Navigation Links
The use of CypHer5E and the IN Cell Analyzer 1000 for live-cell receptor internalization studies

mperature for 15 min.

7. Add 50 µl Krebs-Ringer buffer containing EGF and incubate at 37 °C for 30 min.

8. Image on the IN Cell Analyzer 1000 (20× objective), exposure time 300 ms/field, 620 nm excitation and 700 nm emission for CypHer5E; exposure time of 500 ms/field 360 nm excitation and 535 nm emission for Hoechst.

9. Analyze the images using the Granularity Analysis Module.


Results
IN Cell Analyzer 1000 images of the live-cell β-2 adrenergic assay show agonist induced internalization of CypHer5E into acidic endosomes (Fig 1). Although trafficking destinations are dependent on the receptor, similar patterning was observed with the δ-opioid and EGF receptors (data not shown). In the absence of ligand activation by an agonist, there is very little cell-associated fluorescence (Fig 1a). Gray-scale images show a perinuclear clustering of fluorescent granules in the red channel, which results in an increase in the red signal intensity (Fig 1b). A fused image showing the nucleus (blue) and CypHer5E (red) signal demonstrates the juxtanuclear localization of CypHer5E, indicative of a recycling endosome compartment (Fig 1c).

Agonist-induced internalization of CypHer5E-labeled antibodies is concentration dependent. The internalization was quantitated using the IN Cell Analyzer 1000 Granularity Analysis Module. The analysis routine identifies, counts, and analyzes the grain-like CypHer5E fluorescent structures within cells. A sigmoidal dose response curve was obtained for each agonist by non-linear regression analysis. Dose response curves for the β-2 adrenergic, δ-opioid, and EGF receptors are shown in Figure 2. Calculated EC50 values are comparable to values previously obtained with CypHer5 dye and with other literature values (6).

Receptors of interest will be expressed at the cell membrane at va
'"/>

Source:


Page: All 1 2 3 4 5 6

Related biology technology :

1. Screening for potential beta 2-adrenergic receptor antagonists using CypHer5E and IN Cell Analyzer 1000
2. Analysis of in vitro micronucleus assays using the IN Cell Analyzer 3000 Micronuclei Formation Analysis Module
3. Screening for β2-adrenergic receptor agonists using the pH-sensitive dye,CypHer5, and the IN Cell Analyzer 3000
4. Identification of cells in S phase using the Cell Proliferation Fluorescence Kit and IN Cell Analyzer 1000
5. Rapid, High Throughput Mutation Discovery Using Fluorescent SSCP on the ABI PRISM 3100 and 3100Avant Genetic Analyzers
6. The development of a CXCR2 chemokine receptorbinding assay using the LEADseeker Multimodality Imaging System
7. The development of a melanocortin MC4 receptor binding assay using the LEADseeker Multimodality Imaging System
8. The development of a melanocortin MC5 receptor binding assay using the LEADseeker Multimodality Imaging System
9. The development of a calcitonin-gene related peptide (CGRP) receptor-binding assay using the LEADseeker Multimodality Imaging System
10. The use of CypHer5 for receptor internalization studies in both a range of GPCRs and a non-GPCR
11. Development of a histamine H1 receptor binding assay using the LEADseeker Multimodality Imaging System
Post Your Comments:
(Date:9/19/2014)... 2014 Biotech companies make announcements ... drug systems and advanced Laboratory Instrumentation. Companies in focus ... BDX ), Thermo Fisher Scientific Inc. (NYSE: ... RGDO ), Sangamo Biosciences Inc. (NASDAQ: ... Pressure BioSciences, Inc. (OTCQB: PBIO) ("PBI" or the "Company") ...
(Date:9/19/2014)... -- Pfenex Inc. (NYSE MKT: PFNX), a clinical-stage biotechnology ... to manufacture proteins including biosimilar therapeutics, today announced that ... Annual NewsMakers in the Biotech Industry conference in ... chief executive officer of Pfenex, will provide an overview ... Friday, September 26, 2014 at 10:00 a.m. EDT/7:00 a.m. ...
(Date:9/19/2014)... September 19, 2014 Barnhardt Manufacturing ... Label for its Organic Cotton . The USDA ... of renewable biobased ingredients meets or exceeds levels set ... composed in whole or in significant part of agricultural, ... allow our Organic cotton to be immediately identified as ...
(Date:9/19/2014)... Dublin ... announced the addition of the  "Micro Market Monitor ... offering.       (Logo: http://photos.prnewswire.com/prnh/20130307/600769) , , ... fastest-growing segments in the chromatography market. The market ... expected to reach $2.0 billion by 2019, at ...
Breaking Biology Technology:Medical Instrumentation Automation Advances for Biological Sample Preparation - Company Expects Q4 Revenue Impact from Newest Instrument Systems 2Medical Instrumentation Automation Advances for Biological Sample Preparation - Company Expects Q4 Revenue Impact from Newest Instrument Systems 3Medical Instrumentation Automation Advances for Biological Sample Preparation - Company Expects Q4 Revenue Impact from Newest Instrument Systems 4Medical Instrumentation Automation Advances for Biological Sample Preparation - Company Expects Q4 Revenue Impact from Newest Instrument Systems 5Medical Instrumentation Automation Advances for Biological Sample Preparation - Company Expects Q4 Revenue Impact from Newest Instrument Systems 6Medical Instrumentation Automation Advances for Biological Sample Preparation - Company Expects Q4 Revenue Impact from Newest Instrument Systems 7Pfenex to Present at Upcoming Industry Conference on September 26 2Barnhardt Manufacturing Company Earns USDA Certified Biobased Product Certification and Label for Organic Cotton 2Micro Market Monitor : Global Pre-packed Chromatography Columns 2
... NEOG ) has expanded its line of ... quickest test available for tetracycline residues,in milk. The ... treat,bovine mastitis, and includes tetracycline, chlortetracycline, and,oxytetracycline., ... the company,s widely used BetaStar(R) test for the ...
... Philadelphia Area -, PHILADELPHIA, Feb. 13 ... announced today the appointment of Chris Tihansky as,its ... Paris-based Mauna,Kea Technologies and develops and markets Cellvizio(R), ... care by eliminating the,need for unnecessary biopsies and ...
... Advance Nanotech, Inc.,(OTC Bulletin Board: AVNA), a leader in ... special meeting of shareholders held at,Company headquarters in New ... increase the authorized number of shares of Capital stock ... shares,present at the meeting were voted in favor of ...
Cached Biology Technology:Neogen Launches Quickest Test Available for Tetracyclines in Milk 2Chris Tihansky Appointed President of In Vivo Cellular Imaging Leader Cellvizio Inc. 2Advance Nanotech Announces Shareholder Proxy Vote Results 2Advance Nanotech Announces Shareholder Proxy Vote Results 3
(Date:9/19/2014)... professor will study the unique bioelectric signaling system of ... people,s health. The OU researcher is working to ... produce the signals used to map the world around ... rate of 500-600 discharges a second throughout their lives. ... are extreme, but necessary for survival. , "There is ...
(Date:9/19/2014)... legume plants regulate their symbiotic relationship with soil ... through the plant structure from leaves into the ... in the roots. This collaborative study was conducted ... Biology, the Graduate University for Advanced Studies (SOKENDAI), ... in Japan. , Legumes, an important plant ...
(Date:9/18/2014)... ANTONIO, September 18, 2014 New research into the ... causes a severe hemorrhagic disease in humans similar to ... essential for CCHFV infection. This discovery has the potential ... the pathogen. , The research, reported in ... Pathogens and conducted by scientists at the Texas ...
Breaking Biology News(10 mins):For legume plants, a new route from shoot to root 2Research milestone in CCHF virus could help identify new treatments 2
... engineers have proposed a new environmentally friendly process for ... - potentially available from agricultural and forest waste, providing ... transportation sector." , The new approach modifies conventional methods ... from a "carbon-free" energy source, such as solar or ...
... study in PLoS Medicine has shown that children who ... children, and who have antigens (protein fragments from the ... infectious virus in their blood. Margaret Conner and colleagues ... obtained from hospitalized children with gastroenteritis and compared them ...
... transcendent entity answer prayer? , The answer, according to ... March journal Research on Social Work Practice, is “yes.?David ... the College of Human Services at Arizona State University, ... the effects of intercessory prayer ?or prayer that is ...
Cached Biology News:New biofuels process promises to meet all US transportation needs 2New biofuels process promises to meet all US transportation needs 3New biofuels process promises to meet all US transportation needs 4Does God answer prayer? ASU research says 'yes' 2
...
...
MultiWash III w/ 12-portStandard Manifold...
... Sepharose High Performance, 5 ml. The ... in HiTrap Streptavidin columns for fast, ... packing into Tricorn 5/50 GL, XK ... scale purifications are required.Extremely useful for ...
Biology Products: