Anti-TRIM56 抗体 [EPR10583]

• WB

Lab

Western blot - Anti-TRIM56 antibody [EPR10583] (AB154862)

All lanes:

Western blot - Anti-TRIM56 antibody [EPR10583] (ab154862) at 1/50000 dilution

Lane 1:

HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates at 15 µg

Lane 2:

A375 (Human malignant melanoma epithelial cell) whole cell lysates at 15 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 81 kDa

Observed band size: 81 kDa

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• WB

Supplier Data

Western blot - Anti-TRIM56 antibody [EPR10583] (AB154862)

Lanes 1-4 : Merged signal (red and green). Green - ab154862 observed at 88 kDa. Red - loading control ab8245 observed at 36 kDa.

ab154862 Anti-TRIM56 antibody [EPR10583] was shown to specifically react with TRIM56 in wild-type A549 cells. Loss of signal was observed when knockout cell line ab267063 (knockout cell lysate ab258250) was used. Wild-type and TRIM56 knockout samples were subjected to SDS-PAGE. ab154862 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-TRIM56 antibody [EPR10583] (ab154862) at 1/1000 dilution

Lane 1:

Wild-type A549 (Human lung carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

TRIM56 knockout A549 (Human lung carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

Western blot - Human TRIM56 knockout A549 cell line (<a href='/products/cell-lines/human-trim56-knockout-a549-cell-line-ab267063'>ab267063</a>)

Lane 3:

MCF7 (Human breast adenocarcinoma cell line) whole cell lysate at 20 µg

Lane 4:

A-375 (Human malignant melanoma cell line) whole cell lysate at 20 µg

Secondary

All lanes:

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 1/10000 dilution

Predicted band size: 81 kDa

Observed band size: 88 kDa

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• WB

Unknown

Western blot - Anti-TRIM56 antibody [EPR10583] (AB154862)

All lanes:

Western blot - Anti-TRIM56 antibody [EPR10583] (ab154862) at 1/10000 dilution

Lane 1:

HeLa cell lysate at 10 µg

Lane 2:

A375 cell lysate at 10 µg

Lane 3:

MCF7 cell lysate at 10 µg

Predicted band size: 81 kDa

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• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-TRIM56 antibody [EPR10583] (AB154862)

Intracellular Flow Cytometry analysis of MCF7 (Human breast adenocarcinoma epithelial cell) cells labeling TRIM56 with purified ab154862 at 1/70 dilution (10μg/ml) (red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) secondary antibody was used at 1/2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TRIM56 antibody [EPR10583] (AB154862)

Immunohistochemical analysis of paraffin-embedded Human pancreas tissue labeling TRIM56 with ab154862 (unpurified) at 1/50.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TRIM56 antibody [EPR10583] (AB154862)

Immunohistochemical analysis of paraffin-embedded Human brain tissue labeling TRIM56 with ab154862 (unpurified) at 1/50.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TRIM56 antibody [EPR10583] (AB154862)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human colon cancer tissue sections labeling TRIM56 with purified ab154862 at 1/1000 dilution (0.691 µg/ml). Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-TRIM56 antibody [EPR10583] (AB154862)

Immunocytochemistry/ Immunofluorescence analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling TRIM56 with purified ab154862 at 1/70 dilution (10 μg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) at 1/200 (2.5 μg/ml) dilution. Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody at 1/1000 (2 μg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-TRIM56 antibody [EPR10583] (AB154862)

Immunofluorescent analysis of MCF7 cells labeling TRIM56 with ab154862 (unpurified) at 1/100.

• WB

Lab

Western blot - Anti-TRIM56 antibody [EPR10583] (AB154862)

Lane 1 : Wild-type HAP1 cell lysate (20 μg)
Lane 2 : TRIM56 knockout HAP1 cell lysate (20 μg)
Lane 3 : MCF7 cell lysate (20 μg)
Lane 4 : A375 cell lysate (20 μg)
Lanes 1 - 4 : Merged signal (red and green). Green - ab154862 observed at 88 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab154862 (unpurifed) was shown to recognize TRIM56 when TRIM56 knockout samples were used, along with additional cross-reactive bands. Wild-type and TRIM56 knockout samples were subjected to SDS-PAGE. ab154862 and ab8245 (loading control to GAPDH) were both diluted 1/10000 and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10000 dilution for 1 h at room temperature before imaging.

All lanes:

Western blot - Anti-TRIM56 antibody [EPR10583] (ab154862)

Predicted band size: 81 kDa

false

• WB

Lab

Western blot - Anti-TRIM56 antibody [EPR10583] (AB154862)

Lanes 1-4 : Merged signal (red and green). Green - ab154862 observed at 88 kDa. Red - loading control ab8245 observed at 36 kDa.

ab154862 Anti-TRIM56 antibody [EPR10583] was shown to specifically react with TRIM56 in wild-type A549 cells. Loss of signal was observed when knockout cell line ab267062 (knockout cell lysate ab258249) was used. Wild-type and TRIM56 knockout samples were subjected to SDS-PAGE. ab154862 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-TRIM56 antibody [EPR10583] (ab154862) at 1/1000 dilution

Lane 1:

Wild-type A549 (Human lung carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

TRIM56 knockout A549 (Human lung carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

Western blot - Human TRIM56 knockout A549 cell line (<a href='/products/cell-lines/human-trim56-knockout-a549-cell-line-ab267062'>ab267062</a>)

Lane 3:

MCF7 (Human breast adenocarcinoma cell line) whole cell lysate at 20 µg

Lane 4:

A-375 (Human malignant melanoma cell line) whole cell lysate at 20 µg

Secondary

All lanes:

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 1/10000 dilution

Predicted band size: 81 kDa

Observed band size: 88 kDa

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• WB

CiteAb

Western blot - Anti-TRIM56 antibody [EPR10583] (AB154862)

TRIM56 western blot using anti-TRIM56 antibody [EPR10583] ab154862. Publication image and figure legend from Fiškin, E., Bhogaraju, S., et al., 2017, Nat Commun, PubMed 28084320.

ab154862 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab154862 please see the product overview.

Identification of TRIM56 and TRIM65 as SopA-interacting proteins.(a) Workflow for SILAC-coupled SopA interactome analysis from inducible HeLa Flp-In T-REx GFP-SopA-expressing cells. (b) SopA interacts with TRIM56 and TRIM65. Scatter plot of forward and reverse SILAC SopA interactome. Proteins situated in the upper left quadrant include contaminants. (c) Endogenous TRIM56 and TRIM65 specifically interact with SopA. Lysates from HEK293T cells expressing GFP, GFP-SopA or GFP-NleL constructs were subjected to anti-GFP IP, followed by SDS–PAGE and immunoblot. (d) Bacterially translocated SopA interacts with TRIM56/65. Scatter plot of forward and reverse SILAC interactome experiments from Salmonella-infected HeLa cells. Proteins situated in the upper left quadrant include contaminants. (e) Endogenous TRIM56 interacts with bacterially secreted SopA during infection. Lysates from HeLa cells infected with SL1344 WT, SopA–HA or catalytic-dead SopA C753A-HA-expressing strains were subjected to anti-HA IP, followed by SDS–PAGE and immunoblot.

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