Navigation Links
Primary cells, rat heart muscle

Multiporator Transfection Protocol Protocol No. 4308 915.035 12/1999 Cell line Primary cells: Adult heart muscle cells from rat, 6 days in culture Transfection with Plasmid pCMV--galactosidase (in bidistilled H2O) Electroporation buffer Eppendorf Isoosmolar Electroporation Buffer (PI) Culture medium M199 / HEPES (2 mM carnithin, 5 mM taurine, 10 M cytosine-arabinofur.)+ 20% FCS Outgrowth medium HEPES-buffer (140 mM NaCl, 3.6 mM KCl, 1.2 mM MgSO4, 1 mM CaCl2, 11 mM glucose, 100 U/ml penicillin, 100 g/ml streptomycin, 20 mM HEPES, pH 7.4) without phenol red + 10% FCS Cuvette Eppendorf, 2 mm gap width, 400 l Temperature RT (20-25 C) Reference Dr. Gerhild Taimor Justus-Liebig-Universitt Physiologisches Institut Aulweg 129 D-35392 Gieen
Phone +49 641 9947243 Fax +49 641 9947239 mail: gerhild.taimor@physiologie.med.uni-giessen.de
  1. Harvest the cells by collagenase treatment (0.4 mg/ml, 30 min., 37 C) and centrifuge them (for 2 minutes, 300 x g, at room temperature).
  2. Resuspend the cells in M199 / 0.5% FCS, determine the number of cells and centrifuge them (for 2 minutes, 300 x g, at room temperature). Remove supernatant.

    Note: The overall incubation time in the Eppendorf Electroporation Buffer must not exceed 30 minutes to guarantee a successful electroporation!

  3. Resuspend the cells in Isoosmolar Electroporation Buffer. When doing so, set the cell concentration to 1 x 104 - 105 cells/ml.
  4. Add and mix plasmid DNA (5-20 g/ml final concentration, in bidistilled H2O).
  5. Transfer 400 l cell suspension into electroporation cuvettes (2 mm gap width). The cell suspension must be free of air bubbles.
  6. Electroporation:

    Mode Eukaryotes Voltage (V) 100 V Time constant (T) 50 s No. of pulses (n) 1
  7. After the pulse, allow the cell suspension to stand in the cuvette for 10 minutes at room temperature.
  8. Carefully transfer the cell suspension from the cuvette to 3-5 ml HEPES medium without phenol red / 20% FCS, and cultivate them in a 35 mm culture dish.

Detection methods for transfection:

The expression of the plasmid pCMV--galactosidase can be detected clearly after 24-48 hours with X-gal-staining under a microscope.


'"/>

Source:


Page: All 1 2

Related biology technology :

1. Human Primary Preadipocytes and Differentiated Adipocytes
2. Primary cells, rat heart endothel
3. Efficient Delivery of siRNAs to Human Primary Cells: Electroporation vs. Chemical Transfection
4. Deliver siRNAs Into Primary Cells
5. High Throughput siRNA Delivery In Vitro: From Cell Lines to Primary Cells
6. Electroporation of Primary Bone Marrow Cells
7. Efficient Primary Structure Elucidation of Disulfide-bridged Peptides
8. Embryonic stem cells, mouse
9. Laser Capture Microdissection of muscle fiber populations and expression analysis by RT-PCR
Post Your Comments:
*Name:
*Comment:
*Email:
TAG: Primary cells rat heart muscle

(Date:8/28/2015)... DUBLIN , Aug. 28, 2015 ... announced the addition of the "Global & USA ... offering. The highly awaited US Biosimilar market ... highly awaited filigrastim biologic, Zarxio. How will the global ... The author reveals that biosimilars could save the US ...
(Date:8/28/2015)... India , August 28, 2015 ... by Product (Media, Sera, Cryogenic Storage Systems, Thawing Equipment, ... (Therapeutic, Regenerative Medicine, Clinical Trials), End User - Global ... Market is expected to reach USD 3,731.03 Million by ... a CAGR of 11.65% between 2015 and 2020. ...
(Date:8/28/2015)... , ... August 28, 2015 , ... ... Medicine, presented two studies at the International Spine Intervention Society’ s ... 2015 through to August 1st, 2015. With a focus on evidence-based research and ...
(Date:8/28/2015)... Pittsburgh, PA (PRWEB) , ... August 28, 2015 ... ... Program Chairman, Chuck Gardner, has organized and will present a session on chemical ... September 2. This conference and exposition is Asia’s largest analytical and scientific instruments ...
Breaking Biology Technology:Global & USA BioSimilar Market Analysis Report 2015-2021 2Global & USA BioSimilar Market Analysis Report 2015-2021 3Biopreservation Market Worth 3,731.03 Million USD by 2020 2Biopreservation Market Worth 3,731.03 Million USD by 2020 3Biopreservation Market Worth 3,731.03 Million USD by 2020 4Biopreservation Market Worth 3,731.03 Million USD by 2020 5Proove Biosciences Presents New Research Study at the International Spine Intervention Society’s Annual Scientific Meeting 2Proove Biosciences Presents New Research Study at the International Spine Intervention Society’s Annual Scientific Meeting 3Proove Biosciences Presents New Research Study at the International Spine Intervention Society’s Annual Scientific Meeting 4U.S. Symposium on Imaging Organized by Pittcon’s Program Chairman Chuck Gardner 2U.S. Symposium on Imaging Organized by Pittcon’s Program Chairman Chuck Gardner 3
... The Milwaukee Brewers will soon have their ... at Miller Park, officials announced Thursday. , ,Eastman Kodak ... provide a Directview CR 850 computer-operated radiography ... be used to generate high-resolution digital X-rays of injured ...
... If you had blinked, you would have missed two security ... Web server last Wednesday. , ,Dane Deutsch and Pete Adams, ... used a simple tool to gain complete control of a ... E-Business Institutes conference at the Monona Terrace, without leaving a ...
... Ralph Kauten. Source: Teresa Esser. Ralph is one of ... of bioscience, said Frank Langley, president and CEO of Point ... Kauten to admit that his success in high-tech entrepreneurship is ... Kauten said in an interview. Ive been a lucky guy. ...
Cached Biology Technology:Hacking tools are simple and widely available, security experts say 2Ralph Kauten on how to raise angel financing 2Ralph Kauten on how to raise angel financing 3Ralph Kauten on how to raise angel financing 4
(Date:7/31/2015)... Die 10. internationale Konferenz zu Genomik (ICG-10, ... vom 22. bis 25. Oktober 2015 in ... Die Konferenz feiert in diesem Jahr ihr ... ICG weltweit zu einem der einflussreichsten jährlichen Treffen ... dynamischsten, enthusiastischsten und angenehmsten wissenschaftlichen Zusammentreffen. ...
(Date:7/31/2015)... , July 31, 2015 The 10 th ... held by BGI from October 22-25, 2015, in Shenzhen ... The conference is celebrating its 10 th anniversary this year. ... the world,s most influential annual meetings in the ,omics, fields, ... scientific gatherings. ICG-10 focuses on recent breakthroughs ...
(Date:7/27/2015)... JOSE, Calif. , July 27, 2015 /PRNewswire/ ... developer of human interface solutions, today announced that ... ® touchscreen solution for its stylish smartwatch. ... for its proven reliability, low power and highly ... a wet finger. Huawei designers also required a ...
Breaking Biology News(10 mins):Die 10. internationale Konferenz zu Genomik (ICG-10) beginnt im Oktober 2Die 10. internationale Konferenz zu Genomik (ICG-10) beginnt im Oktober 3The 10th International Conference on Genomics (ICG-10) to Open in October 2Synaptics Touchscreen Solution Powers New Huawei Watch 2Synaptics Touchscreen Solution Powers New Huawei Watch 3
... leader? Traits that we look for typically include a sense ... a recent study suggests that it is not just an ... chosen as leader of a group. Previous research has implicated ... for a male versus female leader may depend on the ...
... years of near-zero growth in atmospheric methane concentrations, levels ... good news for future global warming," says CSIRO,s Dr ... in Geophysical Research Letters, a journal of the American ... important greenhouse gases, namely methane and the CFCs, had ...
... Researchers at Georgetown University Medical Center have discovered a ... to tamoxifen, the world,s largest-selling breast cancer prevention and ... way to identify tamoxifen users who are no longer ... therapy option sooner. , In the November 1 issue ...
Cached Biology News:Gender biases in leadership selection during competitions within and between groups 2Gender biases in leadership selection during competitions within and between groups 3Global methane levels on the rise again 2'New' estrogen receptor found to be key player in tamoxifen resistance 2'New' estrogen receptor found to be key player in tamoxifen resistance 3
... 1 Block MBS Start up kit. Includes ... and choice of 1 MultiBlock satellite ... 30 blocks. User friendly software. ... Adjustable height heated lid. Outstanding uniformity. ...
Monoclonal mouse antibody for immunihistology, clone AD12, Isotype IgG 1, purified, specific for rat pyruvatekinase type L, no cross-reactivities with other pyruvate kinase isoenzymes have been obser...
PxE thermal Cycler. 0.5ml block holds 48 0.5 ml tubes or 96 0.3 ml tubes. Adjustable height heated lid. Outstanding uniformity. Advanced control algorithms for reduced reaction times. User frien...
Carrier Serum...
Biology Products: