Anti-LHX3/LIM antibody [EPR29190-234]

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LHX3/LIM antibody [EPR29190-234] (AB325931)

Immunohistochemical analysis of paraffin-embedded Human pituitary gland tissue labeling LHX3/LIM with ab325931 at 1/100 (5.22 µg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human pituitary gland.

The primary antibody was incubated 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

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LHX3/LIM antibody [EPR29190-234] (AB325931)

Immunohistochemical analysis of paraffin-embedded (A) Y79 (human retinoblastoma cell) cell pellet (B) 293T (human embryonic kidney epithelial cell) cell pellet tissue labeling LHX3/LIM with ab325931 at 1/100 (5.22 µg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on (A) Y79 cell pellet, no staining on (B) 293T cell pellet.

The primary antibody was incubated 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

Lab

Immunocytochemistry/ Immunofluorescence - Anti-LHX3/LIM antibody [EPR29190-234] (AB325931)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized Y79 (human retinoblastoma cell) 293T (human embryonic kidney epithelial cell) cells labelling LHX3/LIM with ab325931 at 1/200 (2.61 µg/ml) dilution , followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green).

Confocal image showing nuclear staining in Y79 cell line (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). Low expression : 293T.

ab7291 Anti-alpha Tubulin mouse monoclonal antibody was used to counterstain tubulin at 1/1000 dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

-ve control 1 : ab325931 at 1/200 dilution, followed by ab150120 at 1/1000 dilution.-ve control 2 : ab7291 at 1/1000 dilution, followed by ab150081 at 1/1000 dilution.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LHX3/LIM antibody [EPR29190-234] (AB325931)

Immunohistochemical analysis of paraffin-embedded Mouse pituitary gland tissue labeling LHX3/LIM with ab325931 at 1/100 (5.22 µg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on mouse pituitary gland (PMID : 9388186).

The primary antibody was incubated 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

• WB

Lab

Western blot - Anti-LHX3/LIM antibody [EPR29190-234] (AB325931)

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

Low expression : 293T

Samples are non-boiled as boiling may cause protein aggregation.

To minimize protein degradation, cells (lanes 4-6) were lysed immediately after harvest and then applied to a gel and transfer membrane for Western blotting as soon as possible.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) (1 : 200000) (36KDa).

All lanes:

Western blot - Anti-LHX3/LIM antibody [EPR29190-234] (ab325931) at 1/1000 dilution

Lane 1:

Y79 (human retinoblastoma cell) whole cell lysate at 20 µg

Lane 2:

WERI-Rb-1 (human Eye, Retina grape-like clusters of round cells) whole cell lysate at 20 µg

Lane 3:

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

Lane 4:

Y79 (human retinoblastoma cell) fresh whole cell lysate at 20 µg

Lane 5:

WERI-Rb-1 (human Eye, Retina grape-like clusters of round cells) fresh whole cell lysate at 20 µg

Lane 6:

293T (human embryonic kidney epithelial cell) fresh 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

Observed band size: 50 kDa,36 kDa

false

Exposure time: 180s

• WB

Lab

Western blot - Anti-LHX3/LIM antibody [EPR29190-234] (AB325931)

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

Low expression : spleen, liver

Samples are non-boiled as boiling may cause protein aggregation.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) (1 : 200000) (36KDa).

All lanes:

Western blot - Anti-LHX3/LIM antibody [EPR29190-234] (ab325931) at 1/1000 dilution

Lane 1:

Mouse pituitary tissue lysate at 20 µg

Lane 2:

Mouse spleen tissue lysate at 20 µg

Lane 3:

Mouse liver 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 1/20000 dilution

Observed band size: 50 kDa,36 kDa

false

Exposure time: 180s

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LHX3/LIM antibody [EPR29190-234] (AB325931)

Immunohistochemical analysis of paraffin-embedded Human spleen tissue labeling LHX3/LIM with ab325931 at 1/100 (5.22 µg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Low expression tissue : No staining on human spleen.

The primary antibody was incubated 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

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LHX3/LIM antibody [EPR29190-234] (AB325931)

Immunohistochemical analysis of paraffin-embedded Human liver tissue labeling LHX3/LIM with ab325931 at 1/100 (5.22 µg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Low expression tissue : No staining on human liver.

The primary antibody was incubated 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

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LHX3/LIM antibody [EPR29190-234] (AB325931)

Immunohistochemical analysis of paraffin-embedded Mouse liver tissue labeling LHX3/LIM with ab325931 at 1/100 (5.22 µg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Low expression tissue : No staining on mouse liver.

The primary antibody was incubated 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

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LHX3/LIM antibody [EPR29190-234] (AB325931)

Immunohistochemical analysis of paraffin-embedded Mouse spleen tissue labeling LHX3/LIM with ab325931 at 1/100 (5.22 µg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Low expression tissue : No staining on mouse spleen.

The primary antibody was incubated 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

• I-ELISA

Lab

Indirect ELISA - Anti-LHX3/LIM antibody [EPR29190-234] (AB325931)

Indirect ELISA analysis of ab325931 at 1000-0 ng/ml. The Secondary antibody used was Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG (H+L) at 1 : 2500 dilution dilution.

Antigen : Human LHX3/LIM protein;Human LHX4 protein;Human LHX5 protein.

Antigen concentration : 1000 ng/ml