AB273378

Human CD74 knockout Raji cell line

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Human CD74 knockout Raji cell line (ab273378) available to order. Recommended control: Human wild-type Raji cell line (ab275473).

- CD74 KO validation: Sanger Sequencing
- Concentration: 1 million cells/vial

View Alternative Names

CD74 antigen, CD74 antigen (invariant polypeptide of major histocompatibility complex, class II antigen-associated), CD74 molecule, CD74 molecule, major histocompatibility complex, class II invariant chain, DHLAG, Gamma chain of class II antigens, HG2A_HUMAN, HLA class II histocompatibility antigen gamma chain, HLA-DR antigens-associated invariant chain, HLA-DR-gamma, HLADG, Ia antigen-associated invariant chain, Ia associated invariant chain, Ia gamma, Ii, Invariant polypeptide of major histocompatibility complex class II antigen associated, MHC HLA-DR gamma chain, Major histocompatibility complex class II invariant chain, P35, Protein 41, la-gamma, p33

10 Images

Lab

Western blot - Human CD74 knockout Raji cell line (AB273378)

Lanes 1 - 4 : Merged signal (red and green). Green - ab270265 observed at 35 kDa. Red - loading control, ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.

ab270265 was shown to react with CD74 in western blot. The band observed in CD74 CRISPR/Cas9 edited cell line ab273378 (CRISPR/Cas9 edited lysate ab275529) below 35 kDa is likely to represent a truncated form. This has not been investigated further. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween^®) before incubation with ab270265 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at 1 μg/ml 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-CD74 antibody [CLIP/3127R] (<a href='/en-us/products/primary-antibodies/cd74-antibody-clip-3127r-ab270265'>ab270265</a>) at 1 µg/mL

Lane 1:

Wild-type Raji cell lysate at 30 µg

Lane 2:

CD74 knockout Raji cell lysate at 30 µg

Lane 2:

Western blot - Human CD74 knockout Raji cell line (ab273378)

Lane 3:

Jurkat cell lysate at 30 µg

Lane 4:

HepG2 cell lysate at 30 µg

Predicted band size: 34 kDa

Observed band size: 35 kDa

false

Flow Cytometry (Intracellular) - Human CD74 knockout Raji cell line (AB273378)

Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Human CD74 knockout Raji cell line (AB273378)

Flow cytometry overlay histogram showing wild-type Raji (green line) and CD74 knockout Raji cells (ab273378) stained with ab108393 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10μg/ml human IgG and 10% normal goat serum to block FC receptors and non-specific protein-protein interaction followed by the antibody (ab108393) (1x10⁶ in 100μl at 0.2 μg/ml) for 30 min at 22°C.

The secondary antibody Goat anti-rabbit IgG H&L (Alexa Fluor^® 488, pre-adsorbed) (ab150081) was used at 1/2000 for 30 min at 22°C.

Isotype control antibody was Rabbit IgG (monoclonal) (ab172730) used at the same concentration and conditions as the primary antibody (wild-type Raji cells - black line; CD74 knockout Raji cells ab273378 - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).

Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

Flow Cytometry - Human CD74 knockout Raji cell line (AB273378)

Flow Cyt

Lab

Flow Cytometry - Human CD74 knockout Raji cell line (AB273378)

Flow cytometry overlay histogram showing wild-type Raji (green line) and CD74 knockout Raji cells (ab273378) stained with ab270265 (red line). The cells were incubated in 1x PBS containing 10μg/ml human IgG and 10% normal goat serum to block FC receptors and non-specific protein-protein interaction followed by the antibody (ab270265) (1x10⁶ in 100μl at 1 μg/ml) for 30 min at 4°C.

The secondary antibody Goat anti-rabbit IgG H&L (Alexa Fluor^® 488 pre-adsorbed) (ab150081) was used at 1/2000 for 30 min at 4°C.

Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Human CD74 knockout Raji cell line (AB273378)

Flow cytometry overlay histogram showing wild-type Raji (green line) and CD74 knockout Raji cells (ab273378) stained with ab9514 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10μg/ml human IgG and 10% normal goat serum to block FC receptors and non-specific protein-protein interaction followed by the antibody (ab9514) (1x10⁶ in 100μl at 1 μg/ml) for 30 min at 22°C.

The secondary antibody Goat anti-mouse IgG H&L (Alexa Fluor^® 488, pre-adsorbed) (ab150117) was used at 1/2000 for 30 min at 22°C.

Isotype control antibody was mouse IgG1κ (ab170190) used at the same concentration and conditions as the primary antibody (wild-type Raji cells - black line; CD74 knockout Raji cells ab273378 - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).

Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

This antibody gave a positive signal in CD74 knockout Raji cells fixed with 4% formaldehyde (10 min) / permeabilized with 0.1% PBS-Triton X-100 for 15 min used under the same conditions.

Lab

Western blot - Human CD74 knockout Raji cell line (AB273378)

Lanes 1 - 4 : Merged signal (red and green). Green - ab9514 observed at 35 kDa. Red - loading control ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) observed at 55 kDa.

ab9514 was shown to react with CD74 in western blot. The band observed in CD74 knockout cell line ab273378 (knockout lysate ab275529) below 35 kDa is likely to represent a truncated form. This has not been investigated further. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab9514 and ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) overnight at 4 ° at 5 μg/ml and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Mouse IgG H&L (IRDye^® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye^® 680RD) preabsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.

All lanes:

Western blot - Anti-CD74 antibody [LN2] (<a href='/en-us/products/primary-antibodies/cd74-antibody-ln2-ab9514'>ab9514</a>) at 5 µg/mL

Lane 1:

Wild-type Raji cell lysate at 30 µg

Lane 2:

CD74 knockout Raji cell lysate at 30 µg

Lane 2:

Western blot - Human CD74 knockout Raji cell lysate (<a href='/en-us/products/cell-lysates/human-cd74-knockout-raji-cell-lysate-ab275529'>ab275529</a>)

Lane 2:

Western blot - Human CD74 knockout Raji cell line (ab273378)

Lane 3:

Jurkat cell lysate at 30 µg

Lane 4:

HepG2 cell lysate at 30 µg

Predicted band size: 34 kDa

Observed band size: 35 kDa

false

Lab

Western blot - Human CD74 knockout Raji cell line (AB273378)

All lanes:

Western blot - Anti-CD74 antibody [PIN.1] (<a href='/en-us/products/primary-antibodies/cd74-antibody-pin1-ab22603'>ab22603</a>) at 1 µg/mL

Lane 1:

Wild-type Raji cell lysate at 30 µg

Lane 2:

Western blot - Human CD74 knockout Raji cell lysate (<a href='/en-us/products/cell-lysates/human-cd74-knockout-raji-cell-lysate-ab275529'>ab275529</a>)

Lane 2:

CD74 knockout Raji cell lysate at 30 µg

Lane 2:

Western blot - Human CD74 knockout Raji cell line (ab273378)

Lane 3:

Jurkat cell lysate at 30 µg

Lane 4:

HepG2 cell lysate at 30 µg

Predicted band size: 34 kDa

Observed band size: 35 kDa

false

Lab

Western blot - Human CD74 knockout Raji cell line (AB273378)

Lanes 1 - 4 : Merged signal (red and green). Green - ab64772 observed at 35 kDa. Red - loading control, ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.

ab64772 was shown to react with CD74 in western blot. The band observed in CD74 knockout cell line ab273378 (knockout lysate ab275529) below 35 kDa is likely to represent a truncated form. This has not been investigated further. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab64772 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at 1 μg/ml 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-CD74 antibody (<a href='/en-us/products/primary-antibodies/cd74-antibody-ab64772'>ab64772</a>) at 1 µg/mL

Lane 1:

Wild-type Raji cell lysate at 30 µg

Lane 2:

CD74 knockout Raji cell lysate at 30 µg

Lane 2:

Western blot - Human CD74 knockout Raji cell lysate (<a href='/en-us/products/cell-lysates/human-cd74-knockout-raji-cell-lysate-ab275529'>ab275529</a>)

Lane 2:

Western blot - Human CD74 knockout Raji cell line (ab273378)

Lane 3:

Jurkat cell lysate at 30 µg

Lane 4:

HepG2 cell lysate at 30 µg

Predicted band size: 34 kDa

Observed band size: 35 kDa

false

Lab

Western blot - Human CD74 knockout Raji cell line (AB273378)

Lanes 1 - 4 : Merged signal (red and green). Green - ab108393 observed at 35 kDa. Red - loading control, ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.

ab108393 was shown to react with CD74 in western blot. The band observed in CD74 knockout cell line ab273378 (knockout lysate ab275529) below 35 kDa is likely to represent a truncated form. This has not been investigated further. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab108393 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C 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-CD74 antibody [EPR4064] (<a href='/en-us/products/primary-antibodies/cd74-antibody-epr4064-ab108393'>ab108393</a>) at 1/1000 dilution

Lane 1:

Wild-type Raji cell lysate at 30 µg

Lane 2:

Western blot - Human CD74 knockout Raji cell lysate (<a href='/en-us/products/cell-lysates/human-cd74-knockout-raji-cell-lysate-ab275529'>ab275529</a>)

Lane 2:

CD74 knockout Raji cell lysate at 30 µg

Lane 2:

Western blot - Human CD74 knockout Raji cell line (ab273378)

Lane 3:

Jurkat cell lysate at 30 µg

Lane 4:

HepG2 cell lysate at 30 µg

Predicted band size: 34 kDa

Observed band size: 35 kDa

false

Flow Cyt

Lab

Flow Cytometry - Human CD74 knockout Raji cell line (AB273378)

Flow cytometry overlay histogram showing wild-type Raji (green line) and CD74 knockout Raji cells (ab273378) stained with ab22606 (red line). The cells were incubated in 1x PBS containing 10μg/ml human IgG and 10% normal goat serum to block FC receptors and non-specific protein-protein interaction followed by the antibody (ab22606) (1x10⁶ in 100μl at 5 μg/ml) for 30 min at 4°C.

The secondary antibody Goat anti-mouse IgG H&L (Alexa Fluor^® 488, pre-adsorbed) (ab150117) was used at 1/2000 for 30 min at 4°C.

Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

Sanger Sequencing - Human CD74 knockout Raji cell line (AB273378)

Sanger seq

Supplier Data

Sanger Sequencing - Human CD74 knockout Raji cell line (AB273378)

Homozygous : 13 bp deletion in exon 2

Key facts

Cell type

Raji

Species or organism

Human

Tissue

Lymphatic

Form

Liquid

form

Knockout validation

Sanger Sequencing

Mutation description

Knockout achieved by using CRISPR/Cas9, Homozygous: 13 bp deletion in exon 2

Disease

Lymphoma

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Reactivity data

{ "title": "Reactivity Data", "filters": { "stats": ["", "Reactivity", "Dilution Info", "Notes"] }, "values": { "Flow Cyt (Intra)": { "reactivity":"TESTED_AND_REACTS", "dilution-info":"", "notes":"" }, "WB": { "reactivity":"TESTED_AND_REACTS", "dilution-info":"", "notes":"Western blot data indicates that the CRISPR gene edit may have resulted in a truncation of the protein of interest. Please see data images." }, "Flow Cyt": { "reactivity":"TESTED_AND_REACTS", "dilution-info":"", "notes":"" } } }

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Product details

We will provide viable cells that proliferate on revival.

Western blot data indicates that the CRISPR gene edit may have resulted in a truncation of the protein of interest. Please see data images.

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

CD74

Gene editing type

Knockout

Gene editing method

CRISPR technology

Knockout validation

Sanger Sequencing

Zygosity

Homozygous

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. Based on cell count, seed cells in an appropriate cell culture flask at a density of 4x10⁵ - 5x10⁵ cells/mL(for initial passages it is recomended to culture the cells in the higher range of recomended seeding density). 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 4x10⁵ cells/mL is recommended.
A maximum of 3x10⁶ viable cells/mL is obtainable.

Culture medium

RPMI + 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.

CD74 also known as the invariant chain or li is a protein with a mass of about 33 kDa. It is widely expressed on the surface of immune cells such as B cells dendritic cells and macrophages. CD74 plays an important role in the immune system by acting as a chaperone for major histocompatibility complex class II (MHC II) molecules guiding their migration to endosomes where they encounter antigenic peptides. The CD74 protein also functions as a cell surface receptor for macrophage migration inhibitory factor (MIF) enhancing the immune response process.

Biological function summary

CD74 facilitates multiple immune regulatory processes. It forms a complex with the MHC II molecules aiding in their proper folding and peptide loading in the endoplasmic reticulum. This complex then travels through the Golgi apparatus towards the lysosomal compartments where antigens are presented to initiate adaptive immune responses. In addition CD74 is involved in signal transduction pathways that regulate cell proliferation and survival influenced by interactions with MIF and other molecules.

Pathways

The role of CD74 extends into both the antigen presentation and MIF signaling pathways. Within the antigen presentation pathway CD74's activity is tightly connected with MHC II and subsequently to CD4+ T cells facilitating the activation of these immune cells. The MIF signaling pathway involves CD74 interacting with CD44 a receptor that further propagates the signaling cascades essential for immune modulation and cell survival.

CD74 is linked to inflammatory and autoimmune conditions such as rheumatoid arthritis and systemic lupus erythematosus. Aberrant expression of CD74 influences the presentation of autoantigens and may contribute to the chronic inflammation seen in these disorders. Its connection to MIF which is a pro-inflammatory cytokine further implicates CD74 in these autoimmune scenarios by promoting persistent immune activation. Researchers are also exploring CD74 as a therapeutic target in certain cancers due to its influence on immune surveillance and tumor progression.

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Quality control

STR analysis

CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX

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Cell culture

Biosafety level

EU: 2 US: 2

Adherent/suspension

Suspension

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 CD74

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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

Human CD74 knockout Raji cell line

Western blot - Human CD74 knockout Raji cell line (AB273378)

Flow Cytometry (Intracellular) - Human CD74 knockout Raji cell line (AB273378)

Flow Cytometry - Human CD74 knockout Raji cell line (AB273378)

Flow Cytometry (Intracellular) - Human CD74 knockout Raji cell line (AB273378)

Western blot - Human CD74 knockout Raji cell line (AB273378)

Western blot - Human CD74 knockout Raji cell line (AB273378)

Western blot - Human CD74 knockout Raji cell line (AB273378)

Western blot - Human CD74 knockout Raji cell line (AB273378)

Flow Cytometry - Human CD74 knockout Raji cell line (AB273378)

Sanger Sequencing - Human CD74 knockout Raji cell line (AB273378)

Key facts

Cell type

Species or organism

Tissue

Form

Knockout validation

Mutation description

Disease

Reactivity data

Product details

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

Subculture guidelines

Culture medium

Cryopreservation medium

Supplementary information

Biological function summary

Pathways

Quality control

STR analysis

Cell culture

Biosafety level

Adherent/suspension

Gender

Product protocols

Target data

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Product promise