Human SCAMP1 knockout HeLa cell line
- Advanced Validation
- What is this?
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(0 Publication)
- WB
Supplier Data
Western blot - Human SCAMP1 knockout HeLa cell line (AB265567)
Lanes 1 - 2 : Merged signal (red and green). Green - ab3430 observed at 36 kDa. Red - loading control, ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa. ab3430 was shown to react with SCAMP1 in wild-type HeLa cells in western blot. The bands observed in SCAMP1 knockout cell line ab265567 (SCAMP1 knockout cell lysate ab258184) below 36 kDa may represent truncated forms and cleaved fragments. This has not been investigated further. HeLa wild-type and SCAMP1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab3430 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4°C at 1 µg/ml and a 1/20000 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 1/20000 dilution for 1 h at room temperature before imaging.
All lanes:
Western blot - Anti-SCAMP1 antibody (<a href='/en-us/products/primary-antibodies/scamp1-antibody-ab3430'>ab3430</a>) at 1 µg/mL
Lane 1:
Wild-type HeLa cell lysate at 20 µg
Lane 2:
Western blot - Human SCAMP1 knockout HeLa cell lysate (<a href='/en-us/products/cell-lysates/human-scamp1-knockout-hela-cell-lysate-ab258184'>ab258184</a>) at 20 µg
Secondary
Lanes 1 - 2:
Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution
Lanes 1 - 2:
Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution
Predicted band size: 38 kDa
Observed band size: 36 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human SCAMP1 knockout HeLa cell line (AB265567)
Allele-1 : 25 bp deletion in exon 2.
- Sanger seq
Unknown
Sanger Sequencing - Human SCAMP1 knockout HeLa cell line (AB265567)
Allele-2 : 1 bp insertion in exon 2.
Complementary Products
Reactivity data
Product details
We will provide viable cells that proliferate on revival.
Western blot data indicates that the CRISPR gene edit may have resulted in a truncation of the protein of interest. Please see data images.
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
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
SCAMP1 is engaged in the regulation of endocytosis and exocytosis within cellular processes. It is not part of a conventional complex but interacts dynamically with other proteins involved in vesicle fusion and release. SCAMP1 associates with cytosolic domains and membrane proteins to orchestrate the trafficking of vesicles ensuring efficient cellular communication and molecular transport across cellular membranes.
Pathways
Proteins related to vesicular transport such as SNAREs interact functionally with SCAMP1. Within this context SCAMP1 is actively involved in the endocytic and exocytic pathways which are important for maintaining cellular homeostasis and neurotransmitter release. These pathways are vital for the release of neurotransmitters during synaptic signaling and hormone secretion in endocrine systems with SCAMP1 aiding in the precise regulation of these events.
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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