In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.
- MeSH
- autofagie * fyziologie MeSH
- autofagozomy MeSH
- biologické markery MeSH
- biotest normy MeSH
- lidé MeSH
- lyzozomy MeSH
- proteiny spojené s autofagií metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
- Research Support, U.S. Gov't, Non-P.H.S. MeSH
- směrnice MeSH
Lipases are enzymes responsible for the conversion of triglycerides and other esterified substrates, they are involved in the basic metabolism of a wide number of organisms, from a simple microorganism and to mammals. They also have broad applicability in many fields from which industrial biotechnology, the production of cleaning agents, and pharmacy are the most important. The use of lipases in analytical chemistry where it can serve as a part of biosensors or bioassays is an application of growing interest and has become another important use. This review is focused on the description of lipases chemistry, their current applications and the methods for their assay measurement. Examples of bioassays and biosensors, including their physical and chemical principles, performance for specific substrates, and discussion of their relevance, are given in this work.
A European round robin test according to ISO 5725-2 was conceptually prepared, realised, and evaluated. The aim was to determine the inter-laboratory variability of the overall process for the ecotoxicological characterization of construction products in eluates and bioassays. To this end, two construction products BAM-G1 (granulate) and HSR-2 (roof sealing sheet), both made of EPDM polymers (rubber), were selected. The granular construction product was eluted in a one stage batch test, the planar product in the Dynamic Surface Leaching test (DSLT). A total of 17 laboratories from 5 countries participated in the round robin test: Germany (12), Austria (2), Belgium (1), Czech Republic (1) and France (1). A test battery of four standardised ecotoxicity tests with algae, daphnia, luminescent bacteria and zebrafish eggs was used. As toxicity measures, EC50 and LID values were calculated. All tests, except the fish egg test, were basically able to demonstrate toxic effects and the level of toxicity. The reproducibility of test results depended on the test specimens and the test organisms. Generally, the variability of the EC50 or LID values increased with the overall level of toxicity. For the very toxic BAM-G1 eluate a relative high variability of CV = 73%-110% was observed for EC50 in all biotests, while for the less toxic HSR-2 eluate the reproducibility of EC50 varied with sensitivity: it was very good (CV = 9.3%) for the daphnia test with the lowest sensitivity, followed by the algae test (CV = 36.4%). The luminescent bacteria test, being the most sensitive bioassay for HSR-2 Eluate, showed the highest variability (CV = 74.8%). When considering the complex overall process the reproducibility of bioassays with eluates from construction products was acceptable.
- MeSH
- Bacteria účinky léků MeSH
- biotest metody normy MeSH
- chemické látky znečišťující vodu analýza toxicita MeSH
- dánio pruhované MeSH
- Daphnia účinky léků MeSH
- ekotoxikologie metody normy MeSH
- elastomery toxicita MeSH
- ethyleny toxicita MeSH
- guma toxicita MeSH
- Heterokontophyta účinky léků MeSH
- odchylka pozorovatele MeSH
- reprodukovatelnost výsledků MeSH
- testy toxicity metody normy MeSH
- vejce MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
OBJECTIVES: Glucose is an important diagnostic biochemical marker of diabetes but also for organophosphates, carbamates, acetaminophens or salicylates poisoning. Hence, innovation of accurate and fast detection assay is still one of priorities in biomedical research. METHODS: Glucose sensor based on magnetic particles (MPs) with immobilized enzymes glucose oxidase (GOx) and horseradish peroxidase (HRP) was developed and the GOx catalyzed reaction was visualized by a smart-phone-integrated camera. RESULTS: Exponential decay concentration curve with correlation coefficient 0.997 and with limit of detection 0.4 mmol/l was achieved. Interfering and matrix substances were measured due to possibility of assay influencing and no effect of the tested substances was observed. Spiked plasma samples were also measured and no influence of plasma matrix on the assay was proved. CONCLUSIONS: The presented assay showed complying results with reference method (standard spectrophotometry based on enzymes glucose oxidase and peroxidase inside plastic cuvettes) with linear dependence and correlation coefficient 0.999 in concentration range between 0 and 4 mmol/l. On the grounds of measured results, method was considered as highly specific, accurate and fast assay for detection of glucose.
- MeSH
- biosenzitivní techniky metody MeSH
- biotest metody normy MeSH
- chytrý telefon využití MeSH
- enzymy imobilizované metabolismus MeSH
- glukosaoxidasa metabolismus MeSH
- křenová peroxidasa metabolismus MeSH
- krevní glukóza metabolismus MeSH
- lidé MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- MeSH
- autofagie * fyziologie MeSH
- biotest metody normy MeSH
- lidé MeSH
- počítačová simulace MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Research Support, N.I.H., Extramural MeSH
- směrnice MeSH
BACKGROUND: Efforts to replace the rabbit skin irritation test have been underway for many years, encouraged by the EU Cosmetics Directive and REACH. Recently various in vitro tests have been developed, evaluated and validated. OBJECTIVE: A key difficulty in confirming the validity of in vitro methods is that animal data are scarce and of limited utility for prediction of human effects, which adversely impacts their acceptance. This study examines whether in vivo or in vitro data most accurately predicted human effects. METHODS: Using the 4-hr human patch test (HPT) we examined a number of chemicals whose EU classification of skin irritancy is known to be borderline, or where in vitro methods provided conflicting results. RESULTS: Of the 16 chemicals classified as irritants in the rabbit, only five substances were found to be significantly irritating to human skin. Concordance of the rabbit test with the 4-hr HPT was only 56%, whereas concordance of human epidermis models with human data was 76% (EpiDerm) and 70% (EPISKIN). CONCLUSIONS: The results confirm observations that rabbits overpredict skin effects in humans. Therefore, when validating in vitro methods, all available information, including human data, should be taken into account before making conclusions about their predictive capacity.
- MeSH
- alternativy testů na zvířatech normy MeSH
- biotest normy MeSH
- dráždivé látky škodlivé účinky diagnostické užití MeSH
- falešně pozitivní reakce MeSH
- kosmetické přípravky škodlivé účinky diagnostické užití MeSH
- králíci MeSH
- kůže účinky léků MeSH
- lidé MeSH
- náplasťové testy normy MeSH
- prediktivní hodnota testů MeSH
- testy kožní dráždivosti normy MeSH
- zvířata MeSH
- Check Tag
- králíci MeSH
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
- validační studie MeSH
The in vivo measurement of the activity deposited in the skeleton is a very useful source of information on human internal contaminations with transuranic elements, e.g. americium 241, especially for long time periods after intake. Measurements are performed on the skull or the larger joints such as the knee or elbow. The paper deals with the construction of an anthropomorphic numerical phantom based on CT scans, its potential for calibration and the estimation of the uncertainties of the detection system. The density of bones, activity distribution and position of the detectors were changed in individual simulations in order to estimate their effects on the result of the measurement. The results from simulations with the numerical phantom were compared with the results of physical phantoms.
- MeSH
- antropometrie metody MeSH
- biologické modely MeSH
- biotest metody normy MeSH
- dávka záření MeSH
- druhová specificita MeSH
- financování organizované MeSH
- hlava fyziologie MeSH
- lidé MeSH
- orgánová specificita MeSH
- počítačová simulace MeSH
- radiometrie metody MeSH
- relativní biologická účinnost MeSH
- senzitivita a specificita MeSH
- tkáňová distribuce MeSH
- uran farmakokinetika MeSH
- Check Tag
- lidé MeSH
