重组Anti-SNX1抗体[EPR7178]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SNX1 antibody [EPR7178] (AB134126)

Immunohistochemical analysis of paraffin embedded Human breast carcinoma labelling SNX1 with ab134126 at 1/250.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-SNX1 antibody [EPR7178] (AB134126)

ab134126 stained HeLa cells. The cells were 4% formaldehyde fixed for 10 minutes at room temperature and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1hour at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab134126 at 1/100 dilution) overnight at +4°C. The secondary antibody (pseudo-colored green) was Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081) used at a 1/1000 dilution for 1hour at room temperature. Alexa Fluor® 594 WGA was used to label plasma membranes (pseudo-colored red) at a 1/200 dilution for 1hour at room temperature. DAPI was used to stain the cell nuclei (pseudo-colored blue) at a concentration of 1.43μM for 1hour at room temperature.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-SNX1 antibody [EPR7178] (AB134126)

Immunofluorescent analysis of 4% Paraformaldehyde fixed 0.1% Triton X-100 permeabilized rat primary neural/glia cells labelling SNX1 with ab134126 at 1/250 (1.864 μg/ml) followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed antibody as secondary at 1/1000 (2 μg/ml) dilution. Confocal image showing cytoplasmic staining in rat primary neural/glia cell (shown in green). ab11267 Anti-MAP2 mouse monoclonal antibody at 1/500 (4μg/ml) was used as a counterstain along with ab150120 as secondary at 1/1000 (2μg/ml) (shown in magenta). Nuclear DNA was labelled with DAPI (shown in blue). Confocal scanning Z step was set as 0.3 μm followed by image processing with maximum Z projection. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-SNX1 antibody [EPR7178] (AB134126)

Immunofluorescent analysis of 4% Paraformaldehyde fixed 0.1% Triton X-100 permeabilized mouse primary neural/glia cells labelling SNX1 with ab134126 at 1/250 (1.864 μg/ml) followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed antibody as secondary at 1/1000 (2 μg/ml) dilution. Confocal image showing cytoplasmic staining in mouse primary neural/glia cell (shown in green). ab11267 Anti-MAP2 mouse monoclonal antibody at 1/500 (4μg/ml) was used as a counterstain along with ab150120 as secondary at 1/1000 (2μg/ml) (shown in magenta). Nuclear DNA was labelled with DAPI (shown in blue). Confocal scanning Z step was set as 0.3 μm followed by image processing with maximum Z projection. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-SNX1 antibody [EPR7178] (AB134126)

Immunofluorescent analysis of 4% Paraformaldehyde fixed 0.1% Triton X-100 permeabilized EL4 (mouse lymphoma T lymphocyte) cells labelling SNX1 with ab134126 at 1/250 (1.864 μg/ml) followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed antibody as secondary at 1/1000 (2 μg/ml) dilution. Negative control : Confocal image showing no staining in EL4 cells (shown in green). ab7291 Anti-alpha Tubulin mouse monoclonal antibody at 1/1000 (1μg/ml) was used as a counterstain along with ab150120 as secondary at 1/1000 (2μg/ml) (shown in magenta). Nuclear DNA was labelled with DAPI (shown in blue). Confocal scanning Z step was set as 0.3 μm followed by image processing with maximum Z projection. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SNX1 antibody [EPR7178] (AB134126)

Immunohistochemical analysis of paraffin-embedded rat spleen tissue labeling SNX1 with ab134126 at 1/5000 dilution.

Granular staining on rat spleen.

The section was incubated with ab134126 for 30 mins at room temperature.

The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.

Counterstained with Hematoxylin.

Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SNX1 antibody [EPR7178] (AB134126)

Immunohistochemical analysis of paraffin-embedded rat skeletal muscle tissue labeling SNX1 with ab134126 at 1/5000 dilution.

Low expression tissue : weak staining on blood vessel of rat skeletal muscle (PMID : 11110793)..

The section was incubated with ab134126 for 30 mins at room temperature.

The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.

Counterstained with Hematoxylin.

Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins.

• WB

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Western blot - Anti-SNX1 antibody [EPR7178] (AB134126)

All lanes:

Western blot - Anti-SNX1 antibody [EPR7178] (ab134126) at 1/10000 dilution

Lane 1:

HeLa cell lysate at 10 µg

Lane 2:

HepG2 cell lysate at 10 µg

Lane 3:

293T cell lysate at 10 µg

Secondary

All lanes:

Goat anti-rabbit HRP conjugated at 1/2000 dilution

Predicted band size: 59 kDa

Observed band size: 74 kDa

false

• WB

Lab

Western blot - Anti-SNX1 antibody [EPR7178] (AB134126)

Western blot : Anti-SNX1 antibody [EPR7178] ab134126 staining at 1/10000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 70 kDa in Wild-type HeLa cell lysates with no signal observed at this size in SNX1 knockout HeLa cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5% Milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-SNX1 antibody [EPR7178] (ab134126) at 1/10000 dilution

Lane 1:

Western blot - Human wild-type HeLa cell line (ab271142) at 20 µg

Lanes 2 and 4:

Empty

Lane 3:

Western blot - Human SNX1 knockout HeLa cell line (<a href='/products/cell-lines/human-snx1-knockout-hela-cell-line-ab273834'>ab273834</a>) at 20 µg

Lane 5:

MCF7 at 20 µg

Lane 6:

A549 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 74 kDa,70 kDa

false

• WB

Lab

Western blot - Anti-SNX1 antibody [EPR7178] (AB134126)

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

All lanes:

Western blot - Anti-SNX1 antibody [EPR7178] (ab134126) at 1/1000 dilution

Lane 1:

Rat brain tissue lysate at 20 µg

Lane 2:

Rat kidney tissue lysate at 20 µg

Lane 3:

Rat spleen tissue lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 63 kDa,74 kDa

false

Exposure time: 8s

• WB

Lab

Western blot - Anti-SNX1 antibody [EPR7178] (AB134126)

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-SNX1 antibody [EPR7178] (ab134126) at 1/1000 dilution

Lane 1:

C6 (rat glial tumor glial cell) transfected with scrambled siRNA control, whole cell lysate at 20 µg

Lane 2:

C6 transfected with siRNA specifically targeting SNX1, whole cell lysate at 20 µg

Secondary

All lanes:

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

Observed band size: 74 kDa,36 kDa

false

Exposure time: 180s

• OI-RD Scanning

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OI-RD Scanning - Anti-SNX1 antibody [EPR7178] (AB134126)

We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody. Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.