Anti-SPHK2 antibody [EPR29189-33] - BSA and Azide free

• WB

Supplier Data

Western blot - Anti-SPHK2 antibody [EPR29189-33] - BSA and Azide free (AB320742)

This data was developed using ab320741, the same antibody clone in a different buffer formulation. Blocking and diluting buffer and concentration : 5% NFDM/TBST. To minimize protein degradation, cells were lysed immediately after harvest and then applied to a gel and transfer membrane for Western blotting as soon as possible (Lane 4). In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control  (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-SPHK2 antibody [EPR29189-33] (<a href='/en-us/products/primary-antibodies/sphk2-antibody-epr29189-33-ab320741'>ab320741</a>) at 1/1000 dilution

Lane 1:

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

Lane 2:

THP-1 (human monocytic leukemia monocyte) whole cell frozen lysate at 20 µg

Lane 3:

NIH/T (mouse embryonic fibroblast) whole cell frozen lysate at 20 µg

Lane 4:

NIH/3T3 (mouse embryonic fibroblast) whole cell fresh 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: 70 kDa,36 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-SPHK2 antibody [EPR29189-33] - BSA and Azide free (AB320742)

This data was developed using ab320741, the same antibody clone in a different buffer formulation. Blocking and diluting buffer and concentration : 5% NFDM/TBST. In Western blot, ab320741 was shown to bind specifically to SPHK2. Target of interest was observed at 70 kDa in wild-type HEK293T cell lysate (Lane 1) with no signal observed at this size in SPHK2 knockout cell line (Lane 2) (Lane 2, knockout cell line ab267293/ knockout cell lysate ab258699). In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control  (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-SPHK2 antibody [EPR29189-33] (<a href='/en-us/products/primary-antibodies/sphk2-antibody-epr29189-33-ab320741'>ab320741</a>) at 1/1000 dilution

Lane 1:

Parental HEK293T(EDWT04) whole cell lysate at 20 µg

Lane 2:

Western blot - Human SPHK2 knockout HEK-293T cell lysate (<a href='/en-us/products/cell-lysates/human-sphk2-knockout-hek-293t-cell-lysate-ab258699'>ab258699</a>) 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: 70 kDa,36 kDa

false

Exposure time: 180s

• Dot

Supplier Data

Dot Blot - Anti-SPHK2 antibody [EPR29189-33] - BSA and Azide free (AB320742)

This data was developed using ab320741, the same antibody clone in a different buffer formulation. Dot blot analysis of SPHK2 using ab320741 at 1 : 1000 (0.475 ug/ml) followed by a Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1 : 100,000 dilution. Lane1 : 293T cells transfected with a human SPHK2 expression vector containing a myc-his-tag whole cell lysate Lane2 : 293T cells transfected with a human SPHK1 expression vector containing a myc-his-tag whole cell lysate Exposure time : 180 seconds. Blocking and diluting buffer and concentration : 5% NFDM/TBST. This antibody does not cross-react with human SPHK1. Anti-His tag® antibody (1 : 10000) as total protein control.

All lanes:

Dot Blot - Anti-SPHK2 antibody [EPR29189-33] (<a href='/en-us/products/primary-antibodies/sphk2-antibody-epr29189-33-ab320741'>ab320741</a>) at 1/1000 dilution

Lane 1:

293T cells transfected with a human SPHK2 expression vector containing a myc-his-tag whole cell lysate

Lane 2:

293T cells transfected with a human SPHK1 expression vector containing a myc-his-tag whole cell lysate

Secondary

All lanes:

Dot 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

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-SPHK2 antibody [EPR29189-33] - BSA and Azide free (AB320742)

This data was developed using ab320741, the same antibody clone in a different buffer formulation. Blocking and diluting buffer and concentration : 5% NFDM/TBST. To minimize protein degradation, cells and tissues were lysed immediately after harvest and loaded onto membranes for Western blotting (Lane 6-9). In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control  (ab181602) staining at 1/200000 dilution. Exposure time : Lane 1-5 : 180 seconds; Lane 6-9 : 70 seconds

All lanes:

Western blot - Anti-SPHK2 antibody [EPR29189-33] (<a href='/en-us/products/primary-antibodies/sphk2-antibody-epr29189-33-ab320741'>ab320741</a>) at 1/1000 dilution

Lane 1:

Mouse liver tissue frozen lysate at 20 µg

Lane 2:

Mouse kidney tissue frozen lysate at 20 µg

Lane 3:

Mouse spleen tissue frozen lysate at 20 µg

Lane 4:

Mouse colon tissue frozen lysate at 20 µg

Lane 5:

Mouse skeletal muscle tissue frozen lysate at 20 µg

Lane 6:

Mouse brain fresh tissue lysate at 20 µg

Lane 7:

Mouse liver fresh tissue lysate at 20 µg

Lane 8:

Mouse testis fresh tissue lysate at 20 µg

Lane 9:

Mouse spleen fresh 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: 70 kDa,36 kDa

false