Anti-Doublecortin antibody (ab18723)

Rabbit polyclonal Doublecortin antibody. Validated in WB, IHC, ICC, ICC/IF and tested in Mouse, Rat, Chicken, Cat, Human, Cynomolgus monkey, Quail, Rhesus monkey. Cited in 177 publication(s).

Overview

  • Product name
    Anti-Doublecortin antibody
    See all Doublecortin primary antibodies
  • Description
    Rabbit polyclonal to Doublecortin
  • Host species
    Rabbit
  • Specificity

    Please note: Low dilutions of this antibody can cause high background in IHC. Please use as high a dilution as possible. Optimal working dilutions are batch dependent.

  • Tested applications
    Suitable for: WB, ICC, ICC/IF, IHC-FrFl, IHC-FoFr, IHC-Fr, IHC-Pmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Chicken, Cat, Human, Cynomolgus monkey, Quail, Rhesus monkey
  • Immunogen

    Synthetic peptide conjugated to KLH derived from within residues 300 to the C-terminus of Human Doublecortin.

    Read Abcam's proprietary immunogen policy (Peptide available as ab19804.)

  • Positive control
    • This antibody gave a positive signal in Brain (Mouse) Tissue Lysate - normal tissue, 0 days old ICC-IF: SHSY5Y cells. IHC-P: FFPE Rat brain 6 weeks/FFPE Mouse Brain 8 weeks.
  • General notes

      

Properties

Applications

Our Abpromise guarantee covers the use of ab18723 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
WB Use a concentration of 1 µg/ml. Detects a band of approximately 45 kDa (predicted molecular weight: 40-45 kDa).
ICC Use at an assay dependent concentration.

We recommend Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081) secondary antibody.

ICC/IF Use a concentration of 1 - 5 µg/ml.
IHC-FrFl 1/1000. Please note: Low dilutions of this antibody can cause high background. Please use as high a dilution as possible. Optimal working dilutions are batch dependent.
IHC-FoFr 1/100 - 1/2000.
IHC-Fr 1/2000 - 1/7000.
IHC-P Use a concentration of 0.05 - 1 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Target

  • Function
    Seems to be required for initial steps of neuronal dispersion and cortex lamination during cerebral cortex development. May act by competing with the putative neuronal protein kinase DCAMKL1 in binding to a target protein. May in that way participate in a signaling pathway that is crucial for neuronal interaction before and during migration, possibly as part of a calcium ion-dependent signal transduction pathway. May be part with LIS-1 of an overlapping, but distinct, signaling pathways that promote neuronal migration.
  • Tissue specificity
    Highly expressed in neuronal cells of fetal brain (in the majority of cells of the cortical plate, intermediate zone and ventricular zone), but not expressed in other fetal tissues. In the adult, highly expressed in the brain frontal lobe, but very low expression in other regions of brain, and not detected in heart, placenta, lung, liver, skeletal muscles, kidney and pancreas.
  • Involvement in disease
    Defects in DCX are the cause of lissencephaly X-linked type 1 (LISX1) [MIM:300067]; also called X-LIS or LIS. LISX1 is a classic lissencephaly characterized by mental retardation and seizures that are more severe in male patients. Affected boys show an abnormally thick cortex with absent or severely reduced gyri. Clinical manifestations include feeding problems, abnormal muscular tone, seizures and severe to profound psychomotor retardation. Female patients display a less severe phenotype referred to as 'doublecortex'.
    Defects in DCX are the cause of subcortical band heterotopia X-linked (SBHX) [MIM:300067]; also known as double cortex or subcortical laminar heterotopia (SCLH). SBHX is a mild brain malformation of the lissencephaly spectrum. It is characterized by bilateral and symmetric plates or bands of gray matter found in the central white matter between the cortex and cerebral ventricles, cerebral convolutions usually appearing normal.
    Note=A chromosomal aberration involving DCX is found in lissencephaly. Translocation t(X;2)(q22.3;p25.1).
  • Sequence similarities
    Contains 2 doublecortin domains.
  • Cellular localization
    Cytoplasm.
  • Information by UniProt
  • Database links
  • Alternative names
    • DBCN antibody
    • Dbct antibody
    • DC antibody
    • DCX antibody
    • DCX_HUMAN antibody
    • Doublecortex antibody
    • Doublin antibody
    • FLJ51296 antibody
    • Lis X antibody
    • Lis-X antibody
    • Lissencephalin X antibody
    • Lissencephalin-X antibody
    • Lissencephaly X linked antibody
    • Lissencephaly X linked doublecortin antibody
    • LISX antibody
    • Neuronal migration protein doublecortin antibody
    • OTTHUMP00000023859 antibody
    • OTTHUMP00000023860 antibody
    • OTTHUMP00000216315 antibody
    • OTTHUMP00000216316 antibody
    • SCLH antibody
    • XLIS antibody
    see all

Images

  • IHC image of ab18723 staining in Mouse 8 weeks brain formalin fixed paraffin embedded tissue section, performed on a Leica BONDTM system using the standard protocol F (with no post primary). The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab18723, 0.1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. Secondary-only control image is shown as insert.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • ab18723 staining Doublecortin in mouse embryonic 15 day brain tissue sections by Immunohistochemistry (PFA perfusion fixed frozen sections). Tissue samples were fixed by perfusion with paraformaldehyde, permeablized with 0.5% Triton X-100 in PBS, blocked with 10% serum for 1 hour at 25°C and antigen retrieval was by heat mediation in citrate buffer, pH 6. The sample was incubated with primary antibody (1/500 in PBS + 0.1% Triton X-100 + 1% serum) for 16 hours at 25°C. An Alexa Fluor® 594-conjugated donkey anti-rabbit IgG polyclonal (1/700) was used as the secondary antibody.

    See Abreview

  • IHC-Fr image of Doublecortin staining in Mouse adult dentate gyrus sections using ab18723(1:100). The sections were fixed in paraformaldehyde and permeabilized using 1x TBST. The sections were then blocked using 10% donkey serum for 1 hour at 25°C. ab18723 was diluted 1:100 and incubated with the sections for 12 hours at 25°C. The secondary antibody used was Donkey anti-rabbit conjugated to Cy3 Dye (1:500). DAPI was used to stain the nuclei.

    See Abreview

  • IHC image of ab18723 staining in rat 6 week brain formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6) for 20 mins. The section was then incubated with ab18723, 0.1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • ab18723 staining 6 week rat brain tissue dentate gyrus (DG) by IHC-P using rabbit-specific EXPOSE IHC detection kit (ab80437). Formalin fixed paraffin embedded tissue sections were pre-treated using heat mediated antigen retrieval (using a pressure cooker) with sodium citrate buffer (pH6) for 30 mins. The section was incubated with ab18723, 0.1µg/ml, for 1 hour at room temperature. DAB was used as the chromogen and the section was counterstained with haematoxylin and mounted with DPX.

  • ab18723 at 1/200 staining mouse E18 body T/S spinal cord tissue sections by IHC-P. The tissue was formaldehyde fixed and a heat mediated antigen retireval step was perfomed before incubation with the antibody for 24 hours. A biotinylated goat antibody was used as the secondary.

    See Abreview

  • ab18723 stained in SHSY5Y cells. Cells were fixed with 100% methanol (5 min) at room temperature and incubated with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% triton for 1h at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab18723 at 5 µg/ml and ab7291 (Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control) at 1/1000 dilution overnight at +4°C. The secondary antibodies were Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody (pseudo-colored red) and Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081) secondary antibody (colored green) used at 1 ug/ml for 1hour at room temperature. DAPI was used to stain the cell nuclei (colored blue) at a concentration of 1.43µM for 1hour at room temperature.

  • Doublecortin antibody (ab18723; green) labeling cell extensions consistent with dendrite morphology in 4 day old cultures. Preincubation of ab18723 with its immunising peptide (ab19804) quenched immunostaining (see review).

    Dorsal root ganglion explants were dissected from 16 day-old rat embryos and cultured for 4 days in vitro with Neurobasal Medium containing 10% fetal calf serum and B27 supplement. Immunocytochemistry: All steps were performed in PBS. Cells or explants were fixed in 4% PFA for 15min, permeabilised with 0.1% TX100 for 10min and blocked with 5% BSA, 0.1% TX100 for 45min. ab18723 was incubated at 5µg/ml for 12h in 5% BSA, 0.1% TX100 at 4°C. For peptide blocking experiments preincubation of the peptide (ab19804; 250µg/ml) and antibody (5µg/ml ) was performed for 2h at 37°C. Cultures were washed (3x) of primary antibody solution. Goat anti-rabbit AlexaFluor 488 was used as secondary antibody (1/400) in 5% BSA, 0.1% TX100 for 2h at R

  • Doublecortin expression in the dentate gyrus of a 1 month-old mouse brain. Doublecortin staining using ab18723 (1/500) in the dentate gyrus of a 1 month-old mouse brain. The mouse has been perfused with paraformaldehyde 4% (50ml). After dissection, the brain has been incubated overnight in sucrose 20%, embedded in OCT and cryosectioned (10 µm). No antigen retrieval was used. The secondary antibody used was a non-Abcam Goat anti-rabbit Alexa488.
  • IHC image of ab18723 staining in rat 6 week brain formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6) for 20 mins. The section was then incubated with ab18723, 0.1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • Immunohistochemistical staining (on formaldehyde/PFA-fixed paraffin-embedded sections) of Doublecortin antibody - Neuronal Marker (ab18723) on Quail Tissue sections (E6/7 brain (Saggital section). Antigen retrieval step: Heat mediated. Blocking step: 1% BSA for 10 mins at RT. Primary Antibody ab18723 incubated at 1/400 for 2 hours RT. Secondary Antibody: Biotin conjugated goat anti rabbit Ig (1/300).
  • ab18723 at 1/2000 staining mouse brain svr: progenitor olfactory neurones by IHC-P. The tissue was formaldehyde fixed and a heat mediated antigen retrieval step was performed before incubation with the antibody for 16 hours. An Alexa-Fluor ® 488 conjugated goat antibody was used as the secondary (green). The tissue was also stained for Ki67 (shown in red).

    The MIP image was derived from Apotome-generated Z-stacks from the greyscale image of each of the channels in the MIP.

    See Abreview

  • ab18723 staining Doublecortin in mouse dentate gyrus hippocampal tissue section by Immunohistochemistry (PFA perfusion fixed frozen sections). Tissue samples were incubated with primary antibody 1/1000 for 16 hours at 4°C.  An Alexa Fluor®488 conjugated goat polyclonal to rabbit IgG at 1/400 dilution was used as secondary.

    See Abreview

  • Anti-Doublecortin antibody (ab18723) at 1 µg/ml + Mouse brain tissue lysate - total protein (0 days) (ab7188) at 10 µg

    Secondary
    Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution

    Performed under reducing conditions.

    Predicted band size: 40-45 kDa
    Observed band size: 45 kDa
    why is the actual band size different from the predicted?

References

This product has been referenced in:
  • Payne SL  et al. Initial cell maturity changes following transplantation in a hyaluronan-based hydrogel and impacts therapeutic success in the stroke-injured rodent brain. Biomaterials 192:309-322 (2019). Read more (PubMed: 30468998) »
  • Nkomozepi P  et al. Age-related changes in Ki-67 and DCX expression in the BALB/ c mouse (Mus Musculus) brain. Int J Dev Neurosci 72:36-47 (2019). Read more (PubMed: 30472241) »
See all 189 Publications for this product

Customer reviews and Q&As

41-50 of 54 Abreviews or Q&A

Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (Adult brain, dentate gyrus)
Specification
Adult brain, dentate gyrus
Fixative
Paraformaldehyde
Permeabilization
Yes - TBST 1x
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Oct 12 2010

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (brain)
Specification
brain
Fixative
Paraformaldehyde
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: citrate
Permeabilization
No
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 21°C

Dr. Diego Gomez-Nicola

Verified customer

Submitted Sep 24 2010

Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (Brain)
Specification
Brain
Fixative
Paraformaldehyde
Permeabilization
Yes - Triton X-100
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 22°C

Abcam user community

Verified customer

Submitted Feb 11 2010

Application
Immunohistochemistry (Frozen sections)
Sample
Rat Tissue sections (Whole embryo)
Specification
Whole embryo
Fixative
Paraformaldehyde
Permeabilization
No
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Dec 14 2009

Application
Immunohistochemistry free floating
Sample
Mouse Tissue sections (Brain, perfusion fixed 4% FA, 40 um)
Specification
Brain, perfusion fixed 4% FA, 40 um

Abcam user community

Verified customer

Submitted Oct 13 2009

Application
Immunohistochemistry free floating
Sample
Mouse Tissue sections (dentate gyrus of hippcampus)
Specification
dentate gyrus of hippcampus

Abcam user community

Verified customer

Submitted Sep 04 2009

Question

BATCH NUMBER -- NOT SPECIFIED -- ORDER NUMBER 719809/2 DESCRIPTION OF THE PROBLEM We have a problem with the abcam rabbit policlonal antibody to doublecortin (ab18723), recently bought from Prodotti Gianni (719809/2 29/11/07, delivered on 05/02/08). Immunoistochemical staining (both immunofluorescence and DAB stainig) on free floating, formalin fixed, 40 micron rat brain sections shows deep differences with the usual DCX staining. In the dentate gyrus, for example, we did not find the typical neuronal labelling and, above of all, we find clearly marked astroglial cells throught the brain, as indicated in the enclosed microphotographs. Obviously we exclude any contamination with other antibodies. Our experimental procedure is in agreement with that indicated in abcam data sheet. Can you help us? SAMPLE Rat brain PRIMARY ANTIBODY anti doublecortin policlonal antibody 1:500 in 3% NGS. DETECTION METHOD Immunofluorescence ABC - DAB POSITIVE AND NEGATIVE CONTROLS USED Positive control: hippocampal dentate gyrus Negetive control: not done ANTIBODY STORAGE CONDITIONS stored at -20° FIXATION OF SAMPLE Formalin ANTIGEN RETRIEVAL No antigen retrieval PERMEABILIZATION STEP No permeabilization step BLOCKING CONDITIONS 1h 3% Normal Goat serum in PBS SECONDARY ANTIBODY Anti rabbit Cy3 1:200 Biotinylated anti rabbit 1:200 HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? three HAVE YOU RUN A "NO PRIMARY" CONTROL? Yes DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes WHAT STEPS HAVE YOU ALTERED? I don't know what do you mean

Read More
Answer

Thank you for contacting us for technical support with the antibody ab18723. We are unable to trace your order from Abcam for this product I'm afraid, and it seems that you have purchased the antibody through a local italian company who is not authorised in reselling our products. Unfortunately as we cannot control the storage and shipping conditions that Prodotti Gianni have used, it may be that the antibody was damaged in their hands. We ship directly to Italy and as this antibody is very popular we have a lot in stock and if you place an order you should receive the vial within a few days. Regarding the protocol you have used and the image you kindly sent to us, it seems that the staining you are observing is very strong neuronally with additional glial cells marked. This non specific binding of the antibody may be due to an excess of primary antibody combined with no permeabilisation. As the protein is at the cytoplasmic level, I would expect permeabilisation to be required. This in turn will mean you do not need as much antibody to achive some staining, therefore reducing non specific binding to other proteins. From your protocol it is not clear how long you postfixed the tissue (I imagine it is perfusion fixed) therefore I would like to highlight that over or under fixation can also lead to non specific staining. Depending on the size and age of the tissue you may need to post fix for 2 hours only, or alternatively postfix overnight so I recommend trying different time points of postfixation. As I mentioned above, the addition of 0.3%tritonx100 to the PBS to dilute the serum, primary and secondary antibody is advisable, and to decrease the concentration of primary and secondary antibody to achieve good but specific staining. Finally, can I please make sure that you rinse extensively in PBS between each step (except between serum and primary antibody) as any remains of fixative can also cause non specific staining. I hope the above suggestions will help you. Should you still experience problems I suggest to contact Prodotti Gianni to obtain a replacement vial from them, as unfortunately I am unable to offer a replacement vial if purchased through non-authorised distributors. Best of luck with your experiments.

Read More
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Quail Tissue sections (E6/7 brain (Saggital section))
Specification
E6/7 brain (Saggital section)
Fixative
Formaldehyde
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: 10mM Citric acid pH6
Permeabilization
No
Blocking step
BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: RT°C

Mr. Carl Hobbs

Verified customer

Submitted Dec 03 2007

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (brain svr: progenitor olfactory neurones)
Specification
brain svr: progenitor olfactory neurones
Fixative
Formaldehyde
Antigen retrieval step
Heat mediated
Blocking step
BSA as blocking agent for 10 minute(s) · Concentration: 1

Mr. Carl Hobbs

Verified customer

Submitted Feb 21 2007

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (E18 body T/S spinal cord)
Specification
E18 body T/S spinal cord
Fixative
Formaldehyde
Antigen retrieval step
Heat mediated
Blocking step
BSA as blocking agent for 10 minute(s) · Concentration: 1%

Mr. Carl Hobbs

Verified customer

Submitted Feb 14 2007

41-50 of 54 Abreviews or Q&A

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