a comprehensive collection of 20 human biofluids. By means of synthetic spike-in controls, we compare RNA content across biofluids, revealing a 10,000-fold difference in concentration. The circular RNA fraction is increased in most biofluids compared to tissues. Each biofluid transcriptome is enriched for RNA molecules derived from specific tissues and cell types. Our atlas enables an informed selection of the most relevant biofluid to monitor particular diseases. To verify the biomarker potential in these biofluids, four validation cohorts representing a broad spectrum of diseases were profiled, revealing numerous differential RNAs between case and control subjects. Spike-normalized data are publicly available in the R2 web portal for further exploration.

• MeSH
• Biomarkers * MeSH
• Cohort Studies MeSH
• Humans MeSH
• RNA genetics   metabolism   MeSH
• Sequence Analysis, RNA methods   MeSH
• Gene Expression Profiling methods   MeSH
• Body Fluids metabolism   MeSH
• Transcriptome * MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, N.I.H., Extramural MeSH

The discovery of α-synuclein (α-syn) as a major component of Lewy bodies, neuropathological hallmark of Parkinson's disease (PD), dementia with Lewy bodies and of glial inclusions in multiple system atrophy initiated the investigation of α-syn as a biomarker in cerebrospinal fluid (CSF). Due to the involvement of the periphery in PD the quantification of α-syn in peripheral fluids such as serum, plasma and saliva has been investigated as well. We review how the development of multiple assays for the quantification of α-syn has yielded novel insights into the variety of α-syn species present in the different fluids; the optimal preanalytical conditions required for robust quantification and the potential clinical value of α-syn as biomarker. We also suggest future approaches to use of CSF α-syn in neurodegenerative diseases.

• MeSH
• alpha-Synuclein MeSH
• Biomarkers MeSH
• Clinical Chemistry Tests MeSH
• Humans MeSH
• Neurodegenerative Diseases diagnosis   genetics   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH

BACKGROUND: Pathogenic heterozygous mutations in the progranulin gene (GRN) are a key cause of frontotemporal dementia (FTD), leading to significantly reduced biofluid concentrations of the progranulin protein (PGRN). This has led to a number of ongoing therapeutic trials aiming to treat this form of FTD by increasing PGRN levels in mutation carriers. However, we currently lack a complete understanding of factors that affect PGRN levels and potential variation in measurement methods. Here, we aimed to address this gap in knowledge by systematically reviewing published literature on biofluid PGRN concentrations. METHODS: Published data including biofluid PGRN concentration, age, sex, diagnosis and GRN mutation were collected for 7071 individuals from 75 publications. The majority of analyses (72%) had focused on plasma PGRN concentrations, with many of these (56%) measured with a single assay type (Adipogen) and so the influence of mutation type, age at onset, sex, and diagnosis were investigated in this subset of the data. RESULTS: We established a plasma PGRN concentration cut-off between pathogenic mutation carriers and non-carriers of 74.8 ng/mL using the Adipogen assay based on 3301 individuals, with a CSF concentration cut-off of 3.43 ng/mL. Plasma PGRN concentration varied by GRN mutation type as well as by clinical diagnosis in those without a GRN mutation. Plasma PGRN concentration was significantly higher in women than men in GRN mutation carriers (p = 0.007) with a trend in non-carriers (p = 0.062), and there was a significant but weak positive correlation with age in both GRN mutation carriers and non-carriers. No significant association was seen with weight or with TMEM106B rs1990622 genotype. However, higher plasma PGRN levels were seen in those with the GRN rs5848 CC genotype in both GRN mutation carriers and non-carriers. CONCLUSIONS: These results further support the usefulness of PGRN concentration for the identification of the large majority of pathogenic mutations in the GRN gene. Furthermore, these results highlight the importance of considering additional factors, such as mutation type, sex and age when interpreting PGRN concentrations. This will be particularly important as we enter the era of trials for progranulin-associated FTD.

• MeSH
• Frontotemporal Dementia * genetics   pathology   MeSH
• Humans MeSH
• Membrane Proteins genetics   MeSH
• Intercellular Signaling Peptides and Proteins genetics   MeSH
• Mutation genetics   MeSH
• Progranulins genetics   MeSH
• Nerve Tissue Proteins genetics   MeSH
• Virulence MeSH
• Check Tag
• Humans MeSH
• Male MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Systematic Review MeSH

With increasing demands on protein analyses in complex biological matrices, the insistence on developing new sample preparation techniques is rising. Recently, we introduced a new displacement electrophoresis technique (epitachophoresis) and instrumentation for preparative concentration and cleaning of DNA samples. This work describes the possibility of applying this device to protein samples. We have developed a method for the epitachophoretic concentration of proteins in a cationic mode and tested it by concentrating and collecting the protein zones from complex biological matrices (urine and growth medium). Under optimized conditions, we have obtained recoveries up to 99%. Furthermore, the applicability of the developed method was proven by concentrating and collecting the cytochrome c zone from a HeLa cell line growth medium, where the protein cytochrome c was released during cell apoptosis.

• MeSH
• Cytochromes c MeSH
• HeLa Cells MeSH
• Isotachophoresis * methods   MeSH
• Humans MeSH
• Proteins MeSH
• Body Fluids * MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

Lung cancer (LC) incidence represents 11.5% of all new cancers, resulting in 1.72 million deaths worldwide in 2015. With the aim to investigate the capability of the electronic nose (e-nose) technology for detecting and differentiating complex mixtures of volatile organic compounds in biofluids ex-vivo, we enrolled 50 patients with suspected LC and 50 matching controls. Tissue biopsy was taken from suspicious lung mass for histopathological evaluation and blood, exhaled breath, and urine samples were collected from all participants and qualitatively processed using e-nose. Odor-print patterns were further analysed using the principal component analysis (PCA) and artificial neural network (ANN) analysis. Adenocarcinoma, non-small cell LC and squamous cell carcinoma were the predominant pathological types among LC patients. PCA cluster-plots showed a clear distinction between LC patients and controls for all biological samples; where the overall success ratios of classification for principal components #1 and #2 were: 95.46, 82.01, and 91.66% for blood, breath and urine samples, respectively. Moreover, ANN showed a better discrimination between LC patients and controls with success ratios of 95.74, 91.67 and 100% for blood, breath and urine samples, respectively. The e-nose is an easy noninvasive tool, capable of identifying LC patients from controls with great precision.

• MeSH
• Breath Tests methods   instrumentation   MeSH
• Electronic Nose MeSH
• Middle Aged MeSH
• Humans MeSH
• Lung Neoplasms * diagnostic imaging   diagnosis   blood   urine   MeSH
• Predictive Value of Tests MeSH
• Prospective Studies MeSH
• Sensitivity and Specificity MeSH
• Case-Control Studies MeSH
• Volatile Organic Compounds analysis   MeSH
• Check Tag
• Middle Aged MeSH
• Humans MeSH
• Male MeSH
• Female MeSH

• MeSH
• Alzheimer Disease * metabolism   MeSH
• Autophagy * physiology   MeSH
• Biomarkers metabolism   MeSH
• Humans MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Review MeSH

A sensitive and specific approach was developed for the determination of Haemophilus influenza using DNA based bio-assay. In this study, citrate capped silver nanoparticle was synthesized and employed for bioconjugation with pDNA toward target sequences detection. In this study, synthesized probe (SH-5'-AAT TTT CCA ACT TTT TCA CCT GCA T-3') of Haemophilus influenza was detected with great sensitivity and selectivity after hybridization with cDNA (5'-ATG CAG GTG AAA AAG TTG GAA AAT T-3'). Regarding to the obtained results, the low limit of quantification (LLOQ) of DNA sample was 1 ZM using 15 μL of probe and 200 μL of Cit/AgNPs. According to ultra-sensitivity of the fabricated optical DNA-based bio-assay, it has potential for bacterial determination both in clinical and environmental specimens. To evaluate the selectivity of developed DNA based biosensor, three mismatch sequences were applied. Finally, the designed genosensor is a significant diagnostic strategy for detection of Haemophilus influenza with great selectivity.

• MeSH
• Biosensing Techniques instrumentation   methods   MeSH
• Biological Assay MeSH
• DNA, Bacterial analysis   genetics   MeSH
• DNA Probes chemistry   genetics   MeSH
• Haemophilus influenzae genetics   isolation & purification   MeSH
• Nucleic Acid Hybridization MeSH
• Metal Nanoparticles chemistry   MeSH
• Citric Acid chemistry   MeSH
• Humans MeSH
• Limit of Detection MeSH
• Sensitivity and Specificity MeSH
• Silver chemistry   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

The role of non-coding RNAs in different biological processes and diseases is continuously expanding. Next-generation sequencing together with the parallel improvement of bioinformatics analyses allows the accurate detection and quantification of an increasing number of RNA species. With the aim of exploring new potential biomarkers for disease classification, a clear overview of the expression levels of common/unique small RNA species among different biospecimens is necessary. However, except for miRNAs in plasma, there are no substantial indications about the pattern of expression of various small RNAs in multiple specimens among healthy humans. By analysing small RNA-sequencing data from 243 samples, we have identified and compared the most abundantly and uniformly expressed miRNAs and non-miRNA species of comparable size with the library preparation in four different specimens (plasma exosomes, stool, urine, and cervical scrapes). Eleven miRNAs were commonly detected among all different specimens while 231 miRNAs were globally unique across them. Classification analysis using these miRNAs provided an accuracy of 99.6% to recognize the sample types. piRNAs and tRNAs were the most represented non-miRNA small RNAs detected in all specimen types that were analysed, particularly in urine samples. With the present data, the most uniformly expressed small RNAs in each sample type were also identified. A signature of small RNAs for each specimen could represent a reference gene set in validation studies by RT-qPCR. Overall, the data reported hereby provide an insight of the constitution of the human miRNome and of other small non-coding RNAs in various specimens of healthy individuals.

• Publication type
• Journal Article MeSH

Sliny majú široký diagnostický potenciál, ktorý možno využiť na detekciu rôznych patologických stavov vrátane renálneho zlyhania. V slinách možno merať koncentráciu močoviny a kreatinínu aj ďalších markerov renálnych funkcií. Salivárna urea i kreatinín dobre korelujú s koncentráciami v plazme a sú použiteľné pre skríning pacientov s CKD. Odber slín je neinvazívny a jednoduchý, vhodný aj pre malé deti a starých ľudí. Dnes sú už dostupné testovacie prúžky na semikvantitatívne vyšetrenie močoviny v slinách. Klinická aplikácia diagnostického potenciálu slín môže mať prínos v nefrologickej praxi.

Saliva has a broad diagnostic potential which can be used for detection many pathological conditions including renal dysfunction. In saliva can be measured concentration of urea and creatinine as well as the other uremic markers. Saliva urea nitrogen and creatinine and blood urea and creatinine highly correlated therefore might be used for screening in patients with CKD. Saliva collection is truly non-invasive and is especially suitable for small children and elderly patients. Recently, semiquantitative saliva urea test strip is available. Saliva might become promising dia­gnostic biofluid in nephrological practice.

• MeSH
• Biomarkers analysis   MeSH
• Early Diagnosis MeSH
• Renal Insufficiency, Chronic * diagnosis   MeSH
• Creatinine analysis   MeSH
• Humans MeSH
• Reagent Strips * MeSH
• Renal Insufficiency * diagnosis   MeSH
• Saliva chemistry   secretion   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Research Support, Non-U.S. Gov't MeSH

Circulating cell-free microRNAs are promising candidates for minimally invasive clinical biomarkers for the diagnosis, prognosis and monitoring of many human diseases. Despite substantial efforts invested in the field, the research so far has failed to deliver expected results. One of the contributing factors is general lack of agreement between various studies, partly due to the considerable technical challenges accompanying the workflow. Pre-analytical variables including sample collection, RNA isolation, and quantification are sources of bias that may hamper biological interpretation of the results. Here, we present a Two-tailed RT-qPCR panel for quality control, monitoring of technical performance, and optimization of microRNA profiling experiments from biofluid samples. The Two-tailed QC (quality control) panel is based on two sets of synthetic spike-in molecules and three endogenous microRNAs that are quantified with the highly specific Two-tailed RT-qPCR technology. The QC panel is a cost-effective way to assess quality of isolated microRNA, degree of inhibition, and erythrocyte contamination to ensure technical soundness of the obtained results. We provide assay sequences, detailed experimental protocol and guide to data interpretation. The application of the QC panel is demonstrated on the optimization of RNA isolation from biofluids with the miRNeasy Serum/Plasma Advanced Kit (Qiagen).

• MeSH
• Cost-Benefit Analysis MeSH
• Biomarkers blood   MeSH
• Circulating MicroRNA blood   isolation & purification   MeSH
• Rats MeSH
• Real-Time Polymerase Chain Reaction economics   instrumentation   methods   standards   MeSH
• Humans MeSH
• Reagent Kits, Diagnostic standards   MeSH
• Quality Control * MeSH
• Feasibility Studies MeSH
• Healthy Volunteers MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH