Anti-Transferrin Receptor antibody [EPR20584]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Immunohistochemical analysis of paraffin-embedded human esophageal cancer tissue labeling Transferrin Receptor with ab214039 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining on human esophageal cancer. The staining intensity of human esophageal cancer tissue was stronger than the human paracarcinoma esophagus tissue. Both paracarcinoma and human esophageal cancer tissues have been taken from the same patient (PMID : 24435655). Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Immunohistochemical analysis of paraffin-embedded human placenta tissue labeling Transferrin Receptor with ab214039 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Positive staining on human placenta (PMID : 27483296). Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Immunohistochemical analysis of paraffin-embedded human cerebrum tissue labeling Transferrin Receptor with ab214039 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Positive staining on capillaries of human cerebrum (PMID : 6095085). Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Immunohistochemical analysis of paraffin-embedded human esophagus tissue labeling Transferrin Receptor with ab214039 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Weakly cytoplasmic staining on human paracarcinoma esophagus. The staining intensity of human paracarcinoma esophagus was weaker than the human esophageal cancer tissue. Both paracarcinoma and human esophageal cancer tissues have been taken from the same patient (PMID : 24435655). Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Immunofluorescent analysis of 100% methanol-fixed K562 (human chronic myelogenous leukemia cell line from bone marrow) cells labeling Transferrin Receptor with ab214039 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing positive staining on K562 cells.

The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) (red) at 1/200 dilution.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Immunohistochemical analysis of formalin fixed paraffin embedded human placenta labelling Transferrin Receptor with ab214039 at a concentration of 0.5µg/ml. The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with a OptiView DAB IHC Detection Kit. Heat mediated antigen retrieval was performed with DISCOVERY cell conditioning solution (CC1) 100°C, pH8.5 for 32mins.

Anti-Transferrin Receptor antibody [EPR20584] ab214039 was incubated for 16mins at 37°C. Sections were counterstained with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control.

Customers are encouraged to optimise antigen retrieval conditions, antibody concentration, incubation times and temperature for best results in their own IHC assay workflow (automated and manual).

• IP

Supplier Data

Immunoprecipitation - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Transferrin Receptor was immunoprecipitated from 0.35 mg of K562 (human chronic myelogenous leukemia cell line from bone marrow) lysate with ab214039 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab214039 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution

Lane 1 : K562 whole cell lysate 10 μg (Input).

Lane 2 : ab214039 IP in K562 whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab214039 in K562 whole cell lysate.

Blocking and dilution buffer and concentration : 5% NFDM/TBST.

Exposure time : 1 second.

All lanes:

Immunoprecipitation - Anti-Transferrin Receptor antibody [EPR20584] (ab214039)

Predicted band size: 84 kDa

false

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Immunofluorescent analysis of 100% methanol-fixed RAW 264.7 (mouse macrophage cell line transformed with Abelson murine leukemia virus) cells labeling Transferrin Receptor with ab214039 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing positive staining on RAW 264.7 cells.

The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) (red) at 1/200 dilution.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Immunohistochemical analysis of paraffin-embedded mouse kidney tissue labeling Transferrin Receptor with ab214039 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining on renal tubules of mouse kidney (PMID : 12538733). Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• WB

Lab

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Blocking and diluting beffer and concentration : 5% NFDM/TBST.

ab181602 was used as a GAPDH loading control at 1/200000 dilution.

The blot of ab269513 is applied with Peroxidase-Conjugated Goat anti-Mouse IgG (H+L) secondary antibody at 1/10000 dilution and the blot of ab214039 is applied with Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) secondary antibody at 1/100000 dilution.

Lanes 1 - 9:

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (ab214039) at 1/1000 dilution

Lanes 1 - 9:

Western blot - Anti-Transferrin Receptor antibody [H68.4] (<a href='/en-us/products/primary-antibodies/transferrin-receptor-antibody-h684-ab269513'>ab269513</a>) at 1/1000 dilution

Lane 1:

Human placenta tissue lysate at 20 µg

Lane 2:

Human small intestine tissue lysate at 20 µg

Lane 3:

HT-29 (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

LoVo (human colorectal adenocarcinoma epithelial cell)whole cell lysate at 20 µg

Lane 5:

Mouse placenta tissue lysate at 20 µg

Lane 6:

Mouse kidney tissue lysate at 20 µg

Lane 7:

Mouse heart tissue lysate at 20 µg

Lane 8:

Rat placenta tissue lysate at 20 µg

Lane 9:

Rat kidney tissue lysate at 20 µg

Predicted band size: 84 kDa

Observed band size: 90 kDa

false

Exposure time: 26s

• WB

Supplier Data

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1 : 2 seconds; Lane 2 : 1 minute; Lane 3 : 1 second.

Lanes 1 - 2:

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (ab214039) at 1/2000 dilution

Lane 3:

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (ab214039) at 1/1000 dilution

Lane 1:

K562 (human chronic myelogenous leukemia cell line from bone marrow) whole cell lysate at 10 µg

Lane 2:

Jurkat (human T cell leukemia cell line from peripheral blood) whole cell lysate at 10 µg

Lane 3:

TF-1 (human bone marrow erythroleukemia cell line) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 84 kDa

Observed band size: 90 kDa

false

• WB

Lab

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

False colour image of Western blot : Anti-Transferrin Receptor antibody [EPR20584] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab214039 was shown to bind specifically to Transferrin Receptor. 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-Transferrin Receptor antibody [EPR20584] (ab214039) at 1/1000 dilution

Lane 1:

HDLM-2 cell lysate at 20 µg

Lane 2:

SH-SY5Y cell lysate at 20 µg

Predicted band size: 84 kDa

Observed band size: 90 kDa

false

• WB

Supplier Data

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1 : 3 seconds; Lane 2 : 15 seconds.

All lanes:

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (ab214039) at 1/1000 dilution

Lane 1:

Human fetal liver lysate at 20 µg

Lane 2:

Human placenta lysate at 20 µg

Secondary

All lanes:

Western blot - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/4000 dilution

Predicted band size: 84 kDa

Observed band size: 90 kDa

false

• WB

Lab

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Blocking buffer and concentration : 5% NFDM/TBST  Diluting buffer and concentration : 5% NFDM/TBST Exposure Time : Lane 1-4 : 3 seconds, Lane 5-6 : 20 seconds. Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) used. Soluble transferrin receptor (sTfR) is cleaved from whole TfR at Arg 100. (PMID : 9575164, PMID : 23390091).

All lanes:

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (ab214039) at 1/1000 dilution

Lane 1:

HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

HUVEC (Human umbilical vein endothelial cell) whole cell lysate at 20 µg

Lane 3:

293T (Human embryonic kidney epithelial cell) whole cell lysate at 20 µg

Lane 4:

Raw 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate at 20 µg

Lane 5:

Mouse lung tissue lysate at 20 µg

Lane 6:

Mouse brain tissue lysate at 20 µg

Secondary

All lanes:

Western blot at 1/20000 dilution

Predicted band size: 85 kDa

Observed band size: 75 kDa,85 kDa

false

• WB

Lab

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Soluble transferrin receptor (sTfR) is cleaved from whole TfR at Arg 100. (PMID : 9575164, PMID : 23390091) TfR is found as a dimer linked by disulfide bonds in cell, so sometimes we could see dimer in a few kinds of lysates. (PMID : 2507316, PMID : 3582362). This dimer was also be detected by ab108985. Blocking buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (ab214039) at 1/1000 dilution

Lane 1:

Daudi (Human Burkitt's lymphoma lymphoblast) whole cell lysate at 20 µg

Lane 2:

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

Secondary

All lanes:

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

false

Exposure time: 3s

• WB

Supplier Data

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (AB214039)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1/4 : 3 seconds; Lane 2 : 30 seconds; Lane 3 : 1 second.

All lanes:

Western blot - Anti-Transferrin Receptor antibody [EPR20584] (ab214039) at 1/1000 dilution

Lane 1:

Mouse spleen lysate at 20 µg

Lane 2:

Mouse testis lysate at 20 µg

Lane 3:

Mouse placenta lysate at 20 µg

Lane 4:

RAW 264.7 (mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 84 kDa

Observed band size: 90 kDa

false