Mouse CCL7 knockout RAW 264.7 cell line
- Advanced Validation
- What is this?
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- WB
Lab
Western blot - Mouse CCL7 knockout RAW 264.7 cell line (AB273755)
Lanes 1 - 6 : Merged signal (red and green). Green - ab228979 observed at 14 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.
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 ab228979 and 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 (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (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] (<a href='/en-us/products/primary-antibodies/mcp3-antibody-epr22649-155-ab228979'>ab228979</a>) 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 (<a href='/en-us/products/biochemicals/brefeldin-a-inhibitor-of-adp-ribosylation-factor-ab120299'>ab120299</a>) treated (5 µg/ml, 5 h) cell lysate at 30 µg
Lane 2:
Western blot - Mouse CCL7 knockout RAW 264.7 cell line (ab273755)
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 (<a href='/en-us/products/biochemicals/brefeldin-a-inhibitor-of-adp-ribosylation-factor-ab120299'>ab120299</a>) 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 (<a href='/en-us/products/biochemicals/brefeldin-a-inhibitor-of-adp-ribosylation-factor-ab120299'>ab120299</a>) 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 (<a href='/en-us/products/biochemicals/brefeldin-a-inhibitor-of-adp-ribosylation-factor-ab120299'>ab120299</a>) treated (5 µg/ml, 5 h) cell lysate at 30 µg
Predicted band size: 11 kDa
Observed band size: 14 kDa
false
- Sanger seq
Supplier Data
Sanger Sequencing - Mouse CCL7 knockout RAW 264.7 cell line (AB273755)
Homozygous : 85 bp deletion in exon 2
Complementary Products
![Immunoprecipitation - Anti-MCP3 antibody [EPR22649-155] (AB228979)](https://content.abcam.com/products/images/mcp3-antibody-epr22649-155-ab228979--immunoprecipitation-img4292.jpg)
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 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.
Supplementary information
This supplementary information is collated from multiple sources and compiled automatically.
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.
Cell culture
Biosafety level
EU: 2 US: 2
Adherent/suspension
Adherent
Gender
Male
Product promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
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