Human S100A4 knockout HeLa cell line (ab265709)
Overview
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Product name
Human S100A4 knockout HeLa cell line
See all S100A4 lysates -
Parental Cell Line
HeLa -
Organism
Human -
Mutation description
Knockout achieved by using CRISPR/Cas9, 1 bp insertion in exon 2 and 5 bp deletion in exon 2 -
Passage number
<20 -
Knockout validation
Sanger Sequencing, Western Blot (WB) -
Tested applications
Suitable for: WB, Sanger Sequencingmore 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, 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.
We will provide viable cells that proliferate on revival.
Properties
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Number of cells
1 x 106 cells/vial, 1 mL -
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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Tissue specificity
Ubiquitously expressed. -
Sequence similarities
Belongs to the S-100 family.
Contains 2 EF-hand domains. - Information by UniProt
Associated products
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Related Products
- Anti-S100A4 antibody [EPR2761(2)] (ab124805)
- Anti-S100A4 antibody [EPR14639(2)] (ab197896)
- Anti-S100A4 antibody [EPR2761(2)] - BSA and Azide free (ab216003)
- Anti-S100A4 antibody [S100A4/1481] (ab218511)
- Anti-S100A4 antibody [EPR14639(2)] - BSA and Azide free (ab220213)
- Anti-S100A4 antibody [EPR2761(2)] - BSA and Azide free (Capture) (ab281227)
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab265709 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 |
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WB |
Use at an assay dependent concentration. Predicted molecular weight: 12 kDa.
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Sanger Sequencing |
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: 12 kDa. |
Sanger Sequencing
Use at an assay dependent concentration. |
Images
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All lanes : Anti-S100A4 antibody [EPR14639(2)] (ab197896) at 1/1000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : S100A4 knockout HeLa cell lysate
Lane 3 : Wild-type A549 cell lysate
Lane 4 : S100A4 knockout A549 cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 12 kDa
Observed band size: 11 kDa why is the actual band size different from the predicted?False colour image of Western blot: Anti-S100A4 antibody [EPR14639(2)] staining at 1/1000 dilution, shown in green; loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) staining at 1/20000 dilution, shown in red. In Western blot, ab197896 was shown to bind specifically to S100A4. A band was observed at 11 kDa in wild-type HeLa and A549 cell lysates with no signal observed at this size in S100A4 knockout HeLa cell line ab265709 (knockout cell lysate ab257046) and S100A4 knockout A549 cell line ab261865 (knockout cell lysate ab261674). To generate this image, wild-type and S100A4 knockout HeLa and S100A4 knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.
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All lanes : Anti-S100A4 antibody [EPR2761(2)] (ab124805) at 1/1000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : S100A4 knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 12 kDa
Observed band size: 11 kDa why is the actual band size different from the predicted?Lanes 1 - 2: Merged signal (red and green). Green - ab124805 observed at 11 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.
ab124805 was shown to react with S100A4 in wild-type HeLa cells in Western blot with loss of signal observed in S100A4 knockout cell line ab265709 (S100A4 knockout cell lysate ab257046). Wild-type HeLa and S100A4 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab124805 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated 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 in 20000 dilution for 1 h at room temperature before imaging.
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All lanes : Anti-S100A4 antibody [EPR14639(2)] (ab197896) at 1/1000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : S100A4 knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 12 kDa
Observed band size: 11 kDa why is the actual band size different from the predicted?Lanes 1 - 2: Merged signal (red and green). Green - ab197896 observed at 11 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.
ab197896 was shown to react with S100A4 in wild-type HeLa cells in Western blot with loss of signal observed in S100A4 knockout cell line ab265709 (S100A4 knockout cell lysate ab257046). Wild-type HeLa and S100A4 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab197896 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated 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 in 20000 dilution for 1 h at room temperature before imaging.
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All lanes : Anti-S100A4 antibody [S100A4/1481] (ab218511) at 0.5 µg/ml
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : S100A4 knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 12 kDa
Observed band size: 11 kDa why is the actual band size different from the predicted?Lanes 1 - 2: Merged signal (red and green). Green - ab218511 observed at 11 kDa. Red - loading control ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) observed at 55 kDa.
ab218511 was shown to react with S100A4 in wild-type HeLa cells in Western blot with loss of signal observed in S100A4 knockout cell line ab265709 (S100A4 knockout cell lysate ab257046). Wild-type HeLa and S100A4 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab218511 and ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) overnight at 4 °C at 0.5 µg/ml and a 1 in 20000 dilution respectively. Blots were incubated 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 in 20000 dilution for 1 h at room temperature before imaging.
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All lanes : Anti-S100A4 antibody [S100A4/1482] (ab218512) at 0.5 µg/ml
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : S100A4 knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 12 kDa
Observed band size: 11 kDa why is the actual band size different from the predicted?Lanes 1 - 2: Merged signal (red and green). Green - ab218512 observed at 11 kDa. Red - loading control ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) observed at 55 kDa.
ab218512 was shown to react with S100A4 in wild-type HeLa cells in Western blot with loss of signal observed in S100A4 knockout cell line ab265709 (S100A4 knockout cell lysate ab257046). Wild-type HeLa and S100A4 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab218512 and ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) overnight at 4 °C at 0.5 µg/ml and a 1 in 20000 dilution respectively. Blots were incubated 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 in 20000 dilution for 1 h at room temperature before imaging.
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Allele-1: 5 bp deletion in exon 2.
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Allele-2: 1 bp insertion in exon 2.
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (0)
ab265709 has not yet been referenced specifically in any publications.