Human ABCC1 (MRP1) knockout HeLa cell line (ab265256)
Overview
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Product name
Human ABCC1 (MRP1) knockout HeLa cell line
See all MRP1 lysates -
Parental Cell Line
HeLa -
Organism
Human -
Mutation description
Knockout achieved by using CRISPR/Cas9, 14 bp deletion in exon 9 and 2 bp deletion in exon 9 -
Passage number
<20 -
Knockout validation
Immunocytochemistry (ICC), Sanger Sequencing, Western Blot (WB) -
Tested applications
Suitable for: WB, Flow Cyt, ICCmore details -
Biosafety level
2 -
General notes
Recommended control: Human wild-type HeLa cell line (ab255928). Please note a wild-type cell line is not automatically included with a knockout cell line order, if required please add recommended wild-type cell line at no additional cost using the code WILDTYPE-TMTK1.
Cryopreservation cell medium: Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Culture medium: DMEM (High Glucose) + 10% FBS
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. Based on cell count, seed cells in an appropriate cell culture 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.
4. Incubate the culture at 37°C incubator with 5% CO2. 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.
- A partial media change 24 hours prior to subculture may be helpful to encourage growth, if required.
- Cells should be passaged when they have achieved 80-90% confluence.
This product is subject to limited use licenses from The Broad Institute and ERS Genomics Limited, and is developed with patented technology. For full details of the limited use licenses and relevant patents please refer to our limited use license and patent pages.
Properties
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Number of cells
1 x 106 cells/vial, 1 mL -
Viability
~80% -
Adherent /Suspension
Adherent -
Tissue
Cervix -
Cell type
epithelial -
Disease
Adenocarcinoma -
Gender
Female -
STR Analysis
Amelogenin X D5S818: 11, 12 D13S317: 12, 13.3 D7S820: 8, 12 D16S539: 9, 10 vWA: 16, 18 TH01: 7 TPOX: 8,12 CSF1PO: 9, 10 -
Antibiotic resistance
Puromycin 1.00µg/ml -
Mycoplasma free
Yes -
Storage instructions
Shipped on Dry Ice. Store in liquid nitrogen. -
Storage buffer
Constituents: 8.7% Dimethylsulfoxide, 2% Cellulose, methyl ether -
Research areas
Target
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Function
Mediates export of organic anions and drugs from the cytoplasm. Mediates ATP-dependent transport of glutathione and glutathione conjugates, leukotriene C4, estradiol-17-beta-o-glucuronide, methotrexate, antiviral drugs and other xenobiotics. Confers resistance to anticancer drugs. Hydrolyzes ATP with low efficiency. -
Tissue specificity
Lung, testis and peripheral blood mononuclear cells. -
Sequence similarities
Belongs to the ABC transporter superfamily. ABCC family. Conjugate transporter (TC 3.A.1.208) subfamily.
Contains 2 ABC transmembrane type-1 domains.
Contains 2 ABC transporter domains. -
Cellular localization
Cell membrane. - Information by UniProt
Associated products
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KO cell lysates
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab265256 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
| Application | Abreviews | Notes |
|---|---|---|
| WB |
Use at an assay dependent concentration. Predicted molecular weight: 171 kDa.
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| Flow Cyt |
Use at an assay dependent concentration.
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| ICC |
Use at an assay dependent concentration.
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| Notes |
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WB
Use at an assay dependent concentration. Predicted molecular weight: 171 kDa. |
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Flow Cyt
Use at an assay dependent concentration. |
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ICC
Use at an assay dependent concentration. |
Images
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Western blot - Human ABCC1 (MRP1) knockout HeLa cell line (ab265256)All lanes : Anti-MRP1 antibody [EPR21062] (ab233383) at 1/1000 dilution
Lane 1 : Wild-type HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 2 : MRP1 knockout HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 3 : Wild-type A549 (Human lung carcinoma cell line) whole cell lysate
Lane 4 : MRP1 knockout A549 (Human lung carcinoma cell line) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/20000 dilution
Performed under reducing conditions.
Predicted band size: 171 kDa
Observed band size: 250 kDa why is the actual band size different from the predicted?Lanes 1-4: Merged signal (red and green). Green - ab233383 observed at 250 kDa. Red - loading control ab7291 observed at 50 kDa.
ab233383 Anti-MRP1 antibody [EPR21062] was shown to specifically react with MRP1 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265256 (knockout cell lysate ab257242) was used. Wild-type and MRP1 knockout samples were subjected to SDS-PAGE. ab233383 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) were incubated overnight at 4°C 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.
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Immunocytochemistry/ Immunofluorescence - Human ABCC1 (MRP1) knockout HeLa cell line (ab265256)ab233383 staining MRP1 in wild-type HeLa cells (top panel) and ABCC1 knockout HeLa cells (ab265256) (bottom panel). The cells were fixed with 4% paraformaldehyde (10 min) then permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab233383 at 1/100 dilution and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8). -
Flow Cytometry (Intracellular) - Human ABCC1 (MRP1) knockout HeLa cell line (ab265256)Flow cytometry overlay histogram showing wild-type HeLa (green line) and ABCC1 knockout HeLa cells (ab265256) stained with ab260038 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10% normal goat serum to block non-specific protein-protein interaction followed by the antibody (ab260038) (1x106 in 100μl at 0.2 μg/ml) for 30 min at 22°C.
The secondary antibody Goat anti-rabbit IgG H&L (Alexa Fluor® 488, pre-adsorbed) (ab150081) was used at 1/2000 for 30 min at 22°C.
Isotype control antibody was Rabbit IgG (monoclonal) (ab172730) used at the same concentration and conditions as the primary antibody (wild-type HeLa - black line ABCC1 knockout HeLa - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).
Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.
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Sanger Sequencing - Human ABCC1 knockout HeLa cell line (ab265256)Allele-1: 14 bp deletion in exon 9.
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Sanger Sequencing - Human ABCC1 knockout HeLa cell line (ab265256)Allele-2: 2 bp deletion in exon 9.
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Cell Culture - Human ABCC1 (MRP1) knockout HeLa cell line (ab265256)Representative images of ABCC1 knockout HeLa cells, low and high confluency examples (top left and right respectively) and wild-type HeLa cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using a EVOS XL Core microscope.
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (0)
ab265256 has not yet been referenced specifically in any publications.