Anti-SIGLEC8 antibody [EPR24419-56] - BSA and Azide free

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-SIGLEC8 antibody [EPR24419-56] - BSA and Azide free (AB305298)

This data was developed using ab305297, the same antibody clone in a different buffer formulation.

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HEK-293T (human embryonic kidney epithelial cell) cells labelling SIGLEC8 with ab305297 at 1/1000 dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in HEK-293T cells transfected with a human SIGLEC8 expression vector containing a myc tag. The image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

Myc-Tag Mouse mAb (Alexa Fluor® 647) was used to counterstain tubulin at 1/100 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 dilution.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-SIGLEC8 antibody [EPR24419-56] - BSA and Azide free (AB305298)

This data was developed using ab305297, the same antibody clone in a different buffer formulation.

Flow cytometric analysis of 2% paraformaldehyde fixed 0.1% Tween-20 permeabilized HEK-293T (human embryonic kidney) cells transfected with a human SIGLEC8 expression vector containing a myc-his tag cells labelling SIGLEC8 with ab305297 at 1/50 dilution (1ug)/ Right compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/2000 dilution was used as the secondary antibody. Live cells were surface stained with ab305297, then fixed with 2% PFA followed by intracellular staining with Myc-tag conjugated to Alexa Fluor® 647.

• IP

Supplier Data

Immunoprecipitation - Anti-SIGLEC8 antibody [EPR24419-56] - BSA and Azide free (AB305298)

This data was developed using ab305297, the same antibody clone in a different buffer formulation. SIGLEC8 was immunoprecipitated from 0.35 mg HEK-293T (human embryonic kidney epithelial cell) cells transfected with a human SIGLEC8 expression vector containing a myc-His-tag®, whole cell lysate, 2 μg with ab305297 at 1/30 dilution (2μg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab305297 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution. Lane 1 : HEK-293T (human embryonic kidney epithelial cell) cells transfected with a human SIGLEC8 expression vector containing a myc-His-tag®, whole cell lysate, 2 μg Lane 2 : ab305297 IP in HEK-293T cells transfected with a human SIGLEC8 expression vector containing a myc-His-tag®, whole cell lysate Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab305297 in HEK-293T cells transfected with a human SIGLEC8 expression vector containing a myc-His-tag®, whole cell lysate Blocking and dilution buffer and concentration : 5% NFDM/TBST. Exposure time : 3 seconds

All lanes:

Immunoprecipitation - Anti-SIGLEC8 antibody [EPR24419-56] (<a href='/en-us/products/primary-antibodies/siglec8-antibody-epr24419-56-ab305297'>ab305297</a>) at 1/30 dilution

All lanes:

HEK-293T (human embryonic kidney epithelial cell)

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

Observed band size: 60 kDa

false

Exposure time: 3s

• WB

Supplier Data

Western blot - Anti-SIGLEC8 antibody [EPR24419-56] - BSA and Azide free (AB305298)

This data was developed using ab305297, the same antibody clone in a different buffer formulation. Blocking and diluting buffer and concentration : 5% NFDM/TBST Exposure time : 1 second

All lanes:

Western blot - Anti-SIGLEC8 antibody [EPR24419-56] (<a href='/en-us/products/primary-antibodies/siglec8-antibody-epr24419-56-ab305297'>ab305297</a>) at 1/1000 dilution

Lane 1:

HEK-293T cells transfected with a human SIGLEC8 expression vector containing a myc-His-tag®, whole cell lysate at 20 µg

Lane 2:

HEK-293T transfected with an empty vector (vector control), 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: 60 kDa

false

Exposure time: 1s