AB280047

Mouse CD68 knockout RAW 264.7 cell line

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CD68 KO cell line available to order. KO validated by Western blot. Free of charge wild type control available. To order both knockout and wild-type control cells: select '2 x 1000000 Cells/vial'. To order only knockout cells: select '1000000 Cells/vial'.

3 Images

Lab

Western blot - Mouse CD68 knockout RAW 264.7 cell line (AB280047)

False colour image of Western blot : Anti-CD68 antibody staining at 0.2 μg/ml shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution shown in red. In Western blot ab125212 was shown to bind specifically to CD68. A band was observed at 95-102 kDa in wild-type RAW 264.7 cell lysates with no signal observed at this size in CD68 knockout cell line ab280047 (knockout cell lysate ab280106). To generate this image wild-type and CD68 knockout RAW 264.7 cell lysates were analysed. First samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with primary antibodies overnight at 4°. Blots were washed four times in TBS-T incubated with secondary antibodies for 1 h at room temperature washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-CD68 antibody (<a href='/en-us/products/primary-antibodies/cd68-antibody-ab125212'>ab125212</a>) at 0.2 µg/mL

Lane 1:

Wild-type RAW 264.7 cell lysate at 20 µg

Lane 2:

CD68 knockout RAW 264.7 cell lysate at 20 µg

Lane 2:

Western blot - Mouse CD68 knockout RAW 264.7 cell line (ab280047)

Lane 3:

Mouse spleen cell lysate at 20 µg

Lane 4:

Neuro-2a cell lysate at 20 µg

Predicted band size: 37 kDa

Observed band size: 95-102 kDa

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Lab

Western blot - Mouse CD68 knockout RAW 264.7 cell line (AB280047)

Western blot : Anti-CD68 antibody [RM1031] (ab303565) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab303565 was shown to bind specifically to CD68. A band was observed at 100 kDa in wild-type RAW 264.7 cell lysates with no signal observed at this size in CD68 knockout cell line. To generate this image, wild-type and CD68 knockout RAW 264.7 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5% milk in TBS-0.1% Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-CD68 antibody [RM1031] (<a href='/en-us/products/primary-antibodies/cd68-antibody-rm1031-ab303565'>ab303565</a>)

Lane 1:

Wild-type RAW 264.7 cell lysate at 20 µg

Lane 2:

CD68 knockout RAW 264.7 cell lysate at 20 µg

Observed band size: 100 kDa

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Sanger Sequencing - Mouse CD68 knockout RAW 264.7 cell line (AB280047)

Sanger seq

Lab

Sanger Sequencing - Mouse CD68 knockout RAW 264.7 cell line (AB280047)

Mouse Cd68 KO in Raw264.7 Cells with 28 bp insertion-13 bp deletion-135 bp insertion-72 bp deletion in Exon 2.

Key facts

Cell type

RAW 264.7

Species or organism

Mouse

Tissue

Lymphatic

Form

Liquid

form

Knockout validation

Sanger Sequencing,Western blot

Disease

Carcinoma

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Reactivity data

{ "title": "Reactivity Data", "filters": { "stats": ["", "Reactivity", "Dilution Info", "Notes"] }, "values": { "WB": { "reactivity":"TESTED_AND_REACTS", "dilution-info":"", "notes":"<p></p>" } } }

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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.

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What's included?

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Properties and storage information

Gene name

CD68

Gene editing type

Knockout

Gene editing method

CRISPR technology

Knockout validation

Sanger Sequencing, Western blot

Shipped at conditions

Dry Ice

Appropriate short-term storage conditions

-196°C

Appropriate long-term storage conditions

-196°C

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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.

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Supplementary information

This supplementary information is collated from multiple sources and compiled automatically.

CD68 is a glycoprotein that functions as a scavenger receptor. It plays an important role in the regulation of cellular debris and apoptotic cell clearance. Researchers often refer to it as KP1 or macrosialin in the literature. CD68 with a molecular weight of approximately 110 kDa is highly expressed in macrophages and monocytes. The mucin-like structure of CD68 allows it to be heavily glycosylated which aids in its function. Researchers detect CD68 in tissues through methods like immunohistochemistry (IHC) immunofluorescence and CD68 staining and it is widely used as a macrophage marker in research.

Biological function summary

CD68 facilitates functions associated with the innate immune response. It serves as an important part of macrophages which contribute to immune surveillance and tissue homeostasis. Though it primarily operates on its own in some contexts CD68 might assist in forming complexes with other receptors to modulate phagocytic activity. This target is significant in atherosclerotic plaque stability as macrophages engulf lipids and cell debris through processes facilitated by CD68.

Pathways

CD68 is involved in pathways connected to inflammation and phagocytosis. The CD68 protein works closely with other scavenger receptors like CD36 to mediate uptake of oxidized low-density lipoproteins (oxLDL) in the lipid metabolism pathway. Additionally CD68 engagement can influence the toll-like receptor (TLR) signaling pathways thereby linking innate immunity and inflammatory responses critical for host defense and disease progression.

CD68 is associated with atherosclerosis and multiple sclerosis. In atherosclerosis its role becomes evident as it accumulates in macrophages within the plaques making it a marker of disease severity. In multiple sclerosis CD68 expression in macrophages and microglia indicates active demyelination and inflammation. It also relates to proteins such as CD36 in atherosclerosis where both mediate the uptake of modified LDL contributing to foam cell formation in plaques.

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Cell culture

Biosafety level

EU: 2 US: 2

Adherent/suspension

Adherent

Gender

Male

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Product protocols

Visit the General protocols
Visit the Troubleshooting

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Target data

See full target information CD68

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Product promise

We are committed to supporting your work with high-quality reagents, and we're here for you every step of the way. In the unlikely event that one of our products does not perform as expected, you're protected by our Product Promise.

For full details, please see our Terms & Conditions

Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.

For licensing inquiries, please contact partnerships@abcam.com

Mouse CD68 knockout RAW 264.7 cell line

Western blot - Mouse CD68 knockout RAW 264.7 cell line (AB280047)

Western blot - Mouse CD68 knockout RAW 264.7 cell line (AB280047)

Sanger Sequencing - Mouse CD68 knockout RAW 264.7 cell line (AB280047)

Key facts

Cell type

Species or organism

Tissue

Form

Knockout validation

Disease

Complementary Products

Primary Antibodies

Primary Antibodies

Primary Antibodies

Primary Antibodies

Reactivity data

Product details

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

Subculture guidelines

Culture medium

Cryopreservation medium

Supplementary information

Biological function summary

Pathways

Cell culture

Biosafety level

Adherent/suspension

Gender

Product protocols

Target data

Product promise