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

>
Materials


Other materials required
HEK293 cell line expressing VSV-G epitope tagged β2-adrenergic receptor

HEK293 cell line expressing VSV-G epitope tagged δ-opioid receptor

HEK293 cell line expressing EGF receptor

HEK Culture medium: MEM (Sigma M2279) containing:
10% (v/v) FBS
Geneticin G418, 200 µg/ml (Sigma)
L-glutamine (Gibco)
Non-essential amino acids (Sigma)

Assay medium: MEM, phenol red free (Invitrogen 51200-038), or Krebs-Ringer buffer (see method)

Agonists: Isoproterenol, DADLE ([D-Ala2, D-Leu5]-Enkephalin), EGF (epidermal growth factor, murine submaxillary gland) (Sigma)

Hoechst 33342 (Molecular Probes)

Rat anti-human EGFR (Serotec, MCA1784)

Phosphate-buffered saline (PBS) solution

Poly-D-lysine, 5 mg (Sigma P6407)

96-well assay plate (Packard Viewplate)


Methods
Assay for β2-adrenergic and δ-opioid receptor internalization

1. Pre-coat 96-well plates with poly-D-lysine, recommended for HEK293 cell lines.

2. Seed cells at a density of ~ 16 000 cells/well.

3. Incubate ~ 48 h at 37 °C with 5% CO2.

4. Replace the culture medium with 100 µl/well of fresh MEM (phenol red free) containing 2.5–5 µg/ml CypHer5E anti-VSV-G antibody and 2.5 µM Hoechst.

5. Incubate at room temperature for 15 min.

6. Add 50 µl assay medium containing agonist and incubate at 37 °C for 30 min.

7. 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 o
'"/>

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:9/2/2015)... Wellesley, Mass. (PRWEB) , ... September 02, 2015 ... ... that includes development of botanical and plant-derived compounds. BCC Research reveals in ... drugs past 2020 because this market defines the category and generates the most ...
(Date:9/2/2015)... Calif. , Sept. 2, 2015 Veracyte, ... Bonnie H. Anderson , president and chief executive officer, ... on Wednesday, September 16, 2015 at 10:30 a.m. ET ... The live audio webcast and subsequent replay ... The webcast will be available shortly after conclusion of ...
(Date:9/2/2015)... ... September 02, 2015 , ... Advancing ... high-risk surgeries. From vital sign monitoring in ICU to 3D skull printing analysis ... to make real-time clinical decisions with simultaneous access to high precision medical images ...
(Date:9/2/2015)... , ... September 02, 2015 , ... ... and life science research, today unveiled Genedata Screener ® version 13.0 at ... in-vitro screening brings to market a rich set of new capabilities. Screener now ...
Breaking Biology Technology:Plant-derived Compounds Poised to Assume Greater Role in Global Drug Markets, According to BCC Research 2Plant-derived Compounds Poised to Assume Greater Role in Global Drug Markets, According to BCC Research 3Veracyte to Present at Morgan Stanley Global Healthcare Conference 2Surgical Workstation Powered by 6th Gen. Intel® Core™ Skylake Processors 2Surgical Workstation Powered by 6th Gen. Intel® Core™ Skylake Processors 3New Genedata Screener 13.0 with Support for High Throughput Flow Cytometry and DMPK Unveiled at ELRIG 2New Genedata Screener 13.0 with Support for High Throughput Flow Cytometry and DMPK Unveiled at ELRIG 3New Genedata Screener 13.0 with Support for High Throughput Flow Cytometry and DMPK Unveiled at ELRIG 4
... NEW YORK, April 3 /PRNewswire-Asia-FirstCall/ -- China Agri-Business, ... agricultural,application products through its operating company in China, ... 31,2008. , Revenue for ... a,decrease of 3.8% as compared to $3,037,414 for ...
... CAMBRIDGE, Mass., April 3 Verenium Corporation (Nasdaq: ... development of high-performance specialty enzymes and next-generation cellulosic ... of Verenium,s demonstration-scale facility in Jennings, Louisiana, and ... with BP, Verenium is now consolidating its research ...
... tiny wires with a diameter as small as ... for devices of the future, both in technology like ... circuits for next generation of electronics. But in order ... must be precisely controlled, and researchers must better understand ...
Cached Biology Technology:China Agri-Business Announces 2008 Operating Results 2China Agri-Business Announces 2008 Operating Results 3China Agri-Business Announces 2008 Operating Results 4China Agri-Business Announces 2008 Operating Results 5Verenium Announces Organizational Changes 2Verenium Announces Organizational Changes 3Researchers peer into nanowires to measure dopant properties 2
(Date:9/2/2015)... , September 2, 2015 ... purpose implied in a system which uses identical ... signature, iris, vein, DNA and finger print. Increasing ... to develop such system which provides high level ... PIN, Patterns and tokens are vulnerable to attacks. ...
(Date:8/31/2015)... 2015 Growing need for data ... digitization projects to drive India biometrics ... Research report, " India Biometrics Market Forecast & Opportunities, 2020 ... projected to grow at a CAGR of over 35% during ... account of extensive use of biometric technology in the government ...
(Date:8/28/2015)... PUNE, India , August 28, 2015 ... "Cognitive Assessment and Training Market by Assessment Type (Pen ... Classroom Learning, Brain Training, Corporate Learning, Academic Research), Vertical ... MarketsandMarkets, Cognitive Assessment and Training Market to grow from ... by 2020, at a Compound Annual Growth Rate (CAGR) ...
Breaking Biology News(10 mins):Biometrics Technology Market is Expected to Reach $13.8 Billion by 2015: Hexa Research Inc. 2Biometrics Technology Market is Expected to Reach $13.8 Billion by 2015: Hexa Research Inc. 3Biometrics Technology Market is Expected to Reach $13.8 Billion by 2015: Hexa Research Inc. 4India Biometrics Market to Grow at Over 35% Through 2020, Says TechSci Research 2India Biometrics Market to Grow at Over 35% Through 2020, Says TechSci Research 3Cognitive Assessment and Training Market Worth 7.5 Billion USD by 2020 2Cognitive Assessment and Training Market Worth 7.5 Billion USD by 2020 3Cognitive Assessment and Training Market Worth 7.5 Billion USD by 2020 4
... The Aeras Global TB Vaccine Foundation is pleased to ... Food and Drug Administration (FDA) to develop new biological ... grant, which supports the coordination of a multi-investigator project ... and South Africa, will provide $362,102 in the first ...
... 4 October, 2010 Elsevier Properties SA today ... Alexander (Sandy) Lawson, for receiving the 2011 Herman ... (ACS) Division of Chemical Information (CINF), the award ... theory and practice of chemical information science and ...
... joint work and scientific adventure, marine explorers from more ... Biological Laboratory (MBL), today delivered the first global Census ... lives and hides in the world,s oceans. In one ... 2,700 scientists spent over 9,000 days at sea on ...
Cached Biology News:Aeras-led research consortium receives FDA support 2Professor Dr. Alexander (Sandy) Lawson is awarded the 2011 Herman Skolnik Award 2MBL scientists reveal findings of World Ocean Microbe Census 2MBL scientists reveal findings of World Ocean Microbe Census 3
Non-Protein Peroxidase Stabilizer , 1 L...
Component of GELase™ Agarose Gel-Digesting Preparation. 1 ml of 50X Buffer is sufficient for replacing the buffer in at least 5 g of gel....
RAT ANTI MOUSE METALLOPHILIC MACROPHAGES...
Request Info...
Biology Products: