Anti-Transferrin antibody [RM2060] - Serum Loading Control

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized Hepa1-6 (mouse hepatoma epithelial cell) cells labelling Transferrin with ab318997 at 1/100 (5.08 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/mL dilution (Green).

Confocal image showing cytoplasmic staining in Hepa1-6 cell line (shown in green). The counterstain was observed in magenta. Nuclear DNA was labeled with DAPI (shown in blue).
Low expression : C2C12
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2 ug/mL dilution.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Immunohistochemical analysis of paraffin-embedded Human non-perfused fixed thyroid carcinoma tissue labeling Transferrin with ab318997 at 1/100 (5.08 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining in plasma of human non-perfused fixed thyroid carcinoma.
The section was incubated with ab318997 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HepG2 (human hepatocellular carcinoma epithelial cell) cells labelling Transferrin with ab318997 at 1/100 (5.08 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/mL dilution (Green).

Confocal image showing cytoplasmic staining in HepG2 cell line (shown in green). The counterstain was observed in magenta. Nuclear DNA was labeled with DAPI (shown in blue).
Low expression : K-562
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2 ug/mL dilution.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Immunohistochemical analysis of paraffin-embedded Human non-perfused fixed colon tissue labeling Transferrin with ab318997 at 1/100 (5.08 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining in plasma of human non-perfused fixed colon.
The section was incubated with ab318997 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HepG2 (human hepatocellular carcinoma epithelial cell) (Right) / K-562 (human chronic myelogenous leukemia lymphoblast) (Left) cells labelling Transferrin with ab318997 at 1/500 dilution (0.1ug )/ Magenta compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue).

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

Low expression : K-562.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Immunohistochemical analysis of paraffin-embedded Human non-perfused fixed skeletal muscle tissue labeling Transferrin with ab318997 at 1/100 (5.08 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Low expression tissue : No staining in human non-perfused fixed skeletal muscle.
The section was incubated with ab318997 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IP

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Immunoprecipitation - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Transferrin was immunoprecipitated from 0.35 mg HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate with ab318997 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab318997 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate
Lane 2 : ab318997 IP in HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab318997 in HepG2 whole cell lysate

All lanes:

Immunoprecipitation - Anti-Transferrin antibody [RM2060] - Serum Loading Control (ab318997) at 1/30 dilution

All lanes:

HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

false

Exposure time: 180s

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Immunohistochemical analysis of paraffin-embedded (A) Mouse perfused fixed liver. (B) Mouse non-perfused fixed liver. tissue labeling Transferrin with ab318997 at 1/2000 (0.254 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining in both (A) perfused fixed and (B) non-perfused fixed mouse liver.
The section was incubated with ab318997 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Immunohistochemical analysis of paraffin-embedded (A) Mouse perfused fixed cardiac muscle. (B) Mouse non-perfused fixed cardiac muscle. tissue labeling Transferrin with ab318997 at 1/2000 (0.254 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Nearly no staining in (A) mouse perfused fixed cardiac muscle; positive staining in plasma of (B) mouse non-perfused fixed cardiac muscle.
The section was incubated with ab318997 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Immunohistochemical analysis of paraffin-embedded Mouse perfused fixed skeletal muscle tissue labeling Transferrin with ab318997 at 1/2000 (0.254 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Low expression tissue : Nearly no staining in mouse perfused fixed skeletal muscle.
The section was incubated with ab318997 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IP

Supplier Data

Immunoprecipitation - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Transferrin was immunoprecipitated from 0.35 mg Mouse liver tissue lysate with ab318997 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab318997 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : Mouse liver tissue lysate
Lane 2 : ab318997 IP in Mouse liver tissue lysate
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab318997 in mouse liver tissue lysate

All lanes:

Immunoprecipitation - Anti-Transferrin antibody [RM2060] - Serum Loading Control (ab318997) at 1/30 dilution

All lanes:

Mouse liver tissue lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

false

Exposure time: 8s

• WB

Supplier Data

Western blot - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Low expression : skeletal muscle (PMID : 3558394).

The identity of the lower MW band at approximately 73 kDa (in lane 5) is unknown.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-Transferrin antibody [RM2060] - Serum Loading Control (ab318997) at 1/1000 dilution

Lane 1:

Human colon tissue lysate at 20 µg

Lane 2:

Human breast tissue lysate at 20 µg

Lane 3:

Human liver tissue lysate at 20 µg

Lane 4:

Human testis tissue lysate at 20 µg

Lane 5:

Human skeletal muscle tissue lysate at 20 µg

Secondary

All lanes:

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

Observed band size: 76 kDa,36 kDa

false

Exposure time: 15s

• WB

Supplier Data

Western blot - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Exposure time : Lane 1 : 114 seconds, lanes 2-3 : 15 seconds

All lanes:

Western blot - Anti-Transferrin antibody [RM2060] - Serum Loading Control (ab318997) at 1/1000 dilution

Lane 1:

Human plasma lysate at 20 µg

Lane 2:

Mouse plasma lysate at 20 µg

Lane 3:

Mouse serum 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

Observed band size: 76 kDa

false

• WB

Supplier Data

Western blot - Anti-Transferrin antibody [RM2060] - Serum Loading Control (AB318997)

Low expression : skeletal muscle (PMID : 3558394).

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-Transferrin antibody [RM2060] - Serum Loading Control (ab318997) at 1/1000 dilution

Lane 1:

Mouse liver tissue lysate at 20 µg

Lane 2:

Mouse heart tissue lysate at 20 µg

Lane 3:

Mouse skeletal muscle tissue 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

Observed band size: 76 kDa,36 kDa

false

Exposure time: 114s