MA5-31979

antibody from Invitrogen Antibodies

Targeting: KRT20 CK20, K20, MGC35423

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Immunohistochemistry (Data presented on provider website)

Flow cytometry (Supportive data in Antibodypedia)

Antibody Data

Product number

MA5-31979

Provider

Invitrogen Antibodies

Product name

Cytokeratin 20 Recombinant Rabbit Monoclonal Antibody (SA35-03)

Antibody type

Monoclonal

Antigen

Synthetic peptide

Description

Recombinant rabbit monoclonal antibodies are produced using in vitro expression systems. The expression systems are developed by cloning in the specific antibody DNA sequences from immunoreactive rabbits. Then, individual clones are screened to select the best candidates for production. The advantages of using recombinant rabbit monoclonal antibodies include: better specificity and sensitivity, lot-to-lot consistency, animal origin-free formulations, and broader immunoreactivity to diverse targets due to larger rabbit immune repertoire.

Reactivity

Human, Rat

Host

Rabbit

Isotype

IgG

Antibody clone number

SA35-03

Vial size

100 µL

Concentration

1 mg/mL

Storage

Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

References

Submitted references

The Interaction between Reactive Peritoneal Mesothelial Cells and Tumor Cells via Extracellular Vesicles Facilitates Colorectal Cancer Dissemination.

Serratì S, Porcelli L, Fragassi F, Garofoli M, Di Fonte R, Fucci L, Iacobazzi RM, Palazzo A, Margheri F, Cristiani G, Albano A, De Luca R, Altomare DF, Simone M, Azzariti A
Cancers 2021 May 20;13(10)

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis of Cytokeratin 20 in different lysates using a Monoclonal antibody (Product #MA5-31979) at a dilution of 1:1,000. Positive control: Lane 1: Lovo, Lane 2: HCT116, Lane 3: CRC.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis of Cytokeratin 20 in different lysates using a Monoclonal antibody (Product #MA5-31979) at a dilution of 1:1,000. Positive control: Lane 1: Lovo, Lane 2: HCT116, Lane 3: CRC.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Knockdown of KRT20 was achieved by transfecting HT-29 cells with KRT20 specific siRNAs (Silencer® select Product # s29040, Product # s29041). Western blot analysis (Fig. a) was performed using whole cell extracts from the KRT20 knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed using KRT20 Monoclonal Antibody (Product # MA5-31979, 1:1000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/ml, 1:4000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to KRT20.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot was performed using Anti-Cytokeratin 20 Monoclonal Antibody (Product # MA5-31979) on whole cell extracts (30 µg lysate) of HT-29 (Lane 1), HCT 116 (Lane 2) and MCF7 (Lane 3) and 48.48 kDa band corresponding to Cytokeratin 20 was observed. Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:1000 dilution) and detected by Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/ml, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunocytochemical analysis of Cytokeratin 20 in CRC cells using a Cytokeratin 20 Monoclonal antibody (Product # MA5-31979) as seen in green. The nuclear counter stain is DAPI (blue). Cells were fixed in paraformaldehyde, permeabilised with 0.25% Triton X100/PBS.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunofluorescence analysis of KRT20 was performed using HT-29 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 1 hour at room temperature. The cells were labeled with Cytokeratin 20 Recombinant Rabbit Monoclonal Antibody (Product # MA5-31979) at 5 µg/mL in 0.1% BSA and incubated overnight at 4 degree and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the composite image showing peripheral membrane localization of NUMB. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Flow cytometry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Flow Cytometric analysis of Cytokeratin 20 in CRC cells using a Cytokeratin 20 Monoclonal Antibody (Product # MA5-31979) at a dilution of 1:50, as seen in blue compared with an unlabelled control (cells without incubation with primary antibody; red). Alexa Fluor 488-conjugated goat anti rabbit IgG was used as the secondary antibody.