Anti-ASIC3 antibody [EPR26557-80]

• ICC

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Immunocytochemistry - Anti-ASIC3 antibody [EPR26557-80] (AB302776)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized 293T (human embryonic kidney epithelial cell) cells labelling ASIC3 with ab302776 at 1/4000 (0.124 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 positive staining in HEK-293T cells transfected with a mouse ASIC3 expression vector containing a myc tag. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). ICC has not been tested with endogenous material. is observed. Myc-Tag Mouse mAb (Alexa Fluor® 647) was used to counterstain tubulin at 1/100 0.38 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.

• IHC

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Immunohistochemistry - Anti-ASIC3 antibody [EPR26557-80] (AB302776)

Immunohistochemical analysis of paraffin-embedded Mouse testis tissue labeling ASIC3 with ab302776 at 1/5000 (0.1 ug/ml) followed by a LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Negative control : no staining on mouse testis (PMID : 11872753). The section was incubated with ab302776 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 LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• IHC

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Immunohistochemistry - Anti-ASIC3 antibody [EPR26557-80] (AB302776)

Immunohistochemical analysis of paraffin-embedded Mouse dorsal root ga tissue labeling ASIC3 with ab302776 at 1/5000 (0.1 ug/ml) followed by a LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Positive staining on mouse dorsal root ganglion. The section was incubated with ab302776 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 LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• IHC

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Immunohistochemistry - Anti-ASIC3 antibody [EPR26557-80] (AB302776)

Immunohistochemical analysis of paraffin-embedded Rat dorsal root gang tissue labeling ASIC3 with ab302776 at 1/5000 (0.1 ug/ml) followed by a LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Positive staining on rat dorsal root ganglion (PMID : 9707631). The section was incubated with ab302776 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 LeicaDS9800 (Bond™ Polymer Refine Detection) was used at Ready to use dilution. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• IP

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Immunoprecipitation - Anti-ASIC3 antibody [EPR26557-80] (AB302776)

ASIC3 was immunoprecipitated from 0.35 mg rat dorsal ganglion tissue lysate with ab302776 at 1/30 dilution (2 µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab302776 at 1/1000 dilution.

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

Samples are non-boiled as boiling may cause protein aggregates in lane 1.

This blot was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

All lanes:

Immunoprecipitation - Anti-ASIC3 antibody [EPR26557-80] (ab302776) at 1/1000 dilution

Lanes 1 - 2:

Rat dorsal ganglion tissue lysate at 10 µg

Lane 3:

Rabbit monoclonal IgG (<a href='/ko/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab302776 in rat dorsal ganglion tissue lysate at 10 µg

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

Predicted band size: 58 kDa

Observed band size: 60 kDa

true

Exposure time: 93s

• WB

Lab

Western blot - Anti-ASIC3 antibody [EPR26557-80] (AB302776)

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

It is recommended to optimize experimental conditions by increasing the sample loading amount, using a lower antibody dilution ratio, and employing femtogram-level sensitivity substrates.

Lanes 1 - 4:

Western blot - Anti-ASIC3 antibody [EPR26557-80] (ab302776) at 1/1000 dilution

Lanes 5 - 8:

Western blot - Anti-ASIC3 antibody [EPR26557-87] (<a href='/ko/products/primary-antibodies/asic3-antibody-epr26557-87-ab305243'>ab305243</a>) at 1/1000 dilution

Lanes 1 and 5:

Mouse dorsal ganglion tissue lysate at 20 µg

Lanes 2 and 6:

Rat dorsal ganglion tissue lysate at 20 µg

Lanes 3 and 7:

Mouse dorsal ganglion tissue lysate at 40 µg

Lanes 4 and 8:

Rat dorsal ganglion tissue lysate at 40 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 1/20000 dilution

Predicted band size: 58 kDa

Observed band size: 60 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-ASIC3 antibody [EPR26557-80] (AB302776)

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

The MW is consistent with what has been described in the literature (PMID : 17012229).

Negative control : rat : hippocampus, spinal cord, kidney, testis (PMID : 11872753).

Samples are non-boiled as boiling may cause protein aggregates.

This blot was developed using a high sensitivity ECL substrate in lane 6 and 7. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

Exposure time :

Lane 1-5 : 180 seconds

Lane 6 and 7 : 37 seconds.

All lanes:

Western blot - Anti-ASIC3 antibody [EPR26557-80] (ab302776) at 1/1000 dilution

Lane 1:

Rat dorsal ganglion tissue lysate at 75 µg

Lane 2:

Rat hippocampus tissue lysate at 75 µg

Lane 3:

Rat spinal cord tissue lysate at 75 µg

Lane 4:

Rat kidney tissue lysate at 75 µg

Lane 5:

Rat testis tissue lysate at 75 µg

Lane 6:

Mouse dorsal ganglion tissue lysate at 75 µg

Lane 7:

Mouse hippocampus tissue lysate at 75 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 1/20000 dilution

Predicted band size: 58 kDa

Observed band size: 60 kDa

true