Anti-P5CS antibody [EPR28827-56]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-P5CS antibody [EPR28827-56] (AB315816)

Immunohistochemical analysis of paraffin-embedded Human kidney tissue labeling P5CS with ab315816 at 1/100 (4.98 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human kidney. The section was incubated with ab315816 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-P5CS antibody [EPR28827-56] (AB315816)

Immunohistochemical analysis of paraffin-embedded Human stomach tissue labeling P5CS with ab315816 at 1/100 (4.98 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human stomach. The section was incubated with ab315816 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-P5CS antibody [EPR28827-56] (AB315816)

Immunohistochemical analysis of paraffin-embedded Human testis tissue labeling P5CS with ab315816 at 1/100 (4.98 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human testis. The section was incubated with ab315816 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-P5CS antibody [EPR28827-56] (AB315816)

Immunohistochemical analysis of paraffin-embedded (A) Wild-type HEK293T (human epithelial cell line from embryonic kidney) cell pellet (B) ALDH18A1 knockout HEK293T cell pellet tissue labeling P5CS with ab315816 at 1/100 (4.98 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on (A) Wild-type HEK293T cell pellet, negative staining on (B) ALDH18A1 knockout HEK293T cell pellet. The section was incubated with ab315816 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)

Supplier Data

Flow Cytometry (Intracellular) - Anti-P5CS antibody [EPR28827-56] (AB315816)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized ALDH18A1 KO HEK293T (human ALDH18A1 knockout embryonic kidney epithelial cell, Left) / Parental HEK293T (Right) cells labelling P5CS with ab315816 at 1/500 dilution (0.1 ug)/Red 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.

• IP

Supplier Data

Immunoprecipitation - Anti-P5CS antibody [EPR28827-56] (AB315816)

P5CS was immunoprecipitated from 0.35 mg HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate with ab315816 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab315816 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

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

All lanes:

Immunoprecipitation - Anti-P5CS antibody [EPR28827-56] (ab315816) at 1/1000 dilution

Lane 1:

HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate

Lane 2:

ab315816 at 1/30 IP in HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate

Lane 3:

Rabbit monoclonal IgG (<a href='/ko/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab315816 in HepG2 whole cell lysate

Secondary

All lanes:

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

false

Exposure time: 3s

• IP

Supplier Data

Immunoprecipitation - Anti-P5CS antibody [EPR28827-56] (AB315816)

P5CS was immunoprecipitated from 0.35 mg NIH/3T3 (mouse embryonic fibroblast) whole cell lysate with ab315816 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab315816 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

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

All lanes:

Immunoprecipitation - Anti-P5CS antibody [EPR28827-56] (ab315816) at 1/1000 dilution

Lane 1:

NIH/3T3 (mouse embryonic fibroblast) whole cell lysate

Lane 2:

ab315816 at 1/30 IP in NIH/3T3 (mouse embryonic fibroblast) whole cell lysate

Lane 3:

Rabbit monoclonal IgG (<a href='/ko/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab315816 in NIH/3T3 whole cell lysate

Secondary

All lanes:

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

false

Exposure time: 6s

• WB

Supplier Data

Western blot - Anti-P5CS antibody [EPR28827-56] (AB315816)

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

In Western blot, ab315816 was shown to bind specifically to P5CS.

Target of interest was observed at 90 KD in wild-type HEK293T cell lysates (lane 3, wild-type cell line ab255449) with no signal observed at this size in ALDH18A1(P5CS) knockout cell line (lane 4, knockout cell line ab266378/ knockout cell lysate ab258300).

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

All lanes:

Western blot - Anti-P5CS antibody [EPR28827-56] (ab315816) at 1/1000 dilution

Lane 1:

HepG2 (human hepatocellar carcinoma epithelial cell) transfected with scrambled siRNA control whole cell lysate at 10 µg

Lane 2:

HepG2 transfected with siRNA specifically targeting P5CS whole cell lysate at 10 µg

Lane 3:

Wild-type HEK293T (human embryonic kidney epithelial cell) whole cell lysate at 10 µg

Lane 4:

ALDH18A1(P5CS) knockout HEK293T whole cell lysate at 10 µg

Secondary

All lanes:

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

Observed band size: 90 kDa,36 kDa

false

Exposure time: 26s

• WB

Supplier Data

Western blot - Anti-P5CS antibody [EPR28827-56] (AB315816)

Exposure time : Lane1-5 : 26 seconds, lane 6-7 : 180 seconds

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

All lanes:

Western blot - Anti-P5CS antibody [EPR28827-56] (ab315816) at 1/1000 dilution

Lane 1:

MCF7 (human breast adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

A431 (human epidermoid carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

A549 (human lung carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

NIH/3T3 (mouse embryonic fibroblast) whole cell lysate at 20 µg

Lane 5:

4T1 (mouse mammary gland carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 6:

Mouse brain tissue lysate at 20 µg

Lane 7:

Rat testis tissue lysate at 20 µg

Secondary

All lanes:

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

Observed band size: 90 kDa

false