Pre-Coating Electrodes with Defined Protein Coats
The gold ECISŪ electrodes are pristine and have no coatings when shipped from Applied BioPhysics. When culture medium is added to these arrays, the proteins and other large molecules in the medium will immediately adsorb to the very wettable gold surface as they do with any uncoated tissue culture dish (note: this will take place even after reacting the gold with cysteine).
Often it is desirable to alter cell behavior by pre-coating the electrodes with fibronectin, laminin or other ECM proteins. We suggest using the following protocol:
(1) Make up a solution of the desired protein at 100 micrograms per ml or more in 0.15M NaCl. If a buffer is required, a mild Tris solution (e.g. 0.01M) is recommended. The electrode arrays are stable under acidic conditions, so when coating with collagen there is no problem using solutions containing acetic acid.
(2) To coat the electrode, simply pipet the protein solution into the well with sufficient volume to flood the substrate surface; usually a volume of 200 microliters is sufficient for most well types. If the protein is valuable, with 1E and 1E+ arrays, it is only necessary to coat the small active electrodes, and a small volume can be applied directly to the electrode surfaces.
(3) Allow adsorption to take place for at least 15 minutes and longer for more dilute solutions.
(4) Once adsorption is complete, a molecular layer of the protein will be coating the surfaces. You can now remove the protein solution and rinse any residual protein from the well with sterile medium, saline or water without concern of removing the adsorbed protein.
(5) The arrays are now ready for other solutions (e.g. cysteine) and inoculation.
We do not recommend:
- Drying protein solutions in place as this may foul or damage the electrodes.
- The use of phosphate buffer (PBS) for adsorption as this has been shown to interfere with the adsorption of some proteins.
- The use of very dilute protein solutions (<50 micrograms per ml). Remember protein may be adsorbing to the walls of the vessels holding solutions at a level as high as 1 microgram/cm2. This can significantly reduce the concentration of protein in very dilute solution.
Applied BioPhysics, Inc. applies the results of biophysical research to provide practical tools for cell research and drug discovery.
ECIS™ or Electric Cell-substrate Impedance Sensing is an impedance based method to study many of the activities of animal cells when grown in tissue culture in real-time. These include morphological changes, cell locomotion, and other behaviors directed by the cell's cytoskeleton.
The ECIS™ approach has been applied to numerous investigations including measurements of the invasive nature of cancer cells, the barrier function of endothelial cells, in vitro toxicity testing as an alternative to animal testing, and signal transduction involving GPCR's for modern drug discovery.
ECIS Application Webinar Series
ECIS application webinars review the topics listed below in 20 to 30 minute, web-based, interactive seminars presented by Applied BioPhysics president and co-founder, Dr. Charles Keese.
Barrier Function Assays - June 24, 2014
Real-time Electroporation and Monitoring - July 8, 2014
Cell Attachment and Spreading Measurements - July 22, 2014
Tradeshows & Events
Gorden Research Conference
Barriers of the CNS
June 15 - 20, 2014Colby-Sawyer College
New London, NH
3D Cell Culture 2014
Advanced Model Systems, Applications & Enabling Technologies
June 25 - 27, 2014
Gorden Research Conference
Endothelial Cell Phenotypes in Health & Disease
July 6 - 11, 2014
Melia Golf Vichy Catalan Business & Convention Center
Girona-Costa Brava, Spain
The Lung Epithelium in Health and Disease
July 27 - August 1, 2014
Saxtons River, VT
11th ISRA: From Molecular Machinery to Clinical Challenges
September 7 - 11, 2014
Korea Society for Molecular and Cellular Biology (KSMCB)
Oct 9th - 11th, 2014
InterContinental Hotels Seoul COEX
Seoul, South Korea
The 87th Annual Meeting of the Japanese Biochemical Society
October 15 - 18, 2014
Kyoto International Conference Center
with Nepa Gene Co., Ltd.
Vascular Biology 2014
Oct 19 - 23, 2014
Asilomar Conference Grounds
American Society for Cell Biology
December 6 - 10, 2014