Recombinant Anti-ADNP antibody [EPR25434-4] (ab300114)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR25434-4] to ADNP
- Suitable for: IHC-Fr, Flow Cyt (Intra), IP, WB, IHC-P, ICC/IF
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
Overview
-
Product name
Anti-ADNP antibody [EPR25434-4]
See all ADNP primary antibodies -
Description
Rabbit monoclonal [EPR25434-4] to ADNP -
Host species
Rabbit -
Tested applications
Suitable for: IHC-Fr, Flow Cyt (Intra), IP, WB, IHC-P, ICC/IFmore details
Unsuitable for: ChIP -
Species reactivity
Reacts with: Mouse, Rat, Human -
Immunogen
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
-
Positive control
- WB: 293T, NIH/3T3, PC-12, HeLa, C6, U-87 MG, SH-SY5Y, Neuro-2a, mouse brain and rat brain lysates. IHC-P: Human colon carcinoma, mouse cerebrum, mouse glioblastoma and rat cerebrum tissues. IHC-Fr: Mouse cerebrum and rat cerebrum tissues. ICC/IF: HeLa and NIH/3T3 cells. Flow Cyt. Intra.: HeLa and NIH/3T3 cells. IP: HeLa and NIH/3T3 cells.
-
General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Properties
-
Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
-
Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR25434-4 -
Isotype
IgG -
Research areas
Associated products
-
Alternative Versions
-
Compatible Secondaries
-
Isotype control
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab300114 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application | Abreviews | Notes |
---|---|---|
IHC-Fr |
1/500.
|
|
Flow Cyt (Intra) |
1/500.
|
|
IP |
1/30.
|
|
WB |
1/1000. Detects a band of approximately 140 kDa (predicted molecular weight: 124 kDa).
|
|
IHC-P |
1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
|
|
ICC/IF |
1/50.
|
Notes |
---|
IHC-Fr
1/500. |
Flow Cyt (Intra)
1/500. |
IP
1/30. |
WB
1/1000. Detects a band of approximately 140 kDa (predicted molecular weight: 124 kDa). |
IHC-P
1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
ICC/IF
1/50. |
Target
- Information by UniProt
-
Database links
- Entrez Gene: 23394 Human
- Entrez Gene: 11538 Mouse
- Entrez Gene: 64622 Rat
- Omim: 611386 Human
- SwissProt: Q9H2P0 Human
- SwissProt: Q9Z103 Mouse
- SwissProt: Q9JKL8 Rat
- Unigene: 570355 Human
see all -
Alternative names
- Activity dependent neuroprotective protein antibody
- Activity dependent neuroprotector antibody
- Activity-dependent neuroprotective protein antibody
see all
Images
-
All lanes : Anti-ADNP antibody [EPR25434-4] (ab300114) at 1/1000 dilution
Lane 1 : 293T (human embryonic kidney epithelial cell) whole cell lysate
Lane 2 : NIH/3T3 (mouse embryonic fibroblast) whole cell lysate
Lane 3 : PC-12 (rat adrenal gland pheochromocytoma) whole cell lysate
Lane 4 : HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 124 kDa
Observed band size: 140 kDa why is the actual band size different from the predicted?Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time:
Lanes 1-3: 114 seconds; Lane 4: 59 seconds.
Lysates should be made freshly and used in WB immediately to minimize protein degradation.
-
All lanes : Anti-ADNP antibody [EPR25434-4] (ab300114) at 1/1000 dilution
Lane 1 : C6 (rat glial tumor glial cell) whole cell lysate
Lane 2 : U-87 MG (human glioblastoma-astrocytoma epithelial cell) whole cell lysate
Lane 3 : SH-SY5Y (human neuroblastoma epithelial cell) whole cell lysate
Lane 4 : Neuro-2a (mouse neuroblastoma neuroblast) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 124 kDa
Observed band size: 140 kDa why is the actual band size different from the predicted?
Exposure time: 3 minutesBlocking and dilution buffer and concentration: 5% NFDM/TBST.
Lysates should be made freshly and used in WB immediately to minimize protein degradation.
-
All lanes : Anti-ADNP antibody [EPR25434-4] (ab300114) at 1/1000 dilution
Lane 1 : Mouse brain tissue lysate
Lane 2 : Rat brain tissue lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 124 kDa
Observed band size: 140 kDa why is the actual band size different from the predicted?
Exposure time: 3 minutesBlocking and dilution buffer and concentration: 5% NFDM/TBST.
Lysates should be made freshly and used in WB immediately to minimize protein degradation. We used fresh mouse brain tissue lysate.
This blot was developed using a higher sensitivity ECL substrate. -
Immunohistochemical analysis of paraffin-embedded Human colon carcinoma tissue labelling ADNP with ab300114 at 1/100 dilution followed by a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection). Nuclear staining on human colon carcinoma is observed (PMID: 27903678). The section was incubated with ab300114 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: PBS was used instead of primary antibody followed by ready to use secondary antibody LeicaDS9800 (BOND™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
-
Immunohistochemical analysis of paraffin-embedded Mouse cerebrum tissue labelling ADNP with ab300114 at 1/100 dilution followed by a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection) was used. Nuclear staining on mouse cerebrum is observed. The section was incubated with ab300114 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument Counterstained with Hematoxylin.
Secondary antibody only control: PBS was used instead of primary antibody followed by ready to use secondary antibody LeicaDS9800 (BOND™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
-
Immunohistochemical analysis of paraffin-embedded mouse glioblastoma tissue labelling ADNP with ab300114 at 1/100 dilution followed by a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection). Nuclear staining on mouse glioblastoma is observed. The section was incubated with ab300114 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: PBS was used instead of primary antibody followed by ready to use secondary antibody LeicaDS9800 (BOND™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
-
Immunohistochemical analysis of paraffin-embedded rat cerebrum tissue labelling ADNP with ab300114 at 1/100 dilution followed by a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection). Nuclear staining on rat cerebrum is observed. The section was incubated with ab300114 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.
Secondary antibody only control: PBS was used instead of primary antibody followed by ready to use secondary antibody LeicaDS9800 (BOND™ Polymer Refine Detection).
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.
-
Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen mouse cerebrum (fresh) tissue labeling ADNP with ab300114 at 1/500 dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (Green). Positive staining on mouse cerebrum is observed. The nuclear counterstain was DAPI (Blue).
Secondary antibody control: PBS was used instead of ab300114 followed by preadsorbed secondary antibody (ab150081) Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.
-
mmunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen rat cerebrum (fresh) tissue labeling ADNP with ab300114 at 1/500 dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (Green). Positive staining on rat cerebrum is observed. The nuclear counterstain was DAPI (Blue).
PBS was used instead of ab300114 followed by preabsorbed secondary antibody (ab150081) Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.
-
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeilized HeLa (human cervix adenocarcinoma epithelial cell) cells lebelling ADNP with ab300114 at 1/50 dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green). Confocal image showing nuclear staining in HeLa cell line. Tubuline was stained with ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: PBS was used instead of ab300114 followed by preabsorbed secondary antibody (ab150081) Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.
-
This data was developed using ab300114, the same antibody clone in a different buffer formulation.
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeilized NIH/3T3 (mouse embryonic fibroblast) cells lebelling ADNP with ab300114 at 1/50 (11.22 µg/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2µg/mL) dilution (Green). Confocal image showing nuclear staining in NIH/3T3 cell line is observed. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 (2.5 µg/ml) dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody control: PBS was used instead of ab300114 followed by preabsorbed secondary antibody (ab150081) Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 (2 µg/mL) dilution.
-
Intracellular flow cytometric analysis of 4% paraformaldehyde fixed and 90% methnol permeabilized HeLa (human cervix adenocarcinoma epithelial cell) cells labelling ADNP with ab300114 at 1/500 dilution (Red) compared with a rabbit monoclonal IgG (ab172730) (Black) isotype control. Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/2000 dilution was used as the secondary antibody.
-
Intracellular flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized NIH/3T3 (mouse embryonic fibroblast) cells labelling ADNP with ab300114 at 1/500 dilution (Red) (Red) compared with a rabbit monoclonal IgG (ab172730) (Black) isotype control. A Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/2000 dilution was used as the secondary antibody.
-
ADNP was immunoprecipitated from 0.35 mg HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate with ab300114 at 1/30 dilution. Western blot was performed on the immunoprecipitate using ab300114 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP) (ab131366) was used at 1/5000 dilution.
Lane 1 (Input): HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate 10 µg
Lane 2 (+): ab300114 IP in HeLa whole cell lysate
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab300114 in HeLa whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 84 seconds
-
ADNP was immunoprecipitated from 0.35 mg NIH/3T3 (mouse embryonic fibroblast) whole cell lysate with ab300114 at 1/30 dilution. Western blot was performed on the immunoprecipitate using ab300114 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP) (ab131366) was used at 1/5000 dilution.
Lane 1 (Inset): NIH/3T3 (mouse embryonic fibroblast) whole cell lysate 10 µg
Lane 2 (+): ab300114 IP in NIH/3T3 whole cell lysate
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab300114 in NIH/3T3 whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 3 minutes.
Protocols
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
Datasheets and documents
-
SDS download
-
Datasheet download
Certificate of Compliance
References (1)
ab300114 has been referenced in 1 publication.
- Wang X et al. ADNP is associated with immune infiltration and radiosensitivity in hepatocellular carcinoma for predicting the prognosis. BMC Med Genomics 16:178 (2023). PubMed: 37525242