重组Anti-TLS/FUS抗体[EPR5812]

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-TLS/FUS antibody [EPR5812] (AB124923)

Flow Cytometry analysis of HepG2 (Human hepatocellular carcinoma epithelial cell) cells labelling TLS/FUS with Purified ab124923 at 1 : 100 dilution (10 µg/ml) (Red). Cells were fixed with 5% Paraformaldehyde and permeabilised with 91% Methanol. A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) secondary antibody was used at 1 : 2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabelled control - Cell without incubation with primary antibody and secondary antibody (Blue).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TLS/FUS antibody [EPR5812] (AB124923)

ab124923, at 1/50 dilution, staining TLS/FUS in formalin-fixed, paraffin-embedded Human kidney tissue, by Immunohistochemistry.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-TLS/FUS antibody [EPR5812] (AB124923)

Immunocytochemistry analysis of HepG2 (Human hepatocellular carcinoma epithelial cell) cells labeling TLS/FUS with Purified ab124923 at 1 : 100 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) 1 : 200 (2.5 µg/ml). 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.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TLS/FUS antibody [EPR5812] (AB124923)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast tissue sections labeling TLS/FUS with Purified ab124923 at 1 : 800 dilution (1.429 µg/mL). Heat mediated antigen retrieval was performed using Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used. PBS instead of the primary antibody was used as the negative control.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-TLS/FUS antibody [EPR5812] (AB124923)

Immunocytochemistry/ Immunofluorescence analysis of K-562 (human chronic myelogenous leukemia lymphoblast) cells labeling TLS/FUS with ab124923 at 1/250 (8.3 μg/mL). Cells were fixed with 4% Paraformaldehyde and permeabilised 0.1% TritonX-100. ab150077, AlexaFluor^®488 Goat anti-Rabbit secondary at 1/1000 (2 μg/mL) was used as the secondary antibody. Nuclei were counterstained with DAPI and are shown in blue.

Confocal image showing nuclear staining in K-562 cells.

• WB

Unknown

Western blot - Anti-TLS/FUS antibody [EPR5812] (AB124923)

We are unsure about the nature of the extra bands.

All lanes:

Western blot - Anti-TLS/FUS antibody [EPR5812] (ab124923) at 1/5000 dilution

Lane 1:

K-562 (Human chronic myelogenous leukemia lymphoblast) whole cell lysate at 20 µg

Lane 2:

Caco-2 (Human colorectal adenocarcinoma epithelial cell) whole cell lysate at 15 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Predicted band size: 53 kDa

Observed band size: 73 kDa

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• WB

Lab

Western blot - Anti-TLS/FUS antibody [EPR5812] (AB124923)

Lanes 1- 2 : Merged signal (red and green). Green - ab124923 observed at 75 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

ab124923 was shown to react with TLS/FUS in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266587 (knockout cell lysate ab257100) was used. Wild-type HEK-293T and FUS knockout HEK-293T cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab124923 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 1000 dilution and a 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-TLS/FUS antibody [EPR5812] (ab124923) at 1/1000 dilution

Lane 1:

Wild-type HEK-293T cell lysate at 20 µg

Lane 2:

FUS knockout HEK-293T cell lysate at 20 µg

Lane 2:

Western blot - Human FUS (TLS) knockout HEK-293T cell line (<a href='/products/cell-lines/human-fus-tls-knockout-hek-293t-cell-line-ab266587'>ab266587</a>)

Predicted band size: 53 kDa

Observed band size: 75 kDa

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• WB

Collaborator

Western blot - Anti-TLS/FUS antibody [EPR5812] (AB124923)

ab124923 was shown to react with FUS in wild-type HeLa cells in Western blot with loss of signal observed in a FUS knockout cell line. Wild-type HeLa and FUS knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab124923 overnight at 4 °C at a 1/5000 dilution. Blots were incubated with secondary antibodies at 1/5000 before imaging. These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

All lanes:

Western blot - Anti-TLS/FUS antibody [EPR5812] (ab124923)

Predicted band size: 53 kDa

false

• WB

Lab

Western blot - Anti-TLS/FUS antibody [EPR5812] (AB124923)

Lanes 1 to 4 : Merged signal (red and green). Green - ab124923 observed at 75 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab124923 was shown to specifically react with TLS/FUS when TLS/FUS knockout samples were used. Wild-type and TLS/FUS knockout samples were subjected to SDS-PAGE. ab124923 and ab28245 (loading control to GAPDH) were both diluted at 1/1000 and 1/10 000 respectively 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 1/10 000 dilution for 1 h at room temperature before imaging.

All lanes:

Western blot - Anti-TLS/FUS antibody [EPR5812] (ab124923) at 1/1000 dilution

Lane 1:

Wild-type HAP1 cell lysate at 20 µg

Lane 2:

TLS/FUS knockout HAP1 cell lysate at 20 µg

Lane 3:

K562 cell lysate at 20 µg

Lane 4:

HepG2 cell lysate at 20 µg

Predicted band size: 53 kDa

false

• WB

Unknown

Western blot - Anti-TLS/FUS antibody [EPR5812] (AB124923)

All lanes:

Western blot - Anti-TLS/FUS antibody [EPR5812] (ab124923) at 1/1000 dilution

Lane 1:

K562 cell lysate at 10 µg

Lane 2:

Human fetal brain lysate at 10 µg

Lane 3:

Caco 2 cell lysate at 10 µg

Lane 4:

HepG2 cell lysate at 10 µg

Secondary

All lanes:

Goat anti-rabbit HRP at 1/2000 dilution

Predicted band size: 53 kDa

Observed band size: 73 kDa

false

• OI-RD Scanning

Unknown

OI-RD Scanning - Anti-TLS/FUS antibody [EPR5812] (AB124923)

We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody. Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.