Human CDK5 knockout HEK-293T cell line
- Advanced Validation
- What is this?
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(0 Publication)
- ICC/IF
Supplier Data
Immunocytochemistry/ Immunofluorescence - Human CDK5 knockout HEK-293T cell line (AB266089)
ab40773 was shown to react with CDK5 in wild-type HEK293T cells in immunocytochemistry with loss of signal observed in CDK5 knockout cell line ab266089. Wild-type and knockout cells were mixed and pelleted at a 1 : 1 ratio on coverslips. The cells were fixed with 4% paraformaldehyde (15 min) then permeabilized with 0.1% Triton X-100 (10min) and then blocked with 1x PBS, 0.01% Triton X-100, 5% BSA, 5% NGS. The cells were then incubated with ab40773 at 1/250 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat anti-rabbit secondary antibody to (Alexa Fluor® 555) at 0.5 μg/ml. Acquisition of the green (wild-type), red (antibody staining) and far-red (knockout) channels was performed. Representative grayscale images of the red channel are shown. Wild-type and knockout cells are outlined with yellow and magenta dashed line, respectively. Schematic representation of the mosaic strategy used is shown on the bottom-right panel. Image was acquired with a Zeiss(LSM-880).
This data was provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.
- WB
Lab
Western blot - Human CDK5 knockout HEK-293T cell line (AB266089)
Lanes 1 - 4 : Merged signal (red and green). Green - ab21249 observed at 32 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55kDa.
ab21249 was shown to react with Cdk5 in wild-type HEK-293T cells in Western blot with loss of signal observed in CDK5 knockout cell line ab266089 (CDK5 knockout cell lysate ab257157). Wild-type HEK-293T and CDK5 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab21249 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.
All lanes:
Anti-CDK5 antibody (<a href='/en-us/products/unavailable/cdk5-antibody-ab21249'>ab21249</a>) at 1/1000 dilution
Lane 1:
Wild-type HEK-293T cell lysate at 20 µg
Lane 2:
CDK5 knockout HEK-293T cell lysate at 20 µg
Lane 2:
Western blot - Human CDK5 knockout HEK-293T cell line (ab266089)
Lane 3:
SH-SY5Y cell lysate at 20 µg
Lane 4:
MCF7 cell lysate at 20 µg
false
- Cell Culture
Unknown
Cell Culture - Human CDK5 knockout HEK-293T cell line (AB266089)
Representative images of CDK5 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 a EVOS XL Core microscope.
- Sanger seq
Unknown
Sanger Sequencing - Human CDK5 knockout HEK-293T cell line (AB266089)
Homozygous : 1 bp insertion in exon 1
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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
Zygosity
Shipped at conditions
Appropriate short-term storage conditions
Appropriate long-term storage conditions
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 2 US: 2
Adherent/suspension
Adherent
Gender
Female
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. <p>1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.<br>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.<br>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.<br>4. Incubate the culture at 37°C incubator with 5% CO<sub>2</sub>. Check the culture one day after revival and continue to check until 80% confluent. Media change can be given if needed.<br>5. Once confluent passage into an appropriate flask at a density of 2x10<sup>4</sup> cells/cm<sup>2</sup>. Seeding density is given as a guide only and should be scaled to align with individual lab schedules. Cultures should be monitored daily.</p>
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.
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