CCL7 KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 85 bp deletion in exon 2.
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Key facts
- Cell type
RAW 264.7
- Species or organism
Mouse
- Tissue
Lymphatic
- Form
Liquid
- Knockout validation
Sanger Sequencing, Western blot
- Mutation description
Knockout achieved by using CRISPR/Cas9, Homozygous: 85 bp deletion in exon 2
Alternative names
C-C motif chemokine 7, CCL7_HUMAN, Chemokine CC motif ligand 7, FIC, MARC, MCP-3, Monocyte chemoattractant protein 3, Monocyte chemotactic protein 3, NC28, RP23-350G1.4, SCYA6, SCYA7, Small-inducible cytokine A7
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CCL7 KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 85 bp deletion in exon 2.
Key facts
- Cell type
RAW 264.7
- Species or organism
Mouse
- Tissue
Lymphatic
- Form
Liquid
- Knockout validation
Sanger Sequencing, Western blot
- Mutation description
Knockout achieved by using CRISPR/Cas9, Homozygous: 85 bp deletion in exon 2
- Disease
Carcinoma
- Concentration
Properties
- Gene name
CCL7
- Gene editing type
Knockout
- Gene editing method
CRISPR technology
- Knockout validation
Sanger Sequencing, Western blot
- Zygosity
Homozygous
Cell culture
- Biosafety level
EU: 2 US: 2
- Adherent/suspension
Adherent
- Gender
Male
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 for bath 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 all remaining cells into an ultra-low attachment 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.
5. Once confluent passage into an appropriate flask at a density of 2x105 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
- Grow in ultra-low attachment flasks.
- Do not wash or use dissociation reagent. Cells are semi-adherent. Tap flask sharply to remove adhered cells.
- All seeding densities should be based on cell counts gained by established methods.
- A guide seeding density of 2x105 cells/mL is recommended.
- Cells should be passaged when they have achieved 80-90% confluence.
- Culture medium
DMEM + 10% FBS
- Cryopreservation medium
Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Storage
- Shipped at conditions
Dry Ice
- Appropriate long-term storage conditions
-196°C
Notes
Recommended control: Mouse wild-type RAW 264.7 cell line (Mouse wild-type RAW 264.7 cell line ab275474). 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.
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.
Supplementary info
This supplementary information is collated from multiple sources and compiled automatically.
- Activity summary
The target MCP-3 also referred to as CCL7 is a chemotactic cytokine belonging to the CC chemokine family. Its molecular weight is approximately 11 kDa. MCP-3 is largely expressed by various cell types including monocytes lymphocytes and endothelial cells. This protein plays an important role in immune system signaling. MCP-3 exhibits chemotactic activity for monocytes T lymphocytes and natural killer (NK) cells contributing to immune cell recruitment at sites of inflammation.
- Biological function summary
MCP-3 interacts with several chemokine receptors such as CCR1 CCR2 and CCR3 allowing for diverse roles in immune response and inflammation. It functions as both a monomer and in conjunction with other chemokines to form heterodimers aiding in the modulation and fine-tuning of the immune response. The presence of MCP-3 in inflammatory tissues indicates its contribution to directing leukocyte migration and positioning therefore supporting both innate and adaptive immunity.
- Pathways
MCP-3 is a component of the chemokine signaling pathway and is significant in the regulation of immune surveillance. It interacts closely with proteins such as CCL2 and CCL3 all of which bind to similar receptors like CCR2 and CCR5. MCP-3's involvement in these pathways supports cellular responses to inflammation and infection and its activity has a direct impact on the mobilization and activation of immune cells.
- Associated diseases and disorders
MCP-3 shows associations with conditions marked by inflammation and immune system dysregulation including rheumatoid arthritis and asthma. The protein's influence in these disorders links to other chemokines like MCP-1 (CCL2) which also recruits monocytes and contributes to chronic inflammation. Disruption in MCP-3 expression or function can aggravate disease states by altering the balance of leukocyte trafficking and exacerbating inflammatory responses.
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2 product images
Western blot - Mouse CCL7 knockout RAW 264.7 cell line (ab273755)
Lanes 1 - 6: Merged signal (red and green). Green - Anti-MCP3 antibody [EPR22649-155] ab228979 observed at 14 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.
Anti-MCP3 antibody [EPR22649-155] ab228979 was shown to react with MCP3 in RAW 264.7 wild-type cells in Western blot with loss of signal observed in CCL7 knockout sample. Wild-type and CCL7 knockout RAW 264.7 cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with Anti-MCP3 antibody [EPR22649-155] ab228979 and Anti-GAPDH antibody [6C5] - Loading Control ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4 ° 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 (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.
All lanes: Western blot - Anti-MCP3 antibody [EPR22649-155] (Anti-MCP3 antibody [EPR22649-155] ab228979) at 1/1000 dilution
Lane 1: Wild-type RAW 264.7 untreated control cell lysate at 30 µg
Lane 2: Wild-type RAW 264.7 PMA treated (80 nM, 24 h) plus LPS treated (100 ng/ml, 6 h) and Brefeldin A (Brefeldin A, Inhibitor of ADP-ribosylation factor ab120299) treated (5 µg/ml, 5 h) cell lysate at 30 µg
Lane 3: CCL7/MCP3 knockout RAW 264.7 untreated cell lysate at 30 µg
Lane 4: CCL7/MCP3 knockout RAW 264.7 PMA treated (80 nM, 24 h) plus LPS treated (100 ng/ml, 6 h) and Brefeldin A (Brefeldin A, Inhibitor of ADP-ribosylation factor ab120299) treated (5 µg/ml, 5 h) cell lysate at 30 µg
Lane 5: J774A.1 Glucose treated (138.8 mMol/L, 8 h) plus Brefeldin A treated (5 µg/ml, 6 h) and Brefeldin A (Brefeldin A, Inhibitor of ADP-ribosylation factor ab120299) treated (5 µg/ml, 5 h) cell lysate at 30 µg
Lane 6: J774A.1 Glucose treated (5.6 mMol/L, 8 h) plus Brefeldin A treated (5 µg/ml, 6 h) and Brefeldin A (Brefeldin A, Inhibitor of ADP-ribosylation factor ab120299) treated (5 µg/ml, 5 h) cell lysate at 30 µg
Performed under reducing conditions.
Predicted band size: 11 kDa
Observed band size: 14 kDa
Sanger Sequencing - Mouse CCL7 knockout RAW 264.7 cell line (ab273755)
Homozygous: 85 bp deletion in exon 2
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