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

    Abcam recommended secondaries - Goat Anti-Rabbit HRP (ab205718) and Goat Anti-Rabbit Alexa Fluor® 488 (ab150077).

    See other anti-rabbit secondary antibodies that can be used with this antibody.

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 178 Publications for this product

Customer reviews and Q&As

31-40 of 54 Abreviews or Q&A

Application
Immunocytochemistry
Sample
Prairie Vole Cell (brain sections)
Specification
brain sections
Fixative
Paraformaldehyde
Permeabilization
No
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Jul 26 2012

Answer

Thank you for kindly confirming these details. I am sorry this vial of antibody has not worked for this customer.

As requested, we are pleased to arrange a free of charge replacement. I can confirm that 1 vial of ab18723 has been added to your order number #####. This has Abcam order reference #####.

I would like to reassure you that this free of charge replacement vial is also covered by our Abpromise guarantee. Should the customer still be experiencing difficulties with the new vial, or if you have any further questions, please do not hesitate to let me know.

Thank you for your help and cooperation with this case. Please do not hesitate to contact me if you need anything further.

Read More

Question

Inquiry: 1)Antibody storage conditions (temperature/reconstitution etc) -20 °C 2)Description of the problem (high background, low signal, non-specific satining etc.): Non-specific staining and no specific signal 3)Sample (Species/Tissue/Cell Type/Cell Line etc.): Rat Brain (hippocampus) 4)Fixation of sample (Ethanol/Methanol/Acetone/Paraformaldehyde/Other/Duration etc.): Paraformladheyde perfusion followed by immersion and sucrose embedding. Frozen 40 micron sections 5)Antigen retrieval (Enzymatic method, Heat mediated technique etc.): No 6)Permeabilization step: Standard with 0.1% Triton in PBS 7)Blocking conditions (Buffer/time period, Blocking agent etc.): 2.5% normal goat serum in PBS-Triton 8)Primary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step): ab 18723, diluted 1:1000 in blocking buffer, overnight incubation in the cold room. 9)Secondary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step): Anti-rabbit HRP conjugared Santa Cruz or anti-rabbit conjugated with Alexa fluor 568, Invitrogen for immunofluorescence, 2 hour incubation ar room temperature. 10)Detection method: DAB for HRP conjugated secondary antibody 11)Positive and negative controls used (please specify): Positive control, differentiating neuroblasts in juvenile hippocampus. Negative control differentiated neuron in adult hippocampus. 12) How many times have you tried the IHC?: Many times with different lots of the same antibody in the past with good results and 4 times with lot GR 55971-1 with bad results. 13)Have you run a "No Primary" control?: No 14)Do you obtain the same results every time?: Yes 15)What steps have you altered? None Additional Notes We have used for more than one year different lots of the same antibody (no bottle conserved) with satisfactory rwsults (see figures marked as old ab). Storage conditions in house, protocol and material used have been always the same.

Read More
Answer

Thank you for taking time to complete our questionnaire and for contacting us. I am sorry to hear this antibody is not providing satisfactory results.

The details provided will enable us to investigate this case and will provide us with vital information for monitoring product quality. I appreciate the time you have spent in the laboratory and understand your concerns. It is regrettable the results have not been successful.

Having reviewed the protocol details, I believe this product should have given satisfactory results. It appears that you may have received a faulty vial.

I apologize for the inconvenience and am pleased to offer you a free of charge replacement with a different lot of ab18723. Or if you would prefer a credit note. If you would like for me to organise this could you please confirm the order number (or purchase order number) of ab18723.

Thank you for your cooperation. I look forward to hearing from you with details of how you would like to proceed.

Read More

Answer

I have been talking to the lab about ab18723 and they have told me that theres been no change in purification with this ab but it is a tricky in terms of characterising and looking at the notes, it seems there is some variability from AP to AP regarding background staining. We don't batch test this in house in FrFl IHC but we do batch test this in FFPE IHC. The 2 AP's GR55971-1 and GR52629-1 were ones that passed in the hands of a collaborator. For lot GR55971-1, it was used at a 1/1250 dilution and their notes state that a lot of tweaking was necessary to obtain a good signal in relation to background. Lot GR52629-1 was used at 1/750 dilution but also showed a little background. I think generally it's just a case of lot to variability and both these lots were passed on the basis that the specific staining is more intense than the background. I hope that helps but if you need anything else then just let me know.

Read More

Answer

I got the email with images attached, thank you for sending that information. I just want to make sure that I am looking at these images correctly. Are you saying that Lot GR55971-1 is showing specific staining in the SCGZ, but the surrounding staining is non-specific? While lot #GR49156-2 (bottom images) are only showing specific staining. Is that correct? I just want to make sure that I am reading your images correctly before I forward them to the lab to have take a look. I look forward to your reply.

Read More

Answer

Thank you for contacting us. I am sorry to hear you have had difficulty obtaining satisfactory results from this antibody.  Regarding the image sent it seems to be high background and no specific staining on the samples. In order to improve the results from this antibody, I would like to offer a suggestion. It would be worth using different more diluted antibody dilutions, to try reducing the background staining. I would suggest 1/1500 or 1/2000.  I would also appreciate if you could send me the order number, so I can check there has not been any issue during the shipping. In any case, if the antibody was purchased within the last 6 months, it is covered by our Abpromise guarantee, and therefore, I’ll be more than happy to offer you a free replacement of ab18723 form a different lot in case there has been any problem with this particular lot. I hope this information is useful. Please do not hesitate to contact us if you need further advice or information.

Read More

Answer

Thanks for your enquiry. I have attached a Certificate of Compliance for ab18723 as you requested. Please let me know if you need anything further.

Read More
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (brain)
Specification
brain
Fixative
Formaldehyde
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: EDTA pH9.0, 20 min, 100C
Permeabilization
No

Abcam user community

Verified customer

Submitted Oct 11 2011

Answer

Thank you very much for your feedback concerning these two antibodies. I would like to assure you that we are not misrepresenting information. Both antibodies are polyclonal and produced using the same immunogen and were originally considered one product. Each batch (production round) is batch tested by our lab and we noticed that some batches are not suitable for IHC-P. Therefore we split this antibody into two products: ab77450 which is not suitable for IHC-P and ab18723 which is tested suitable for IHC-P. The original WB image was generated before the batches were split into two products and is therefore valid and the same for both. Batch testing for WB was always successful for both batches (products). We do have many additional WB images for each batch and will soon upload more images to prevent further confusion. I would like to thank you for spotting this confusing data and making us aware that we have to provide more information to our users about our (batch-) testing and quality control procedures. I hope I was able to resolve this misunderstanding with this information. Please feel free to contact me again with any further questions.

Read More

Answer

Thank you for contacting us with your question. Although we generally don't recommend using a primary antibody raised in mouse to stain mouse tissue, there are steps that can reduce the background when this kind of staining is necessary. For mouse on mouse staining, the endogenous IgG molecules in the tissue should be blocked by anti-mouse IgG Fab fragments, such as ab6668. The protocol would be as follows: 1) Prepare tissue or cell samples as usual and perform the usual protein block (10% non-mouse serum for 1 hour). 2) Wash 3x2 minutes in TBS and 0.025% Triton x-100 with gentle agitation. 3) Incubate sections with an unconjugated anti-mouse IgG Fab fragment for 1 hour at room temperature or overnight at 4C. 4) Proceed with antibody staining. I hope this information helps, but please let me know if your customer has any further questions.

Read More

31-40 of 54 Abreviews or Q&A

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"
For licensing inquiries, please contact partnerships@abcam.com

Sign up