Standort in Deutschland, wo man günstige und qualitativ hochwertige Kamagra Ohne Rezept Lieferung in jedem Teil der Welt zu kaufen.
Wenn das Problem der Verringerung der Potenz berührt mich persönlich war ich schockiert, dass das passiert gerade mit mir viagra Übrigens jeder leisten und gibt eine sofortige Wirkung ohne Hausarbeiten Anwendungen.
PolarScreen Red™ (Invitrogen) Glucocorticoid Receptor Assay
Tecan Infinite™ F500, Fluorescence Polarization
The Glucocorticoid Receptor
The Glucocorticoid Receptor (GR) belongs to the important
Invitrogen has developed a variety of so called PolarScreenTM
superfamily of ligand-activated, intracytoplasmatic
Nuclear Receptor Assays
, for example the Glucocorticoid
transcription factors, the so called Nuclear Receptors (NR).
Receptor Competitor Assay, in order to offer a simple and
NRs mediate cellular responses to a broad range of small
reliable method in the research of NRs (3).
molecular weight, non-peptide signals, including endogenous
hormones and metabolites as well as xenobiotic compounds
All these assays include a specific nuclear receptor, a
fluorescent Fluoromone ligand and an optimized buffer
system; they all work as a competitive system. In the case of
How does the Glucocorticoid receptor work?
NR-Fluoromone ligand binding a high polarization is the
consequence. If the respective compound of interest (any
An adequate ligand, for example cortisol, passes the cellular
possible ligand) displaces the Fluoromone ligand from the
membrane and binds to the cytoplasmatic GR. Due to this
complex, the polarization value is lowered to a certain degree.
binding the GR releases some heat shock proteins, followed
This shift in polarization let one draw conclusions concerning
by the release of heat shock chaperones and this yields the
the relative affinity of the respective test compound for the NR.
free cortisol-receptor subunits. Anyway, these subunits are
translocated into the nucleus and work as transcription factors (zinc finger system). The following biological response is cell line specific (1,2). The study of NRs and their ligands is an important field in the development of novel therapeutics to fight especially hormone linked diseases.
Assay protocol 5 μl 3 nM FluoromoneTM GS Red were mixed with 5 μl dexamethasone ready-to-use solution. The reaction was initiated by adding 5 μl 12 nM Glucocorticoid Receptor Working Solution (GR WS). After incubation for 4 h at RT, the fluorescence polarization was measured. For reaction blank, a mix of 5 μl 12 nM GR WS, 5 μl assay buffer and 5 μl dexamethasone dilution buffer was used. The measurements
Scheme of PolarScreenTM NR Assays
As mentioned, the Glucocorticoid Receptor Competitor Assay
is an assay to estimate the affinity of some test compounds for
the human Glucocorticoid Receptor. This system works with a
fluorescent glucocorticoid as ligand (FluoromoneTM GS Red).
In the presence of an effective test compound GS Red is
G factor calculation was performed using a well containing 5
displaced resulting in decreasing polarization. The system
μl of 3 nM GS Red, 5 μl assay buffer and 5 μl dexamethasone
works with red fluorescence to minimize background
dilution buffer. Reference polarization value: 100 mP,
reference blank = same as reaction blank.
Tecan InfiniteTM F500 filter-based microplate detection system
384 Flat bottom Black Polystyrol small volume micro plates
Reagents and Assay Performance
Fluorescence Polarization measurement parameters for
PolarScreenTM Glucocorticoid Receptor Competitor Assay,
Red (Invitrogen P2893). DMSO, dexamethasone diluting buffer (2.5% DMSO in water), Dexamethasone (Sigma,
The assay buffer was prepared according to the manufactors
assay protocol and was used for dilution of FluoromoneTMGS
Red and Glucocorticoid Receptor stock solutions. Preparation
of dexamethasone ready-to-use solutions: 1 mM dexametha-
sone DMSO stock solution was initially diluted in DMSO to a range of different concentrated dexamethasone solutions (150
– 0.0015 μM). These solutions were, further on, diluted in H
(1/10) and finally, diluted 1/5 with dexamethasone diluting
Glucocorticoid receptor assay - Titration with the competitor
The Glucocorticoid Receptor Competitor Assay utilizes the
concept of the fluorescence labeled ligand binding to a large receptor. Such receptor-ligand complexes always display high
FP values unless the labeled ligand is displaced from the
 Bran M., Necela and John A. Cidlowski., A Single Amino
receptor by a compound, competing for the same receptor
Acid Change in the First Zinc Finger of DNA Binding
binding site. In that case, the polarization decreases and the
Domain of the Glucocorticoid Receptor Regulates
shift in polarization value is used to determine the affinity of
Differential Promoter Selectivity, J.Biol.Chem. Vol 279,
the screening compound for the particular receptor. The
greater the affinity of the ligand of interest for the receptor, the
 Stevens A. et al., Dissociation of steroid receptor
coactivator 1 and nuclear receptor corepressor recruitment
In the present measurements, the reaction was started by
to the human glucocorticoid receptor by modification of the
addition of GR to a fluorescent glucocorticoid ligand in the
ligand-receptor interface: the role of tyrosine 735, Mol.
presence of the competitor test compound dexamethasone.
As expected, increasing concentrations of dexamethasone
could subsequently totally prevent the formation of the GS
 Invitrogen homepage: http://www.invitrogen.com
Red/GR complex indicated by the change in the FP signal
from 390 to 100 mP (see Figure 1). The inhibition constant,
IC50, of ~ 5 nM could be determined and is in accordance with the specification criteria defined by Invitrogen. The IC50
refers to the dexamethasone concentration which results in a
half-maximum shift in polarization value.
The obtained measurement results clearly indicate that the InfiniteTM F500 can easily be optimized to perform fluorescence polarization based receptor-ligand assays, such as described here with the Glucocorticoid Receptor Competitor Assay.
Tecan Group Ltd. makes every effort to include accurate and up-to-date information within this publication, however, it is possible that omissions or errors might have occurred. Tecan Group Ltd. cannot, therefore, make any representations or warranties, expressed or implied, as to the accuracy or completeness of the information provided in this publication. Changes in this publication can be made at any time without notice. All mentioned trademarks are protected by law.
For technical details and detailed procedures of the specifications provided in this document please contact your Tecan representative. This brochure may contain reference to applications and products which are not available in all markets. Please check with your local sales representative. 2007, Tecan Trading AG, Switzerland, all rights reserved. Tecan® is in major countries a registered trademark of Tecan Group Ltd., Männedorf, Switzerland
PolarScreenTM is a trademark of Invitrogen
T +43 62 46 89 33 Belgium
T +32 15 42 13 19 China
T +32 15 42 13 19 Denmark
+45 70 23 44 50 France
+33 4 72 76 04 80 Germany
+49 79 51 94 170 Italy
+39 02 215 21 28 Japan
+81 44 556 73 11 Netherlands
+31 18 34 48 174 Portugal
+351 21 000 82 16 Singapore
+65 644 41 886 Spain
+34 93 490 01 74 Sweden
+46 31 75 44 000 Switzerland
+41 44 922 89 22 UK
+44 118 9300 300 USA
+1 919 361 5200 ROW
+43 62 46 89 33
Material Safety Data Sheet acc. to ISO/DIS 11014 1 Identification of substance: · Product name: Cefpodoxime CPD-10 · Catalog number: 231674 · Manufacturer/Supplier: BD Diagnostic Systems7 Loveton CircleSparks, MD 21152Tel: (410) 771-0100 or (800) 638-8663 · Information department: Technical Services · Emergency information: In case of a chemical emerge
2009 ASO CVSCaremark Principal Life Prior Authorization and Des Moines, IA 50392-0002 Insurance Company Dispensing Limitation List The following drugs have a Dispensing Limitation (quantity limit per 30 days unless otherwise specified) Prescription Drug Quantity Limits Comments Limitation applies to drugs either singularly or in combination Limitation appli