Methylation-Blocked Enzymatic Recycling Amplification for Highly Sensitive Fluorescence Sensing of DNA Methyltransferase Activity.
Methylation-blocked enzymatic recycling amplification for highly sensitive fluorescence sensing of DNA methyltransferase activity.
Filed under: Drug and Alcohol Rehabilitation
Analyst. 2012 Nov 8;
Chen F, Zhao Y
Herein, using DNA adenine methylation (Dam) methyltransferase (MTase) as a model analyte, a novel fluorescence sensing strategy was developed for facile, rapid and highly sensitive detection of the activity and inhibition of the target based on methylation-blocked enzymatic recycling amplification. In this sensing system, nicking endonuclease Nt.AlwI with the methylation-sensitive property was selected to achieve signal amplification. In addition, a DNA heteroduplex probe is specially designed to contain the recognition sequences for both Dam MTase and Nt.AlwI. In the absence of Dam MTase, Nt.AlwI cleaves the DNA heteroduplex at only the top strand. At the reaction temperature, the cleaved heteroduplex is unstable and readily separates. The released bottom strand can hybridize with the molecular beacons (MB) and subsequently trigger Nt.AlwI-mediated recycling cleavage of MBs, providing a dramatically amplified fluorescence signal. However, when the heteroduplex is methylated by Dam MTase, the cleaving operation is blocked, resulting in an inconspicuous fluorescence enhancement. Unlike existing signal amplified assays which use at least two enzymes, only one is involved in this amplified strategy. Under optimized conditions, the sensing system reveals a detection limit of 0.05 U mL(-1) in a short assay time (65 min), which is much superior to all presently reported methods except for two electrochemical biosensors (0.04 U mL(-1)). Furthermore, the application of the assay in human serum and screening of Dam MTase inhibition were demonstrated with satisfactory results. Overall, the proposed sensing system shows great potential for further application in biological research, early clinical diagnosis and designed drug therapy.
HubMed – drug
[Pharmacoepidemiological study of the use of psychotropics in hospitalized patients].
Filed under: Drug and Alcohol Rehabilitation
Vertex. 2012 Mar-Apr; 23(102): 98-103
Bolaños R, Bazerque PM, Nordaby M, Miceli MB, Saxton R, Pastore F, Estrin MA
Objective: To assess consumption and the exposition of patients admitted to the Universitary Hospital of the Interamerican Open University (UAI), between October and December of 2007-2009. Methods: Descriptive observational study. The information was obtained of the Pharmacia Sector. We analyze the dispensations (Units); the exposition was evaluated using the defined daily dose per 100 beds/day. We analyzed how many of the psychotropics used were considerated essentials drugs for the WHO. We assessed the adverse drug reactions registered. Results: The psychopharmacological drugs represented the 43%, 41%, and the 44% (period 2007-2009) of the drugs used for the Central Nervous System. The benzodiazepines represented more of the 50% of the psychopharmacological drugs used. The exposure was respectively of 57%, 66%, and 40% (for all the patients admitted to the hospital on period 2007-2009). The essential medicines used fluctuate between 41% and 48%. From 32 adverse reactions connected with the Central Nervous System, 20 (62.5%) were attributed to psychotropics. Conclusions: 1. We established the quantity of the psychotropics dispensations, as well as the level of the exposure. 2. The psychotropic utilization of the essentials medicines list of the WHO was above 40%.
HubMed – drug
Stratification substantially reduces behavioral variability in the hypoxic-ischemic stroke model.
Filed under: Drug and Alcohol Rehabilitation
Brain Behav. 2012 Sep; 2(5): 698-706
Pollak J, Doyle KP, Mamer L, Shamloo M, Buckwalter MS
Stroke is the most common cause of long-term disability, and there are no known drug therapies to improve recovery after stroke. To understand how successful recovery occurs, dissect candidate molecular pathways, and test new therapies, there is a need for multiple distinct mouse stroke models, in which the parameters of recovery after stroke are well defined. Hypoxic-ischemic stroke is a well-established stroke model, but behavioral recovery in this model is not well described. We therefore examined a panel of behavioral tests to see whether they could be used to quantify functional recovery after hypoxic-ischemic stroke. We found that in C57BL/6J mice this stroke model produces high mortality (approximately one-third) and variable stroke sizes, but is fast and easy to perform on a large number of mice. Horizontal ladder test performance on day 1 after stroke was highly and reproducibly correlated with stroke size (P < 0.0001, R(2) = 0.7652), and allowed for functional stratification of mice into a group with >18% foot faults and 2.1-fold larger strokes. This group exhibited significant functional deficits for as long as 3 weeks on the horizontal ladder test and through the last day of testing on automated gait analysis (33 days), rotarod (30 days), and elevated body swing test (EBST) (36 days). No deficits were observed in an automated activity chamber. We conclude that stratification by horizontal ladder test performance on day 1 identifies a subset of mice in which functional recovery from hypoxic-ischemic stroke can be studied.
HubMed – drug
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