AB273759

Human ANPEP (CD13) knockout THP-1 cell line

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

別名を表示する

AMPN_HUMAN, ANPEP, AP-M, AP-N, Alanyl (membrane) aminopeptidase, Alanyl aminopeptidase, Aminopeptidase M, Aminopeptidase N, CD13 antigen, LAP 1, Microsomal aminopeptidase, Myeloid plasma membrane glycoprotein CD13, P150, PEPN, gp150, hAPN

10 Images

Immunocytochemistry/ Immunofluorescence - Human ANPEP (CD13) knockout THP-1 cell line (AB273759)

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Immunocytochemistry/ Immunofluorescence - Human ANPEP (CD13) knockout THP-1 cell line (AB273759)

ab7417 staining CD13 in wild-type THP-1 cells (top panel) and ANPEP knockout THP-1 cells (bottom panel) (ab273759). The cells were fixed with 4% paraformaldehyde (10 min) then permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab7417 at 2.5μg/ml concentration and ab6046 (Rabbit polyclonal to beta Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to mouse IgG (Alexa Fluor^® 488) (ab150117) at 2 μg/ml (shown in green) and a goat secondary antibody to rabbit IgG (Alexa Fluor^® 594) (ab150080) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).

This image was generated from the hybridoma version of the product.

ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Human ANPEP (CD13) knockout THP-1 cell line (AB273759)

ab108382 staining CD13 in wild-type THP-1 cells (top panel) and ANPEP knockout THP-1 cells (bottom panel) (ab273759). The cells were fixed with 100% methanol (5 min) then permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab108382 at 1/1000 dilution and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor^® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor^® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).

ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Human ANPEP (CD13) knockout THP-1 cell line (AB273759)

ab108310 staining CD13 in wild-type THP-1 cells (top panel) and ANPEP knockout THP-1 cells (bottom panel) (ab273759). The cells were fixed with 4% paraformaldehyde (10 min) then permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab108310 at 1/500 dilution and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor^® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor^® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).

Flow Cytometry - Human ANPEP (CD13) knockout THP-1 cell line (AB273759)

Flow Cyt

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Flow Cytometry - Human ANPEP (CD13) knockout THP-1 cell line (AB273759)

Flow cytometry overlay histogram showing wild-type THP1 (green line) and ANPEP knockout THP1 cells (ab273759) stained with ab20136 (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 (ab20136) (1x10⁶ in 100μl at 1 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.

Isotype control antibody was mouse IgG2aκ (ab18413) used at the same concentration and conditions as the primary antibody (wild-type THP1 cells - black line; ANPEP knockout THP1 cells ab273759 - 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 Cyt

Lab

Flow Cytometry - Human ANPEP (CD13) knockout THP-1 cell line (AB273759)

Flow cytometry overlay histogram showing wild-type THP1 (green line) and ANPEP knockout THP1 cells (ab273759) stained with ab7417 (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 (ab7417) (1x10⁶ in 100μl at 1 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.

Isotype control antibody was mouse IgG1κ (ab170190) used at the same concentration and conditions as the primary antibody (wild-type THP1 cells - black line; ANPEP knockout THP1 cells ab273759 - 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 image was generated from the hybridoma version of the product.

ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Human ANPEP (CD13) knockout THP-1 cell line (AB273759)

ab227663 staining CD13 in wild-type THP-1 cells (top panel) and ANPEP knockout THP-1 cells (bottom panel) (ab273759). The cells were fixed with 4% paraformaldehyde (10 min) then permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab227663 at 1μg/ml concentration and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor^® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor^® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).

Lab

Western blot - Human ANPEP (CD13) knockout THP-1 cell line (AB273759)

Lanes 1 - 4 : Merged signal (red and green). Green - ab108310 observed at 160 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.

ab108310 was shown to react with CD13 in wild-type THP-1 cells in western blot with loss of signal observed in ANPEP knockout cell line ab273759 (knockout cell lysate ab275505). Wild-type and ANPEP knockout THP-1 cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab108310 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 hour at room temperature before imaging.

All lanes:

Western blot - Anti-CD13 antibody [EPR4058] (<a href='/products/primary-antibodies/cd13-antibody-epr4058-ab108310'>ab108310</a>) at 1/1000 dilution

Lane 1:

Wild-type THP-1 cell lysate at 30 µg

Lane 2:

ANPEP knockout THP-1 cell lysate at 30 µg

Lane 2:

Western blot - Human ANPEP (CD13) knockout THP-1 cell line (ab273759)

Lane 3:

PANC-1 cell lysate at 30 µg

Lane 4:

HEK-293 cell lysate at 30 µg

Predicted band size: 109 kDa

Observed band size: 160 kDa

false

Lab

Western blot - Human ANPEP (CD13) knockout THP-1 cell line (AB273759)

Lanes 1 - 4 : Merged signal (red and green). Green - ab108382 observed at 160 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.

ab108382 was shown to react with CD13 in wild-type THP-1 cells in western blot with loss of signal observed in ANPEP knockout cell line ab273759 (knockout cell lysate ab275505). Wild-type and ANPEP knockout THP-1 cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween^®) before incubation with ab108382 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 hour at room temperature before imaging.

All lanes:

Western blot - Anti-CD13 antibody [EPR4059] (<a href='/products/primary-antibodies/cd13-antibody-epr4059-ab108382'>ab108382</a>) at 1/1000 dilution

Lane 1:

Wild-type THP-1 cell lysate at 30 µg

Lane 2:

ANPEP knockout THP-1 cell lysate at 30 µg

Lane 2:

Western blot - Human ANPEP (CD13) knockout THP-1 cell line (ab273759)

Lane 3:

PANC-1 cell lysate at 30 µg

Lane 4:

HEK-293 cell lysate at 30 µg

Predicted band size: 109 kDa

Observed band size: 160 kDa

false

Lab

Western blot - Human ANPEP (CD13) knockout THP-1 cell line (AB273759)

Lanes 1 - 4 : Merged signal (red and green). Green - ab227663 observed at 160 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.

ab227663 was shown to react with CD13 in wild-type THP-1 cells in western blot with loss of signal observed in ANPEP knockout cell line ab273759 (knockout cell lysate ab275505). Wild-type and ANPEP knockout THP-1 cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween^®) before incubation with ab227663 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 400 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 hour at room temperature before imaging.

All lanes:

Western blot - Anti-CD13 antibody [SP187] (<a href='/products/primary-antibodies/cd13-antibody-sp187-ab227663'>ab227663</a>) at 1/400 dilution

Lane 1:

Wild-type THP-1 cell lysate at 30 µg

Lane 2:

Western blot - Human ANPEP (CD13) knockout THP-1 cell lysate (<a href='/products/cell-lysates/human-anpep-cd13-knockout-thp-1-cell-lysate-ab275505'>ab275505</a>) at 30 µg

Lane 3:

PANC-1 cell lysate at 30 µg

Lane 4:

HEK-293 cell lysate at 30 µg

Secondary

Lanes 1 - 4:

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

Lanes 1 - 4:

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

Predicted band size: 109 kDa

Observed band size: 160 kDa,37 kDa

false

Sanger Sequencing - Human ANPEP (CD13) knockout THP-1 cell line (AB273759)

Sanger seq

Supplier Data

Sanger Sequencing - Human ANPEP (CD13) knockout THP-1 cell line (AB273759)

Homozygous : 2 bp deletion in exon 2

Key facts

細胞タイプ

THP-1

生物種

Human

組織

Blood

製品の状態

Liquid

form

ノックアウト検証方法

Immunocytochemistry,Sanger Sequencing,Western blot

ノックアウト変異

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

疾病

Acute Monocytic Leukemia

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Reactivity data

{ "title": "Reactivity Data", "filters": { "stats": ["", "Reactivity", "Dilution Info", "Notes"] }, "values": { "ICC": { "reactivity":"TESTED_AND_REACTS", "dilution-info":"", "notes":"" }, "WB": { "reactivity":"TESTED_AND_REACTS", "dilution-info":"", "notes":"" }, "Flow Cyt": { "reactivity":"TESTED_AND_REACTS", "dilution-info":"", "notes":"" } } }

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製品の詳細

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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製品内容

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出荷温度及び保存条件

遺伝子名

ANPEP

遺伝子編集のタイプ

Knockout

遺伝子編集の方法

CRISPR technology

ノックアウト検証方法

Immunocytochemistry, Sanger Sequencing, Western blot

接合型

Homozygous

出荷温度

Dry Ice

短期保存温度

-196°C

長期保存温度

-196°C

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取り扱い方法

初回取り扱いガイドライン

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. Based on cell count, seed cells in an appropriate cell culture flask at a density of 4x10⁵ cells/mL. 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.
5. THP-1 cells recover slowly from cryopreservation and therefore may not be ready for subculture for a number of days. Cells should be left as much as possible over this time and only subcultured when the cell density reaches 8x10⁵ cells/mL.Small amounts of fresh media can be added until cell number/viability improves.

継代培養ガイドライン

All seeding densities should be based on cell counts gained by established methods.
Cells should be seeded at 2x10⁵ - 3x10⁵ cells/mL and subcultured when they have reached 8x10⁵ cells/mL.
It is not recommended to allow the cell density to exceed 1x10⁶ cells/mL.

培養培地

RPMI + 10% FBS + 0.05 mM beta-mercaptoethanol

凍結保存培地

Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.

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補足情報

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

CD13 also known as aminopeptidase N (ANPEP) is a transmembrane protein with a molecular weight of approximately 150 kDa. It functions as a zinc-dependent metalloenzyme which cleaves N-terminal amino acids from peptides and proteins. CD13 protein is expressed on various cell types including myeloid cells epithelial cells endothelial cells and fibroblasts. Its presence is significantly observed in the brush border of the small intestinal mucosa and the renal proximal tubule. Researchers can study CD13 using anti-CD13 antibodies and CD13 ELISA kits.

Biological function summary

CD13 regulates peptide-mediated signaling and controls the maturation and catabolism of bioactive peptides. The enzymatic function of CD13 influences processes such as cell proliferation motility and angiogenesis. It does not operate as part of a larger complex but its activity modulates several cellular and systemic functions. CD13's role in these biological processes highlights its importance in modulating local and systemic peptide pools which contributes to its diverse physiological effects.

Pathways

CD13 plays significant roles in the renin-angiotensin system and the regulation of inflammatory responses. In the renin-angiotensin system CD13 modulates the activity of angiotensin influencing blood pressure and fluid balance. Through its enzymatic activity CD13 interacts with proteins such as ACE another critical player in this pathway. In inflammation CD13 regulates the availability of chemotactic peptides affecting leukocyte migration and adhesion. The proteins it works with in inflammatory pathways include cytokines which CD13 indirectly modulates by altering chemokine activity.

Aberrant CD13 expression and activity are implicated in cancer and inflammatory diseases. In cancer overexpression of CD13 correlates with tumor growth and metastasis particularly in cancers like renal cell carcinoma and prostate cancer. It can confer an increased malignant phenotype by promoting angiogenesis and immune evasion. In inflammatory disorders dysregulation of CD13 exacerbates diseases such as rheumatoid arthritis through its impact on peptide-mediated signaling and immune cell movement. Its connection with inflammatory cytokines like IL-8 indicates its important role in mediating inflammatory responses.

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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

Suspension

Gender

Male

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製品プロトコール

Visit the General protocols
Visit the Troubleshooting

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ターゲットの情報

See full target information ANPEP

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文献 (1)

Recent publications for all applications. Explore the full list and refine your search

iScience 26:108219 PubMed37942010

2023

Cancer-specific glycosylation of CD13 impacts its detection and activity in preclinical cancer tissues.

Applications

Unspecified application

Species

Unspecified reactive species

Francis M Barnieh,Sebastian P Galuska,Paul M Loadman,Simon Ward,Robert A Falconer,Sherif F El-Khamisy

View all publications

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Abcam product promise

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保証に関する詳細については利用規約をご確認ください。

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

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Human ANPEP (CD13) knockout THP-1 cell line