Navigation Links
Laser Capture Microdissection of Cells Labeled with Enhanced Green Fluorescent Protein

equence Detection System (Applied Biosystems, Foster City, CA)


Special Considerations
Excessive fixation of the tissue prior to embedding prevents efficient extraction of RNA from captured cells. Illumination of tissue sections under epifluorescence optics should be minimized in order to reduce photo-bleaching and prolong EGFP fluorescence.


Method
CRYOSECTIONING AND LCM
1. Fix freshly dissected tissues in 4% paraformaldehyde in phosphate buffered saline (PBS) for 5 min. Mild fixation by 4% PFA retains EGFP within the cells during embedding and sectioning without affecting subsequent RNA extraction. Wash the tissue twice in PBS for 5 min, then freeze in OCT compound on dry ice.

2. Pre-clean the internal surfaces of the cryostat with 100% ethanol. Perform cryosectioning onto silanized glass slides in a cryostat at 19 C using a fresh disposable low-profile blade under RNase-free conditions. Mount sections onto slides at room temperature but subsequent storage should be inside the cryostat (short term) or at 70 C (long term).

3. Process tissue sections for LCM within 2 days of cryosectioning. Allow mounted tissue sections to thaw for no longer than 20 seconds before beginning dehydration. Dehydrate sections in 70%, 95% and 100% ethanol for 30 seconds each, followed by 5 min in xylene. Air-dry sections for 2 minutes before commencing LCM.

4. Visualize EGFP-expressing cells in the tissue sections using the EGFP-optimized emission filters with an integration setting of 0.12-0.2 using an Arcturus PixCell IIe system equipped with epifluorescence optics.

5. A spot size of 7.5 m with power settings of 60-70 mA and a pulse duration of 1.3 msec ensured efficient capture of cells.

6. The laser spot overwhelms green fluorescence, so comparison of an image map acquired under fluorescence with the live camera image (under brightfiel
'"/>

Source:


Page: All 1 2 3 4 5

Related biology technology :

1. Isolate and Analyze Total RNA from Cells Harvested by Laser Capture Microdissection
2. Laser microdissection and nucleic acid purification - a Leica - QIAGEN cooperation
3. Working With Laser Capture Microdissection Samples
4. Laser Capture Microdissection (LCM) for the Analysis of Macrophage Gene Expression from Atherosclerotic Lesions
5. Laser Capture Microdissection (LCM) and Expression Profiling of Long-Range Projection Neurons
6. Differential Gene Expression Analysis of Pure Cell Populations from Frozen UterineTissue Using Laser Capture Microdissection (LCM) and cDNA Microarrays
7. Laser Capture Microdissection of muscle fiber populations and expression analysis by RT-PCR
8. An Epitope Tagging Vector for Gene Expression in Mammalian Cells
9. Versatile Vectors for Ponasterone A- Inducible Control of Gene Expression in Mammalian Cells
10. Cell Proliferation Kit Outperforms the Competition in the Range of Cells Typically Counted
11. XL10-Gold Cells Supplied in New Single-Reaction Format
Post Your Comments:
(Date:12/22/2014)... , Dec. 22, 2014  Alternative Energy & ... it has signed a letter of intent to ... developed and patented a nanotechnology-based development platform used ... that enable rapid on-site collection and testing to ... health issues in an immediate, non-invasive and cost-effective ...
(Date:12/22/2014)... December 22, 2014 ITRA Global, one ... occupiers of commercial real estate, has further expanded its ... in Perth and Brisbane, Western Australia, reports Mylinda Vick, ... , ITRA Global / ACORPP (Australian Corporate Property ... consultancy company providing property services to a wide range ...
(Date:12/19/2014)... (PRWEB) December 19, 2014 BioPlus ... specialty pharmacies, announces the promotion of Nick Maroulis, Pharm.D. ... Specialty Pharmacy Services. , In this position, Dr. ... while also managing the directors of our multi-site pharmacies ... BioPlus since 1997 and during that time he has ...
(Date:12/19/2014)... , Dec. 19, 2014 Decision Resources Group ... grow at a 12 percent compound annual growth rate ... the aging population and the increasing adoption of dental ... will also spur demand for dental biomaterials ... implant and periodontal treatments. Other key findings ...
Breaking Biology Technology:ALNE Announces Intention To Acquire BioTechPharma 2ITRA Global Continues International Expansion in Perth and Brisbane, Australia 2BioPlus Specialty Pharmacy Promotes Dr. Nick Maroulis to Vice President of Specialty Pharmacy Services 2Latin American Dental Biomaterial Market Will Expand at a 12 Percent Annual Growth Rate Through 2023 2Latin American Dental Biomaterial Market Will Expand at a 12 Percent Annual Growth Rate Through 2023 3
... Dissolve Powder Antacid Delivers Faster Relief, ... Increased Convenience and Great Taste, PITTSBURGH, April ... integrated marketing campaign,for the new TUMS(R) QuikPak(TM), the first and ... QuikPak campaign, part of,a larger new "Bring It On" brand ...
... - MIGENIX Inc.,(TSX: MGI, OTC: MGIFF), a clinical-stage ... Dorsman to its Board of Directors. Mr.,Dorsman is ... a,financial consulting firm in Vancouver, which advises companies ... overall business,strategies. Prior to founding his consulting firm ...
... Sirion Therapeutics, Inc., a,privately held ophthalmic-focused biopharmaceutical company, ... targeted for the,treatment of retinal diseases will be ... for Research in Vision and Ophthalmology (ARVO),April 27 ... http://www.newscom.com/cgi-bin/prnh/20080220/CLW064LOGO ), "We are very ...
Cached Biology Technology:New TUMS(R) QuikPak(TM) Campaign Offers Innovative Heartburn Relief with a Little Help from the Jetsons 2MIGENIX Appoints Pieter Dorsman to Board of Directors 2Sirion Therapeutics Preclinical Studies: Two New Potential Advances in Treatment of Retinal Diseases to be Presented at ARVO 2Sirion Therapeutics Preclinical Studies: Two New Potential Advances in Treatment of Retinal Diseases to be Presented at ARVO 3
(Date:12/10/2014)... , Dec. 9, 2014  Wake Forest Baptist Medical ... building for its School of Medicine. Funding for this ... capital campaign that will be publicly launched next summer. ... located in the former 60 series R.J. Reynolds Tobacco ... Quarter. Construction will begin immediately with plans to be ...
(Date:12/10/2014)... Research and Markets ( http://www.researchandmarkets.com/research/xclcvw/biometrics_market ... "Biometrics Market in Japan 2014-2018" report to ... The integration of biometrics and ... and upgradation of the driver,s license is one ... Besides the aforementioned projects, biometrics is being used ...
(Date:12/5/2014)... 4, 2014 Increased emphasis on product ... new testing and inspection technologies. The convergence of ... a range of innovative test solutions. This trend ... Y, which is generally more inclined towards communication, ... development strategies of test equipment vendors will cater ...
Breaking Biology News(10 mins):Wake Forest Baptist to Build New Medical Education Facility In Wake Forest Innovation Quarter 2Wake Forest Baptist to Build New Medical Education Facility In Wake Forest Innovation Quarter 3Wake Forest Baptist to Build New Medical Education Facility In Wake Forest Innovation Quarter 4Biometrics Market in Japan 2014-2018: Key Vendors are DDS, Fujitsu, Hitachi and NEC 2Emerging Technologies Enable Smarter Monitoring of Assets, Push Industrial Testing, Inspection and Monitoring Market 2Emerging Technologies Enable Smarter Monitoring of Assets, Push Industrial Testing, Inspection and Monitoring Market 3Emerging Technologies Enable Smarter Monitoring of Assets, Push Industrial Testing, Inspection and Monitoring Market 4
... Researchers at the University of Alabama at Birmingham (UAB) ... grow and remodel themselves to stay strong, a finding ... osteoporosis drugs, the researchers said The coupling factor ... or TGF beta-1. Previously, scientists had searched for but ...
... "Push your finger as hard as you can against the ... slowly - follow the ticking clock. Now faster. Now faster." ... experiment that casts doubt on old ideas about mechanisms to ... explain why manual dexterity is so vulnerable to aging and ...
... Ore. A 700-mile security wall under construction along ... the movement and "connectivity" of wildlife, biologists say, and ... of some species. However, technology and alterations to ... to move more freely between the two countries, the ...
Cached Biology News:UAB study reveals bone coupling factor key to skeletal health 21-finger exercise reveals unexpected limits to dexterity 21-finger exercise reveals unexpected limits to dexterity 3US-Mexico border wall could threaten wildlife species 2US-Mexico border wall could threaten wildlife species 3US-Mexico border wall could threaten wildlife species 4
... system is used for single-color real-time detection ... or JOE fluorophores and features a real-time ... data from 96 wells, a gene expression ... analysis. The system includes the optical module ...
... PCR detection system offers an affordable alternative ... dyes such as FAM and SYBR Green ... iCycler thermal cycler, offering superior features such ... The MyiQ real-time PCR detection system is ...
... systems are highly sensitive laser confocal systems ... are advanced scanners that meet your expectations, ... level, and optimizing the signal on the ... of the range for increased sensitivity. Results ...
Bio-Plex Manager software, version 3.0, 5 license netshare, is a five-user software package that provides access to data from the Bio-Plex suspension array system from a server. This version does not...
Biology Products: