AB266949

Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line

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IDO1 KO cell line available to order. KO validated by Western blot. Free of charge wild type control available. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp deletion in exon 3. To order both knockout and wild-type control cells: select '2 x 1000000 Cells/vial'. To order only knockout cells: select '1000000 Cells/vial'.

8 Images

Lab

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Western blot : Anti-IDO1 antibody [EPR28349-89] (ab311847) staining at 1/1000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab311847 was shown to bind specifically to IDO1. A band was observed at 42 kDa in treated wild-type A549 cell lysates with no signal observed at this size in IDO1 knockout cell line. To generate this image, wild-type and IDO1 knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % 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-Indoleamine 2, 3-dioxygenase antibody [EPR28349-89] (<a href='/en-us/products/primary-antibodies/indoleamine-2-3-dioxygenase-antibody-epr28349-89-ab311847'>ab311847</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 Treated IFN gamma (25 ng/mL, 48 h) ab281500 cell lysate at 20 µg

Lane 2:

Wild-type A549 Vehicle Control IFN gamma (0 ng/mL, 48 h) ab255554 cell lysate at 20 µg

Lane 3:

IDO1 knockout A549 Treated IFN gamma (25 ng/mL, 48 h) ab281489 cell lysate at 20 µg

Lane 4:

IDO1 knockout A549 Vehicle Control IFN gamma (0 ng/mL, 48 h) ab263515 cell lysate at 20 µg

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Predicted band size: 45 kDa

Observed band size: 42 kDa

false

Lab

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Lanes 1 - 6 : Merged signal (red and green). Green - ab211017 observed at 40 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.

ab211017 was shown to react with Indoleamine 2, 3-dioxygenase in treated wild-type A549 cells in Western blot with no signal observed in treated IDO1 knockout cell line ab266949 (IDO1 knockout cell lysate ab256948). Wild-type A549 and IDO1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween^®) before incubation with ab211017 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-Indoleamine 2, 3-dioxygenase antibody [EPR20374] (<a href='/en-us/products/primary-antibodies/indoleamine-2-3-dioxygenase-antibody-epr20374-ab211017'>ab211017</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 Vehicle Control IFN gamma (0 ng/ml, 48 h) cell lysate at 20 µg

Lane 2:

Wild-type A549 Treated IFN gamma (25 ng/ml, 48 h) cell lysate at 20 µg

Lane 2:

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (ab266949)

Lane 3:

IDO1 knockout A549 Vehicle Control IFN gamma (0 ng/ml, 48 h) cell lysate at 20 µg

Lane 4:

IDO1 knockout A549 Treated IFN gamma (25 ng/ml, 48 h) cell lysate at 20 µg

Lane 5:

SK-OV-3 cell lysate at 20 µg

Lane 6:

MCF7 cell lysate at 20 µg

Predicted band size: 45 kDa

Observed band size: 40 kDa

false

Lab

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Lanes 1 - 6 : Merged signal (red and green). Green - ab156787 observed at 40 kDa. Red - loading control ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) observed at 55 kDa.

ab156787 was shown to react with Indoleamine 2, 3-dioxygenase in treated wild-type A549 cells in Western blot with no signal observed in treated IDO1 knockout cell line ab266949 (IDO1 knockout cell lysate ab256948). Wild-type A549 and IDO1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween^®) before incubation with ab156787 and ab52866 (Rabbit anti-alpha Tubulin antibody [EP1332Y]) overnight at 4 °C at a 1 in 4000 dilution 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-Indoleamine 2, 3-dioxygenase antibody [OTI1A3] (<a href='/en-us/products/primary-antibodies/indoleamine-2-3-dioxygenase-antibody-oti1a3-ab156787'>ab156787</a>) at 1/4000 dilution

Lane 1:

Wild-type A549 Vehicle Control IFN gamma (0 ng/ml, 48 h) cell lysate at 20 µg

Lane 2:

Wild-type A549 Treated IFN gamma (25 ng/ml, 48 h) cell lysate at 20 µg

Lane 2:

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (ab266949)

Lane 3:

IDO1 knockout A549 Vehicle Control IFN gamma (0 ng/ml, 48 h) cell lysate at 20 µg

Lane 4:

IDO1 knockout A549 Treated IFN gamma (25 ng/ml, 48 h) cell lysate at 20 µg

Lane 5:

SK-OV-3 cell lysate at 20 µg

Lane 6:

MCF7 cell lysate at 20 µg

Predicted band size: 45 kDa

Observed band size: 40 kDa

false

Lab

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Lanes 1 - 6 : Merged signal (red and green). Green - ab76157 observed at 40 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.

ab76157 was shown to react with Indoleamine 2, 3-dioxygenase in treated wild-type A549 cells in Western blot with loss of signal observed in treated IDO1 knockout cell line ab266949 (IDO1 knockout cell lysate ab256948). Wild-type A549 and IDO1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween^®) before incubation with ab76157 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at a 1 in 2500 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-Indoleamine 2, 3-dioxygenase antibody [EPR1230Y] (<a href='/en-us/products/primary-antibodies/indoleamine-2-3-dioxygenase-antibody-epr1230y-ab76157'>ab76157</a>) at 1/2500 dilution

Lane 1:

Wild-type A549 Vehicle Control IFN gamma (0 ng/ml, 48 h) cell lysate at 20 µg

Lane 2:

Wild-type A549 Treated IFN gamma (25 ng/ml, 48 h) cell lysate at 20 µg

Lane 3:

IDO1 knockout A549 Vehicle Control IFN gamma (0 ng/ml, 48 h) cell lysate at 20 µg

Lane 4:

IDO1 knockout A549 Treated IFN gamma (25 ng/ml, 48 h) cell lysate at 20 µg

Lane 5:

SK-OV-3 cell lysate at 20 µg

Lane 6:

MCF7 cell lysate at 20 µg

Secondary

Lanes 1 - 6:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Lanes 1 - 6:

Western blot - Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>) at 1/20000 dilution

Predicted band size: 45 kDa

Observed band size: 40 kDa,55 kDa

false

Lab

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Western blot : Anti-IDO1 antibody [EPR1230Y] (ab76157) staining at 1/2500 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab76157 was shown to bind specifically to IDO1. A band was observed at 42/56 kDa in treated wild-type A549 cell lysates with no signal observed at this size in IDO1 knockout cell line. To generate this image, wild-type and IDO1 knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % 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:

Lane 1:

Wild-type A549 Treated IFN gamma (25 ng/mL, 48 h) ab281500 cell lysate at 20 µg

Lane 2:

Wild-type A549 Vehicle Control IFN gamma (0 ng/mL, 48 h) ab255554 cell lysate at 20 µg

Lane 3:

IDO1 knockout A549 Treated IFN gamma (25 ng/mL, 48 h) ab281489 cell lysate at 20 µg

Lane 4:

IDO1 knockout A549 Vehicle Control IFN gamma (0 ng/mL, 48 h) ab263515 cell lysate at 20 µg

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Predicted band size: 45 kDa

Observed band size: 42 kDa,56 kDa

false

Lab

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Western blot : Anti-IDO1 antibody [EPR24032-22] (ab277522) staining at 1/1000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab277522 was shown to bind specifically to IDO1. A band was observed at 35/42 kDa in treated wild-type A549 cell lysates with no signal observed at this size in IDO1 knockout cell line. To generate this image, wild-type and IDO1 knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % 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-Indoleamine 2, 3-dioxygenase antibody [EPR24032-22] (<a href='/en-us/products/primary-antibodies/indoleamine-2-3-dioxygenase-antibody-epr24032-22-ab277522'>ab277522</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 Treated IFN gamma (25 ng/mL, 48 h) ab281500 cell lysate at 20 µg

Lane 2:

Wild-type A549 Vehicle Control IFN gamma (0 ng/mL, 48 h) ab255554 cell lysate at 20 µg

Lane 3:

IDO1 knockout A549 Treated IFN gamma (25 ng/mL, 48 h) ab281489 cell lysate at 20 µg

Lane 4:

IDO1 knockout A549 Vehicle Control IFN gamma (0 ng/mL, 48 h) ab263515 cell lysate at 20 µg

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Predicted band size: 45 kDa

Observed band size: 35 kDa,42 kDa

false

Lab

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Western blot : Anti-IDO1 antibody [SP260] (ab228468) staining at 1/1000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab228468 was shown to bind specifically to IDO1. A band was observed at 42 kDa in treated wild-type A549 cell lysates with no signal observed at this size in IDO1 knockout cell line. To generate this image, wild-type and IDO1 knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % 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-Indoleamine 2, 3-dioxygenase antibody [SP260] - C-terminal (<a href='/en-us/products/primary-antibodies/indoleamine-2-3-dioxygenase-antibody-sp260-c-terminal-ab228468'>ab228468</a>) at 1/1000 dilution

Lane 1:

Wild-type A549 Treated IFN gamma (25 ng/mL, 48 h) ab281500 cell lysate at 20 µg

Lane 2:

Wild-type A549 Vehicle Control IFN gamma (0 ng/mL, 48 h) ab255554 cell lysate at 20 µg

Lane 3:

IDO1 knockout A549 Treated IFN gamma (25 ng/mL, 48 h) ab281489 cell lysate at 20 µg

Lane 4:

IDO1 knockout A549 Vehicle Control IFN gamma (0 ng/mL, 48 h) ab263515 cell lysate at 20 µg

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Predicted band size: 45 kDa

Observed band size: 42 kDa

false

Sanger Sequencing - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Sanger seq

Unknown

Sanger Sequencing - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Homozygous : 1 bp deletion in exon3

Key facts

Cell type

A549

Species or organism

Human

Tissue

Lung

Form

Liquid

form

Knockout validation

Sanger Sequencing,Western blot

Mutation description

Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp deletion in exon 3

Antibiotic resistance

Puromycin 1µg/mL

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>" }, "Sanger seq": { "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?

{ "values": { "2x1000000Cellsvial": { "sellingSize": "2 x 1000000 Cells/vial", "publicAssetCode":"ab266949-2x1000000Cells_vial", "assetComponentDetails": [ { "size":"1 x 1000000 Cells/vial", "name":"ab266949 Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line", "number":"AB266949-CMP01" }, { "size":"1 x 1000000 Cells/vial", "name":"ab255450 Human wild-type A549 cell line", "number":"AB266949-CMP02" } ] }, "1000000Cellsvial": { "sellingSize": "1000000 Cells/vial", "publicAssetCode":"ab266949-1000000Cells_vial", "assetComponentDetails": [ { "size":"1 x 1000000 Cells/vial", "name":"ab266949 Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line", "number":"AB266949-CMP01", "productcode":"" } ] } } }

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Properties and storage information

Gene name

IDO1

Gene editing type

Knockout

Gene editing method

CRISPR technology

Knockout validation

Sanger Sequencing, Western blot

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 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 seed all remaining cells into a 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. Media change can be given if needed.
5. Once confluent passage into an appropriate 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. 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.
Cells should be passaged when they have achieved 80-90% confluence.
Do not allow the cell density to exceed 7x10⁴ cells/cm².

Culture medium

F-12K + 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.

Indoleamine 2 3-dioxygenase (IDO) also known as IDO1 is an enzyme involved in the catabolism of tryptophan to kynurenine. It weighs approximately 45 kDa and it is an important target in immunoregulatory processes. IDO is highly expressed in antigen-presenting cells such as dendritic cells and macrophages as well as in various tumor cells where it helps modulate immune responses. This enzyme can also be measured by using assays like IDO ELISA.

Biological function summary

IDO plays a role in immune response regulation by degrading tryptophan an essential amino acid needed for T-cell proliferation. The depletion of tryptophan and the accumulation of its metabolites such as kynurenine cause immunosuppressive effects within the tissue microenvironment. While IDO functions mostly as a standalone enzyme its activity influences the cellular surroundings by altering the balance of local immune reactions.

Pathways

IDO integrates into the tryptophan metabolism pathway and is important in the kynurenine pathway. Through this pathway it maintains immune homeostasis and cell defense mechanisms. Proteins like kynureninase and kynurenine 3-monooxygenase also participate in the same metabolic processes which further extend the effects of tryptophan metabolism on immune cell behavior.

IDO is linked with cancer and chronic inflammatory conditions. In cancer elevated IDO expression suppresses anti-tumor immunity aiding tumor cells to evade immune surveillance. It also plays a role in autoimmune disorders by regulating excessive immune activity. During cancer progression IDO often works in conjunction with immune checkpoint proteins like PD-L1 which further enhances its immunosuppressive capabilities within the tumor microenvironment.

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Quality control

STR analysis

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

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Cell culture

Biosafety level

EU: 1 US: 1

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 IDO1

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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 IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Western blot - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Sanger Sequencing - Human IDO1 (Indoleamine 2, 3-dioxygenase) knockout A549 cell line (AB266949)

Key facts

Cell type

Species or organism

Tissue

Form

Knockout validation

Mutation description

Antibiotic resistance

Disease

Complementary Products

Assay Kits

Primary Antibodies

Proteins & Peptides

Assay Kits

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

Product promise