Human CTSH (Cathepsin H) knockout HEK-293T cell line
- Advanced Validation
- What is this?
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(0 Publication)
- WB
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Western blot - Human CTSH (Cathepsin H) knockout HEK-293T cell line (AB266245)
Lanes 1-3 : Merged signal (red and green). Green - ab88960 observed at 27 kDa. Red - loading control ab181602 observed at 36 kDa.
ab88960 Anti-Cathepsin H antibody [AF14D7] was shown to specifically react with Cathepsin H in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab266245 (knockout cell lysate ab258387) was used. Wild-type and Cathepsin H knockout samples were subjected to SDS-PAGE. ab88960 and Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) 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® 680RD) preadsorbed (ab216777) and Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes:
Anti-Cathepsin H antibody [AF14D7] (<a href='/en-us/products/unavailable/cathepsin-h-antibody-af14d7-ab88960'>ab88960</a>) at 1/1000 dilution
Lane 1:
Wild-type HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg
Lane 2:
CTSH knockout HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg
Lane 3:
Western blot - Human CTSH (Cathepsin H) knockout HEK-293T cell lysate (<a href='/en-us/products/cell-lysates/human-ctsh-cathepsin-h-knockout-hek-293t-cell-lysate-ab258387'>ab258387</a>) at 20 µg
Secondary
Lanes 1 - 3:
Western blot - Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-680rd-preadsorbed-ab216777'>ab216777</a>) at 1/10000 dilution
Lanes 1 - 3:
Western blot - Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-800cw-preadsorbed-ab216772'>ab216772</a>) at 1/20000 dilution
Observed band size: 37 kDa
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- Cell Culture
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Cell Culture - Human CTSH (Cathepsin H) knockout HEK-293T cell line (AB266245)
Representative images of CTSH 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 M5000 microscope.
- Sanger seq
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Sanger Sequencing - Human CTSH (Cathepsin H) knockout HEK-293T cell line (AB266245)
Homozygous : 11 bp deletion in exon3
Complementary Products
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 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.
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
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
As a member of the cathepsin family Cathepsin H contributes to the regulation of protein turnover and homeostasis. It performs a dual role as both an aminopeptidase and endopeptidase. In addition to its function in protein degradation the enzyme involves itself in antigen processing during immune responses. Reactivity of Cathepsin H in immune complexes shows its importance in shaping immune tolerance and responses.
Pathways
Cathepsin H interacts with the lysosomal degradation pathway and plays a role in the MHC class II-mediated antigen presentation. This pathway is pivotal for the regulation of immune responses. Relation to other proteins like Cathepsin L and Cathepsin B highlights Cathepsin H's complementary function in proteolytic processing within lysosomes. Its role and expression play an integral part in balancing protein synthesis and degradation cycles.
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'.
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