Anti-SPTLC1 antibody [EPR26226-221] - BSA and Azide free

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SPTLC1 antibody [EPR26226-221] - BSA and Azide free (AB315159)

This data was developed using ab315158, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Human skeletal muscle tissue labeling SPTLC1 with ab315158 at 1/2000 (0.249 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : No staining on human skeletal muscle. The section was incubated with ab315158 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-SPTLC1 antibody [EPR26226-221] - BSA and Azide free (AB315159)

This data was developed using ab315158, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Human placenta tissue labeling SPTLC1 with ab315158 at 1/2000 (0.249 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human placenta. The section was incubated with ab315158 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-SPTLC1 antibody [EPR26226-221] - BSA and Azide free (AB315159)

This data was developed using ab315158, the same antibody clone in a different buffer formulation.

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized 293T (human embryonic kidney epithelial cell) cells labelling SPTLC1 with ab315158 at 1/50 (9.94 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 293T cell line. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

Anti-KDEL mouse monoclonal antibody was used to counterstain tubulin at 1/100 (1 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.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-SPTLC1 antibody [EPR26226-221] - BSA and Azide free (AB315159)

This data was developed using ab315158, the same antibody clone in a different buffer formulation.

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HeLa (human cervical adenocarcinoma epithelial cell) cells labelling SPTLC1 with ab315158 at 1/50 (9.94 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 HeLa cell line. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

Anti-KDEL mouse monoclonal antibody was used to counterstain tubulin at 1/100 (1 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.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-SPTLC1 antibody [EPR26226-221] - BSA and Azide free (AB315159)

This data was developed using ab315158, the same antibody clone in a different buffer formulation.

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized 293T (human embryonic kidney epithelial cell) cells labelling SPTLC1 with ab315158 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.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-SPTLC1 antibody [EPR26226-221] - BSA and Azide free (AB315159)

This data was developed using ab315158, the same antibody clone in a different buffer formulation.

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HeLa (human cervical adenocarcinoma epithelial cell) cells labelling SPTLC1 with ab315158 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-SPTLC1 antibody [EPR26226-221] - BSA and Azide free (AB315159)

This data was developed using ab315158, the same antibody clone in a different buffer formulation.

SPTLC1 was immunoprecipitated from 0.35 mg HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate with ab315158 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab315158 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 : ab315158 IP in HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab315158 in HepG2 whole cell lysate

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

All lanes:

Immunoprecipitation - Anti-SPTLC1 antibody [EPR26226-221] (<a href='/en-us/products/primary-antibodies/sptlc1-antibody-epr26226-221-ab315158'>ab315158</a>) 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: 24s

• WB

Lab

Western blot - Anti-SPTLC1 antibody [EPR26226-221] - BSA and Azide free (AB315159)

This data was developed using ab315158, the same antibody clone in a different buffer formulation.

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

Compared with ab307552, ab315158 has higher sensitivity. We recommend ab315158 as an alternative.

Lanes 1 - 4:

Western blot - Anti-SPTLC1 antibody [EPR26226-136] (<a href='/en-us/products/primary-antibodies/sptlc1-antibody-epr26226-136-ab307552'>ab307552</a>) at 1/1000 dilution

Lanes 5 - 8:

Western blot - Anti-SPTLC1 antibody [EPR26226-221] (<a href='/en-us/products/primary-antibodies/sptlc1-antibody-epr26226-221-ab315158'>ab315158</a>) at 1/1000 dilution

Lanes 1 and 5:

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

Lanes 2 and 6:

293T (human embryonic kidney epithelial cell) whole cell lysate

Lanes 3 and 7:

HepG2 (human hepatocellar carcinoma epithelial cell) whole cell lysate at 20 µg

Lanes 4 and 8:

Human small intestine 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: 54 kDa

false

Exposure time: 60s

• WB

Supplier Data

Western blot - Anti-SPTLC1 antibody [EPR26226-221] - BSA and Azide free (AB315159)

This data was developed using ab315158, the same antibody clone in a different buffer formulation.

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

Low expression : human small intestine, skeletal muscle (PMID : 17023427).

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

All lanes:

Western blot - Anti-SPTLC1 antibody [EPR26226-221] (<a href='/en-us/products/primary-antibodies/sptlc1-antibody-epr26226-221-ab315158'>ab315158</a>) at 1/1000 dilution

Lane 1:

Human placenta tissue lysate at 20 µg

Lane 2:

Human skeletal muscle tissue lysate at 20 µg

Lane 3:

Human small intestine tissue lysate at 20 µg

Observed band size: 53 kDa,36 kDa

false

Exposure time: 48s

• WB

Supplier Data

Western blot - Anti-SPTLC1 antibody [EPR26226-221] - BSA and Azide free (AB315159)

This data was developed using ab315158, the same antibody clone in a different buffer formulation.

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

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

All lanes:

Western blot - Anti-SPTLC1 antibody [EPR26226-221] (<a href='/en-us/products/primary-antibodies/sptlc1-antibody-epr26226-221-ab315158'>ab315158</a>) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) transfected with scrambled siRNA control whole cell lysate at 20 µg

Lane 2:

HeLa transfected with siRNA specifically targeting SPTLC1 whole cell lysate at 20 µg

Lane 3:

Jurkat (human t cell leukemia t lymphocyte from peripheral blood) whole cell lysate at 20 µg

Lane 4:

HepG2 (human hepatocellular carcinoma epithelial cell) 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: 53 kDa,36 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-SPTLC1 antibody [EPR26226-221] - BSA and Azide free (AB315159)

This data was developed using ab315158, the same antibody clone in a different buffer formulation.

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

All lanes:

Western blot - Anti-SPTLC1 antibody [EPR26226-221] (<a href='/en-us/products/primary-antibodies/sptlc1-antibody-epr26226-221-ab315158'>ab315158</a>) at 1/1000 dilution

All lanes:

293T (human embryonic kidney epithelial cell) 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: 53 kDa

false

Exposure time: 15s

• WB

Supplier Data

Western blot - Anti-SPTLC1 antibody [EPR26226-221] - BSA and Azide free (AB315159)

This data was developed using ab315158, the same antibody clone in a different buffer formulation.

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

This antibody does not cross-react with human SPTLC2 and SPTLC3.

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

All lanes:

Western blot - Anti-SPTLC1 antibody [EPR26226-221] (<a href='/en-us/products/primary-antibodies/sptlc1-antibody-epr26226-221-ab315158'>ab315158</a>) at 1/1000 dilution

Lane 1:

His-tagged human SPTLC1 recombinant protein fragment at 10 ng

Lane 2:

His-tagged human SPTLC2 recombinant protein fragment at 10 ng

Lane 3:

His-tagged human SPTLC3 recombinant protein fragment at 10 ng

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: 46 kDa

false

Exposure time: 15s