Anti-TIM 1 antibody [EPR28341-72]

• Flow Cyt

Lab

Flow Cytometry - Anti-TIM 1 antibody [EPR28341-72] (AB316854)

Flow cytometric analysis of MEF (mouse embryo fibroblast, Left) / A20 (mouse reticulum sarcoma B lymphocyte, Right) cells labelling TIM 1 with ab316854 at 1/500 dilution (1µg) (Red) followed by a Goat anti-Rabbit IgG (Alexa Fluor^® 647, ab150083) secondary antibody used at a 1/5000 dilution compared with a Rabbit monoclonal IgG (ab172730) ( Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

Low expression : MEF.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TIM 1 antibody [EPR28341-72] (AB316854)

Immunohistochemical analysis of paraffin-embedded Mouse spleen tissue labeling TIM 1 with ab316854 at 1/2000 (0.252 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : No staining on mouse spleen.
The section was incubated with ab316854 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

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TIM 1 antibody [EPR28341-72] (AB316854)

Immunohistochemical analysis of paraffin-embedded Mouse normal kidney tissue labeling TIM 1 with ab316854 at 1/2000 (0.252 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : No staining on mouse normal kidney (PMID : 17277130).
The section was incubated with ab316854 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-TIM 1 antibody [EPR28341-72] (AB316854)

Immunohistochemical analysis of paraffin-embedded (A) Injured mouse kidney (cisplatin-induced acute kidney injury). (B) Control mouse kidney. tissue labeling TIM 1 with ab316854 at 1/2000 (0.252 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on (A) Injured mouse kidney (cisplatin-induced acute kidney injury), no staining on (B)control kidney (PMID : 34298880).
The section was incubated with ab316854 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

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Immunocytochemistry/ Immunofluorescence - Anti-TIM 1 antibody [EPR28341-72] (AB316854)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized A20 (mouse reticulum sarcoma B lymphocyte) cells labelling TIM 1 with ab316854 at 1/100 (5.03 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 membranous staining in A20 cell line (shown in green). The counterstain was observed in red. Nuclear DNA was labeled with DAPI (shown in blue).
Low expression : MEF.
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 (2.5 ug/ml) dilution (Red). 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.

• IP

Supplier Data

Immunoprecipitation - Anti-TIM 1 antibody [EPR28341-72] (AB316854)

TIM 1 was immunoprecipitated from 0.35 mg A20 (mouse reticulum sarcoma B lymphocyte) whole cell lysate with ab316854 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab316854 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : A20 (mouse reticulum sarcoma B lymphocyte) whole cell lysate
Lane 2 : ab316854 IP in A20 (mouse reticulum sarcoma B lymphocyte) whole cell lysate
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab316854 in A20 whole cell lysate

All lanes:

Immunoprecipitation - Anti-TIM 1 antibody [EPR28341-72] (ab316854) at 1/30 dilution

All lanes:

A20 (mouse reticulum sarcoma B lymphocyte) whole cell lysate at 10 µg

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: 93s

• WB

Supplier Data

Western blot - Anti-TIM 1 antibody [EPR28341-72] (AB316854)

Negative control : normal kidney (PMID : 21576461).

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

All lanes:

Western blot - Anti-TIM 1 antibody [EPR28341-72] (ab316854) at 1/1000 dilution

Lane 1:

Mouse injured kidney tissue lysate (cisplatin-induced acute kidney injury) at 20 µg

Lane 2:

Mouse kidney 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: 75 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-TIM 1 antibody [EPR28341-72] (AB316854)

Low expression : MEF.

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

All lanes:

Western blot - Anti-TIM 1 antibody [EPR28341-72] (ab316854) at 1/1000 dilution

Lane 1:

A20 (mouse reticum sarcoma B lymphocyte) whole cell lysate at 20 µg

Lane 2:

MEF (mouse embryonic fibroblast (immortalized)) 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

Observed band size: 75 kDa,36 kDa

true

Exposure time: 70s

• WB

Supplier Data

Western blot - Anti-TIM 1 antibody [EPR28341-72] (AB316854)

This antibody does not cross-react with mouse Timd2.

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

In Western blot, Anti-6X His tag® antibody [EPR20547] - ChIP Grade (ab213204) staining at 1/5000 dilution.

All lanes:

Western blot - Anti-TIM 1 antibody [EPR28341-72] (ab316854) at 1/1000 dilution

Lane 1:

293T (human embryonic kidney epithelial cell) transfected with an empty vector containing a myc-His-tag®, whole cell lysate at 20 µg

Lane 2:

293T transfected with a mouse TIM 1 expression vector containing a myc-His-tag®, whole cell lysate at 20 µg

Lane 3:

293T transfected with a mouse Timd2 expression vector containing a myc-His-tag®, 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

Observed band size: 20-250 kDa,36 kDa

false

Exposure time: 180s