Human ABCC2 knockout A549 cell line (ab261855)
Overview
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Product name
Human ABCC2 knockout A549 cell line -
Parental Cell Line
A549 -
Organism
Human -
Mutation description
Knockout achieved by CRISPR/Cas9; X = 11 bp deletion; Frameshift = 97% -
Passage number
<20 -
Knockout validation
Next Generation Sequencing (NGS), Western Blot (WB) -
Tested applications
Suitable for: WBmore details -
Biosafety level
1 -
General notes
Recommended control: Human wild-type A549 cell line (ab259777). 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:Hams F12 + 5% 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 2x103-1x104 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 6x104 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.
- Do not exceed 7x104 cells/cm2.
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
Lung -
Cell type
epithelial -
Disease
Carcinoma -
Gender
Male -
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 hepatobiliary excretion of numerous organic anions. May function as a cellular cisplatin transporter. -
Tissue specificity
Found on the apical membrane of polarized cells in liver, kidney and intestine. The highest expression is found in liver. -
Involvement in disease
Defects in ABCC2 are the cause of Dubin-Johnson syndrome (DJS) [MIM:237500]. DJS is an autosomal recessive disorder characterized by conjugated hyperbilirubinemia, an increase in the urinary excretion of coproporphyrin isomer I, deposition of melanin-like pigment in hepatocytes, and prolonged retention of sulfobromophthalein, but otherwise normal liver function. -
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
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 ab261855 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.
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| Notes |
|---|
|
WB
Use at an assay dependent concentration. |
Images
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Western blot - Human ABCC2 knockout A549 cell line (ab261855)All lanes : Anti-MRP2 antibody [M2III-5] (ab15603) at 1/20 dilution
Lane 1 : Wild-type A549 (Human lung carcinoma cell line) whole cell lysate
Lane 2 : ABCC2 knockout A549 (Human lung carcinoma cell line) whole cell lysate
Performed under reducing conditions.Lanes 1 - 2: Merged signal (red and green). Green - ab15603 observed at 210 kDa. Red - loading control, ab52866, observed at 50 kDa.
ab15603 was shown to recognize ABCC2 in wild-type A549 cells as signal was lost at the expected MW in ABCC2 knockout cell line ab261855 (knockout cell lysate ab261663). Additional cross-reactive bands were observed in the wild-type and knockout samples. Wild-type and ABCC2 knockout samples were subjected to SDS-PAGE. The membrane was blocked with 3% milk. Ab15603 and ab52866 (Rabbit anti-alpha Tubulin loading control) were incubated overnight at 4°C at 1/20 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed ab216772 and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed ab216777 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.
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Western blot - Human ABCC2 knockout A549 cell line (ab261855)All lanes : Anti-MRP2 antibody [M2 III-6] (ab3373) at 1/20 dilution
Lane 1 : Wild-type A549 (Human lung carcinoma cell line) whole cell lysate
Lane 2 : ABCC2 knockout A549 (Human lung carcinoma cell line) whole cell lysate
Lane 3 : Hep G2 (Human liver hepatocellular carcinoma cell line) whole cell lysate
Performed under reducing conditions.Lanes 1 - 3: Merged signal (red and green). Green - ab3373 observed at 210 kDa. Red - loading control, ab52866, observed at 50 kDa.
ab3373 was shown to recognize ABCC2 in wild-type A549 cells as signal was lost at the expected MW in ABCC2 knockout cell line ab261855 (knockout cell lysate ab261663). Additional cross-reactive bands were observed in the wild-type and knockout samples. Wild-type and ABCC2 knockout samples were subjected to SDS-PAGE. The membrane was blocked with 3% milk. Ab3373 and ab52866 (Rabbit anti-alpha Tubulin loading control) were incubated overnight at 4°C at 1/20 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed ab216772 and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed ab216777 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.
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Western blot - Human ABCC2 knockout A549 cell line (ab261855)All lanes : Anti-MRP2 antibody [EPR10997(2)] (ab187644) at 1/1000 dilution
Lane 1 : Wild-type A549 (Human lung carcinoma cell line) whole cell lysate
Lane 2 : ABCC2 knockout A549 (Human lung carcinoma cell line) whole cell lysate
Lane 3 : Hep G2 (Human liver hepatocellular carcinoma cell line) whole cell lysate
Lane 4 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.Lanes 1 - 4: Merged signal (red and green). Green - ab187644 observed at 210 kDa. Red - loading control, ab130007, observed at 130 kDa.
ab187644 was shown to recognize MRP2 in wild-type A549 cells as signal was lost at the expected MW in ABCC2 knockout cell line ab261855 (knockout cell lysate ab261663). Additional cross-reactive bands were observed in the wild-type and knockout samples. Wild-type and ABCC2 knockout samples were subjected to SDS-PAGE. The membrane was blocked with 3% milk. Ab187644 and ab130007 (Mouse anti-Vinculin loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/1000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.
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Western blot - Human ABCC2 knockout A549 cell line (ab261855)All lanes : Anti-MRP2 antibody [EPR10998] (ab172630) at 1/1000 dilution
Lane 1 : Wild-type A549 (Human lung carcinoma cell line) whole cell lysate
Lane 2 : ABCC2 knockout A549 (Human lung carcinoma cell line) whole cell lysate
Lane 3 : Hep G2 (Human liver hepatocellular carcinoma cell line) whole cell lysate
Lane 4 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.Lanes 1 - 4: Merged signal (red and green). Green - ab172630 observed at 210 kDa. Red - loading control, ab130007, observed at 130 kDa.
ab172630 was shown to recognize MRP2 in wild-type A549 cells as signal was lost at the expected MW in ABCC2 knockout cell line ab261855 (knockout cell lysate ab261663). Additional cross-reactive bands were observed in the wild-type and knockout samples. Wild-type and ABCC2 knockout samples were subjected to SDS-PAGE. The membrane was blocked with 3% milk. Ab172630 and ab130007 (Mouse anti-Vinculin loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/1000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.
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Next Generation Sequencing - Human ABCC2 knockout A549 cell line (ab261855)X = 11 bp deletion
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (0)
ab261855 has not yet been referenced specifically in any publications.