Human MDH2 knockout HEK-293T cell line
- Advanced Validation
- What is this?
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(0 Publication)
- WB
Lab
Western blot - Human MDH2 knockout HEK-293T cell line (AB266449)
Lanes 1-4 : Merged signal (red and green). Green - ab181857 observed at 36 kDa. Red - loading control ab7291 observed at 50 kDa.
ab181857 Anti-MDH2 antibody [EPR14883(B)] was shown to specifically react with MDH2 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266449 (knockout cell lysate ab257533) was used. Wild-type and MDH2 knockout samples were subjected to SDS-PAGE. ab181857 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) were incubated overnight at 4° at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes:
Western blot - Anti-MDH2 antibody [EPR14883(B)] (<a href='/en-us/products/primary-antibodies/mdh2-antibody-epr14883b-ab181857'>ab181857</a>) at 1/1000 dilution
Lane 1:
Wild-type HEK293T cell lysate at 20 µg
Lane 2:
MDH2 knockout HEK293T cell lysate at 20 µg
Lane 2:
Western blot - Human MDH2 knockout HEK-293T cell line (ab266449)
Lane 3:
K-562 cell lysate at 20 µg
Lane 4:
Human eyeball tissue lysate at 20 µg
Secondary
All lanes:
Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution
Predicted band size: 36 kDa
Observed band size: 36 kDa
false
- WB
Lab
Western blot - Human MDH2 knockout HEK-293T cell line (AB266449)
Lanes 1-4 : Merged signal (red and green). Green - ab181873 observed at 36 kDa. Red - loading control ab7291 observed at 50 kDa.
ab181873 Anti-MDH2 antibody [EPR14882(B)] was shown to specifically react with MDH2 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266449 (knockout cell lysate ab257533) was used. Wild-type and MDH2 knockout samples were subjected to SDS-PAGE. ab181873 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) were incubated overnight at 4° at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes:
Western blot - Anti-MDH2 antibody [EPR14882(B)] (<a href='/en-us/products/primary-antibodies/mdh2-antibody-epr14882b-ab181873'>ab181873</a>) at 1/1000 dilution
Lane 1:
Wild-type HEK293T cell lysate at 20 µg
Lane 2:
MDH2 knockout HEK293T cell lysate at 20 µg
Lane 2:
Western blot - Human MDH2 knockout HEK-293T cell line (ab266449)
Lane 3:
K-562 cell lysate at 20 µg
Lane 4:
Human eyeball tissue lysate at 20 µg
Secondary
All lanes:
Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution
Predicted band size: 36 kDa
Observed band size: 36 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human MDH2 knockout HEK-293T cell line (AB266449)
Allele-1 : 4 bp deletion in exon 4
- Sanger seq
Unknown
Sanger Sequencing - Human MDH2 knockout HEK-293T cell line (AB266449)
Allele-2 : Insertion of the selection cassette in exon 4.
- Cell Culture
Unknown
Cell Culture - Human MDH2 knockout HEK-293T cell line (AB266449)
Representative images of MDH2 knockout HEK293T cells, low and high confluency examples (top left and right respectively) and wild-type HEK293T cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using an EVOS M5000 microscope.
Complementary Products
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MDH2 antibody [EPR14882(B)] (AB181873)](https://content.abcam.com/products/images/mdh2-antibody-epr14882b-ab181873--immunohistochemistry-formalin-pfa-fixed-paraffin-embedded-sections-img133692.jpg)
Reactivity data
Product details
We will provide viable cells that proliferate on revival.
This product is subject to limited use licenses from The Broad Institute, ERS Genomics Limited and Sigma-Aldrich Co. LLC, and is developed with patented technology. For full details of the licenses and patents please refer to our limited use license and patent pages.
What's included?
Properties and storage information
Gene name
Gene editing type
Gene editing method
Knockout validation
Shipped at conditions
Appropriate short-term storage conditions
Appropriate long-term storage conditions
Handling procedures
Initial handling guidelines
Upon arrival, the vial should be stored in liquid nitrogen vapor phase and not at -80°C. Storage at -80°C may result in loss of viability.
1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.
2. Transfer the cell suspension (0.8 mL) to a 15 mL/50 mL conical sterile polypropylene centrifuge tube containing 8.4 mL pre-warmed culture medium, wash vial with an additional 0.8 mL culture medium (total volume 10 mL) to collect remaining cells, and centrifuge at 201 x g (rcf) for 5 minutes at room temperature. 10 mL represents minimum recommended dilution. 20 mL represents maximum recommended dilution.
3. Resuspend the cell pellet in 5 mL pre-warmed culture medium and count using a haemocytometer or alternative cell counting method seed all remaining cells into a T25.
4. Incubate the culture at 37°C incubator with 5% CO2. Check the culture one day after revival and continue to check until 80% confluent. Media change can be given if needed.
5. Once confluent passage into an appropriate flask at a density of 2x104 cells/cm2. Seeding density is given as a guide only and should be scaled to align with individual lab schedules. Cultures should be monitored daily.
Subculture guidelines
- All seeding densities should be based on cell counts gained by established methods.
- A guide seeding density of 2x104 cells/cm2 is recommended.
- Cells should be passaged when they have achieved 80-90% confluence.
Culture medium
DMEM (High Glucose) + 10% FBS
Cryopreservation medium
Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Supplementary information
This supplementary information is collated from multiple sources and compiled automatically.
Biological function summary
MDH2 participates in the critical process of energy production within the cell. While it does not form a complex itself its activity is intimately connected with other enzymes in mitochondrial energy metabolism. The malate dehydrogenase assay often measures the activity of MDH2 to understand the metabolic status of cells. By facilitating the oxidation of malate MDH2 aids in maintaining the efficiency of the mitochondrial electron transport chain by regenerating NADH.
Pathways
The enzyme is essential in the tricarboxylic acid (TCA) cycle and the malate-aspartate shuttle. In the TCA cycle MDH2 collaborates with enzymes like citrate synthase and isocitrate dehydrogenase to assist in the conversion of acetyl-CoA into energy-rich molecules. The malate-aspartate shuttle on the other hand involves MDH2 working closely with aspartate transaminase to transfer reducing equivalents into the mitochondria. These pathways highlight MDH2's importance in cellular energy homeostasis.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 2 US: 2
Adherent/suspension
Adherent
Gender
Female
Target data
Product promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
For licensing inquiries, please contact partnerships@abcam.com