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An overview of technical considerations for Western blotting applications to physiological research.Scandinavian Journal of Medicine &... Jan 2017The applications of Western/immunoblotting (WB) techniques have reached multiple layers of the scientific community and are now considered routine procedures in the... (Review)
Review
The applications of Western/immunoblotting (WB) techniques have reached multiple layers of the scientific community and are now considered routine procedures in the field of physiology. This is none more so than in relation to skeletal muscle physiology (i.e., resolving the mechanisms underpinning adaptations to exercise). Indeed, the inclusion of WB data is now considered an essential aspect of many such physiological publications to provide mechanistic insight into regulatory processes. Despite this popularity, and due to the ubiquitous and relatively inexpensive availability of WB equipment, the quality of WB in publications and subsequent analysis and interpretation of the data can be variable, perhaps resulting in spurious conclusions. This may be due to poor laboratory technique and/or lack of comprehension of the critical steps involved in WB and what quality control procedures should be in place to ensure robust data generation. The present review aims to provide a detailed description and critique of WB procedures and technicalities, from sample collection through preparation, blotting and detection, to analysis of the data collected. We aim to provide the reader with improved expertise to critically conduct, evaluate, and troubleshoot the WB process, to produce reproducible and reliable blots.
Topics: Blotting, Western; Data Accuracy; Humans; Muscle, Skeletal; Physiology; Specimen Handling
PubMed: 27263489
DOI: 10.1111/sms.12702 -
Virchows Archiv : An International... Aug 2021The term "biobanking" is often misapplied to any collection of human biological materials (biospecimens) regardless of requirements related to ethical and legal issues... (Review)
Review
The term "biobanking" is often misapplied to any collection of human biological materials (biospecimens) regardless of requirements related to ethical and legal issues or the standardization of different processes involved in tissue collection. A proper definition of biobanks is large collections of biospecimens linked to relevant personal and health information (health records, family history, lifestyle, genetic information) that are held predominantly for use in health and medical research. In addition, the International Organization for Standardization, in illustrating the requirements for biobanking (ISO 20387:2018), stresses the concept of biobanks being legal entities driving the process of acquisition and storage together with some or all of the activities related to collection, preparation, preservation, testing, analysing and distributing defined biological material as well as related information and data. In this review article, we aim to discuss the basic principles of biobanking, spanning from definitions to classification systems, standardization processes and documents, sustainability and ethical and legal requirements. We also deal with emerging specimens that are currently being generated and shaping the so-called next-generation biobanking, and we provide pragmatic examples of cancer-associated biobanking by discussing the process behind the construction of a biobank and the infrastructures supporting the implementation of biobanking in scientific research.
Topics: Accreditation; Biological Specimen Banks; Biomedical Research; Guidelines as Topic; Humans; Policy Making; Precision Medicine; Specimen Handling; Stakeholder Participation; Terminology as Topic
PubMed: 34255145
DOI: 10.1007/s00428-021-03151-0 -
Clinical Microbiology and Infection :... Apr 2020Blood culture contamination carries risks for patients, such as unnecessary antimicrobial therapy and other additional hazards and costs. One method shown to be... (Randomized Controlled Trial)
Randomized Controlled Trial
OBJECTIVES
Blood culture contamination carries risks for patients, such as unnecessary antimicrobial therapy and other additional hazards and costs. One method shown to be effective in reducing contamination is initial blood specimen diversion during collection. We hypothesized that initial blood specimen diversion without a designated device or procedure would suffice for reduction in blood culture contamination rate.
METHODS
From 1 September 2017 through to 6 September 2018, we conducted a randomized controlled trial to assess the effect of an initial-specimen diversion technique (ISDT) on the rate of blood-culture contamination by changing the order of sampling using regular vacuum specimen tubes instead of commercially available sterile diversion devices. We included adults from whom the treating physician planned to take blood cultures and additional blood chemistry tests. Additionally, we evaluated the potential economic benefits of an ISDT. This was a researcher-initiated trial, Clinicaltrials.gov NCT03088865.
RESULTS
In all, 756 patients were enrolled. This method, compared with the standard procedure in use at our medical centre, reduced contamination by 66% (95% CI 17%-86%), from 20/400 (5%) with the standard method to 6/356 (1.6%) with the ISDT, without compromising detection of true bloodstream infection and at no additional cost. Hospital-wide implementation of ISDT was associated with a 1.1% saving in hospitalization days.
CONCLUSIONS
We offer this novel approach as a simple, cost-effective measure to reduce risks to patient safety from contaminated blood cultures, without the need for using costly devices.
Topics: Adolescent; Adult; Aged; Aged, 80 and over; Blood Culture; Blood Specimen Collection; Costs and Cost Analysis; Female; Hospitals; Humans; Male; Middle Aged; Prospective Studies; Specimen Handling; Young Adult
PubMed: 31539635
DOI: 10.1016/j.cmi.2019.09.005 -
EMBO Molecular Medicine Nov 2019Plasma and serum are rich sources of information regarding an individual's health state, and protein tests inform medical decision making. Despite major investments, few...
Plasma and serum are rich sources of information regarding an individual's health state, and protein tests inform medical decision making. Despite major investments, few new biomarkers have reached the clinic. Mass spectrometry (MS)-based proteomics now allows highly specific and quantitative readout of the plasma proteome. Here, we employ Plasma Proteome Profiling to define quality marker panels to assess plasma samples and the likelihood that suggested biomarkers are instead artifacts related to sample handling and processing. We acquire deep reference proteomes of erythrocytes, platelets, plasma, and whole blood of 20 individuals (> 6,000 proteins), and compare serum and plasma proteomes. Based on spike-in experiments, we determine sample quality-associated proteins, many of which have been reported as biomarker candidates as revealed by a comprehensive literature survey. We provide sample preparation guidelines and an online resource ( www.plasmaproteomeprofiling.org) to assess overall sample-related bias in clinical studies and to prevent costly miss-assignment of biomarker candidates.
Topics: Bias; Biomarkers; Female; Germany; Healthy Volunteers; Humans; Male; Plasma; Proteome; Proteomics; Specimen Handling
PubMed: 31566909
DOI: 10.15252/emmm.201910427 -
Current Opinion in Structural Biology Jun 2024Specimen preparation is a critical but challenging step in high-resolution cryogenic electron microscopy (cryo-EM) structural analysis of macromolecules. In the past... (Review)
Review
Specimen preparation is a critical but challenging step in high-resolution cryogenic electron microscopy (cryo-EM) structural analysis of macromolecules. In the past decade, graphene has gained much recognition as the supporting substrate to optimize cryo-EM specimen preparation. It improves macromolecule embedding in ice, reduces beam-induced motion, while imposing negligible background noise. Various types of graphene-coated cryo-EM grids were implemented to improve the robustness and efficiency of specimen preparation. Graphene functionalization by different means has been proved specifically useful in addressing challenges related to the air-water interface (AWI), such as preferential orientation and sample denaturation. Graphene sandwich specimen preparation sets a new direction to explore in cryo-EM analysis of biological specimens. In this review, we discuss the current challenges and future prospects of graphene application in cryo-EM analysis of macromolecules.
Topics: Cryoelectron Microscopy; Graphite; Macromolecular Substances; Specimen Handling
PubMed: 38688075
DOI: 10.1016/j.sbi.2024.102823 -
The Urologic Clinics of North America Nov 2015Pediatric urinary tract infection (UTI) costs the health care system more than $180 million annually, and accounts for more than 1.5 million clinician visits per year.... (Review)
Review
Pediatric urinary tract infection (UTI) costs the health care system more than $180 million annually, and accounts for more than 1.5 million clinician visits per year. Accurate and timely diagnosis of these infections is important for determining appropriate treatment and preventing long-term complications such as renal scarring, hypertension, and end-stage renal disease. After the first 12 months, girls are more likely to be diagnosed with a UTI. About half of boys with UTI are diagnosed within the first 12 months of life. Diagnosis of UTI is made based on history and examination findings and confirmed by urine testing.
Topics: Adolescent; Anti-Bacterial Agents; Child; Child, Preschool; Diagnostic Imaging; Female; Humans; Infant; Male; Medical History Taking; Physical Examination; Specimen Handling; Urinalysis; Urinary Tract Infections
PubMed: 26475948
DOI: 10.1016/j.ucl.2015.05.011 -
Journal of Virological Methods Mar 2021The worldwide COVID-19 pandemic outburst has caused a serious public health issue with increasing needs of accurate and rapid diagnostic and screening testing. This... (Review)
Review
The worldwide COVID-19 pandemic outburst has caused a serious public health issue with increasing needs of accurate and rapid diagnostic and screening testing. This situation requires an optimized management of the chemical reagents, the consumables, and the human resources, in order to respond accurately and effectively, controlling the spread of the disease. Testing on pooled samples maximizes the number of tested samples, by minimizing the time and the lab supplies needed. The general conceptualization of the pooling method is based on mixing samples together in a batch. Individual testing is needed only if a specific pool exhibits a positive result. The development of alternative hybrid methods, based on "in house" protocols, utilizing commercially available consumables, in combination with a reliable pooling method would provide a solution, focusing on the better exploitation of the personnel and the lab supplies, allowing for rapid screening of a population in a reasonably short time.
Topics: COVID-19; COVID-19 Testing; Diagnostic Tests, Routine; Humans; Pandemics; SARS-CoV-2; Specimen Handling
PubMed: 33316285
DOI: 10.1016/j.jviromet.2020.114044 -
British Dental Journal Jun 2022
Topics: Female; Humans; Papillomaviridae; Specimen Handling; Vaginal Smears
PubMed: 35689043
DOI: 10.1038/s41415-022-4361-0 -
The Veterinary Clinics of North... Jan 2017Synovial fluid analysis is a key component of the minimum database needed to diagnose and manage primary and secondary articular joint disorders. Unfortunately,... (Review)
Review
Synovial fluid analysis is a key component of the minimum database needed to diagnose and manage primary and secondary articular joint disorders. Unfortunately, preanalytical variables can drastically alter samples submitted for evaluation to veterinary laboratories and it is considered the stage at which most laboratory error occurs. This article addresses common sources of preanalytical variability and error that are seen in veterinary medicine. With consistent quality control and reporting of specimens, downstream clinical decision making and management of patients can be accelerated and improved.
Topics: Animals; Cytological Techniques; Specimen Handling; Synovial Fluid
PubMed: 27720280
DOI: 10.1016/j.cvsm.2016.07.007 -
Trends in Ecology & Evolution Nov 2021Historical DNA (hDNA), obtained from museum and herbarium specimens, has yielded spectacular new insights into the history of organisms. This includes documenting... (Review)
Review
Historical DNA (hDNA), obtained from museum and herbarium specimens, has yielded spectacular new insights into the history of organisms. This includes documenting historical genetic erosion and extinction, discovering species new to science, resolving evolutionary relationships, investigating epigenetic effects, and determining origins of infectious diseases. However, the development of best-practices in isolating, processing, and analyzing hDNA remain under-explored, due to the substantial diversity of specimen preparation types, tissue sources, archival ages, and collecting histories. Thus, for hDNA to reach its full potential, and justify the destructive sampling of the rarest specimens, more experimental work using time-series collections, and the development of improved methods to correct for data asymmetries and biases due to DNA degradation are required.
Topics: Biological Evolution; DNA; Museums; Sequence Analysis, DNA; Specimen Handling
PubMed: 34456066
DOI: 10.1016/j.tree.2021.07.009