Human PPIF (Cyclophilin F) knockout HEK-293T cell line (ab266077)
Overview
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Product name
Human PPIF (Cyclophilin F) knockout HEK-293T cell line -
Parental Cell Line
HEK293T -
Organism
Human -
Mutation description
Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 1 -
Passage number
<20 -
Knockout validation
Sanger Sequencing, Western Blot (WB) -
Tested applications
Suitable for: WBmore details -
Biosafety level
2 -
General notes
Recommended control: Human wild-type HEK293T cell line (ab255449). 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.
Properties
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Number of cells
1 x 106 cells/vial, 1 mL -
Viability
~80% -
Adherent /Suspension
Adherent -
Tissue
Kidney -
Cell type
epithelial -
STR Analysis
Amelogenin X D5S818: 8, 9 D13S317: 12, 14 D7S820: 11 D16S539: 9, 13 vWA: 16, 19 TH01: 7, 9.3 TPOX: 11 CSF1PO: 11, 12 -
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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Function
PPIases accelerate the folding of proteins. It catalyzes the cis-trans isomerization of proline imidic peptide bonds in oligopeptides. -
Sequence similarities
Belongs to the cyclophilin-type PPIase family.
Contains 1 PPIase cyclophilin-type domain. -
Cellular localization
Mitochondrion matrix. - Information by UniProt
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Form
This gene encodes a 178 aa mature protein that is found in the mitochondrion and may participate in the permeability transition pore. While technically this protein is Cyclophilin F, literature references commonly refer to this protein as 'cyclophilin D’ or ‘CypD’. A different cytoplasmic protein of 370 aa, represented by Entrez GeneID 5481, is identified as Cyclophilin D. This antibody does not react with this 370 aa cytoplasmic protein.
Associated products
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KO cell lysates
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab266077 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: 22 kDa.
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Notes |
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WB
Use at an assay dependent concentration. Predicted molecular weight: 22 kDa. |
Images
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All lanes : Anti-Cyclophilin F antibody [E11AE12BD4] (ab110324) at 1/10000 dilution
Lane 1 : Wild-type HEK-293T cell lysate
Lane 2 : PPIF knockout HEK-293T cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 22 kDa
Observed band size: 23 kDa why is the actual band size different from the predicted?Lanes 1- 2: Merged signal (red and green). Green - ab110324 observed at 23 kDa. Red - Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) observed at 37 kDa.
ab110324 was shown to react with Cyclophilin F in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266077 (knockout cell lysate ab257039) was used. Wild-type HEK-293T and PPIF knockout HEK-293T 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. ab110324 and Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) overnight at 4°C at a 1 in 10000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye®800CW) preadsorbed (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye®680RD) preadsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
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All lanes : Anti-Cyclophilin F antibody [EPR11311-121] (ab231155) at 1/1000 dilution
Lane 1 : Wild-type HEK-293T cell lysate
Lane 2 : PPIF knockout HEK-293T cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 22 kDa
Observed band size: 23 kDa why is the actual band size different from the predicted?Lanes 1- 2: Merged signal (red and green). Green - ab231155 observed at 23 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.
ab231155 was shown to react with Cyclophilin F in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266077 (knockout cell lysate ab257039) was used. Wild-type HEK-293T and PPIF knockout HEK-293T 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. ab231155 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C 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.
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Homozygous: 1 bp insertion in exon 1
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (0)
ab266077 has not yet been referenced specifically in any publications.