Human TSPO (PBR) knockout HCT116 cell line
- Advanced Validation
- What is this?
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- WB
Lab
Western blot - Human TSPO (PBR) knockout HCT116 cell line (AB266878)
Lanes 1- 2 : Merged signal (red and green). Green - ab109497 observed at 17 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.
ab109497 was shown to react with PBR in wild-type HCT116 cells in western blot. Loss of signal was observed when knockout cell line ab266878 (knockout cell lysate ab257067) was used. Wild-type HCT116 and TSPO knockout HCT116 cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab109497 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4° at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye®800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye®680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes:
Western blot - Anti-PBR antibody [EPR5384] (<a href='/en-us/products/primary-antibodies/pbr-antibody-epr5384-ab109497'>ab109497</a>) at 1/1000 dilution
Lane 1:
Wild-type HCT116 cell lysate at 20 µg
Lane 2:
Western blot - Human TSPO (PBR) knockout HCT116 cell lysate (<a href='/en-us/products/cell-lysates/human-tspo-pbr-knockout-hct116-cell-lysate-ab257067'>ab257067</a>) at 20 µg
Predicted band size: 19 kDa
Observed band size: 17 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human TSPO (PBR) knockout HCT116 cell line (AB266878)
Homozygous : Insertion of the selection cassette in exon2
Complementary Products
Primary Antibodies
AB109497
RabMAb|Recombinant|KO Validated|20ul selling size
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PBR antibody [EPR5384] (AB109497)](https://content.abcam.com/products/images/pbr-antibody-epr5384-ab109497--immunohistochemistry-formalin-pfa-fixed-paraffin-embedded-sections-img454145.jpg)
- 5
- 15
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
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
McCoY5a + 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
PBR interacts with cholesterol for the synthesis of steroid hormones making it important for steroidogenesis. PBR is a part of the Mitochondrial Permeability Transition Pore complex (MPTP) involved in regulating the transport of molecules across the mitochondrial membrane. Through its association with the MPTP PBR plays a significant role in mitochondrial functions such as apoptosis and energy metabolism. The interaction with other molecules also includes the binding with benzodiazepines impacting processes like immune response and cell proliferation.
Pathways
PBR plays a role in the steroid biosynthesis and apoptosis pathways. It interfaces with the StAR (Steroidogenic Acute Regulatory) protein to facilitate cholesterol transport into mitochondria the initial step in steroid hormone production. PBR is also involved in pathways that regulate apoptosis and mitochondrial function linking it to different cellular processes through interactions with proteins like VDAC (Voltage-Dependent Anion Channel).
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 1 US: 1
Adherent/suspension
Adherent
Gender
Male
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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