Overview

  • Product name
    Anti-Doublecortin antibody - Neuronal Marker
    See all Doublecortin primary antibodies
  • Description
    Rabbit polyclonal to Doublecortin - Neuronal Marker
  • Host species
    Rabbit
  • Tested applications
    Suitable for: WB, ICC/IFmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
    Predicted to work with: Chicken
  • Immunogen

    Synthetic peptide corresponding to Human Doublecortin aa 350 to the C-terminus conjugated to keyhole limpet haemocyanin.
    Database link: O43602
    (Peptide available as ab19804)

  • Positive control
    • This antibody gave a positive signal in WB on Mouse 0 Day Brain Tissue lysate. ICC/IF: SH-SY5Y cells.
  • General notes
    ab77450 is raised using the same immunogen as ab18723. Batches sold under ab77450 have passed QC testing in Western Blot and ICC/IF. In our hands, this product did not work in IHC-P. For further information please contact our Customer Services team.

Properties

Applications

Our Abpromise guarantee covers the use of ab77450 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).Can be blocked with Human Doublecortin peptide (ab19804).
ICC/IF Use a concentration of 1 - 5 µg/ml.

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

  • ICC/IF image of ab77450 stained SH-SY5Y cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal Goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab77450, 5µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 Goat anti-Rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue).
  • Anti-Doublecortin antibody - Neuronal Marker (ab77450) at 1 µg/ml + Mouse brain tissue lysate - total protein (0 days) (ab7188) at 10 µg

    Secondary
    Rabbit IgG secondary antibody (ab28446) at 1/10000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 40-45 kDa


    Exposure time: 1 minute
  • Anti-Doublecortin antibody - Neuronal Marker (ab77450) at 1 µg/ml + Human Doublecortin full length protein at 0.01 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 40-45 kDa


    Exposure time: 10 seconds

References

This product has been referenced in:
  • Zhao YH  et al. Combined treatment of sodium ferulate, n-butylidenephthalide, and ADSCs rehabilitates neurovascular unit in rats after photothrombotic stroke. J Cell Mol Med 23:126-142 (2019). Read more (PubMed: 30421523) »
  • Feat-Vetel J  et al. Multiple effects of the herbicide glufosinate-ammonium and its main metabolite on neural stem cells from the subventricular zone of newborn mice. Neurotoxicology 69:152-163 (2018). Read more (PubMed: 30312642) »
See all 14 Publications for this product

Customer reviews and Q&As

1-10 of 10 Abreviews or Q&A

Answer

This is a complaint from earlier this year. I had been awaiting some further information, however I have not received it. I am so sorry that I have not followed up with you and the customer meanwhile on this problem. Do you have any updates? How has the customer proceed so far?

Please provide me with the order number as well of the original vial and depending on your answer, I will see what I can do.

I apologize again for not having followed up earlier. Please let me know whether and how you would like to proceed.

Thank you very much for your cooperation.

Read More

Answer

ab77450 is raised using the same immunogen as ab18723 and therefore the products are essentially identical. The only difference between these products is that batches sold under ab77450 have passed QC testing in Western Blot and ICC/IF however, in our hands, this product did not work in IHC-P. ab18723 on the other hand has been used successfully in IHC-P.

I have compared the immunogen of these antibodies with human DCLK1 and DCLK2 and found 31% homology therefore due to this low homology, we would not expect these antibodies to cross-react with DCLK1 and DCLK2.

Read More

Answer

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

As requested, we are pleased to arrange a free of charge replacement. I can confirm that 1 vial each of ab6789, ab72439 and ab77450 have been added to your order number [BO] NL-396-Dt:-11-12-2012. This has Abcam order reference 1224924.

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 vials, 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

Answer

Thank you for your message and for providing this further information.

I am sorry to hear the suggestions made have not improved the results on this occasion.

I appreciate the time the customer has spent on these experiments, and would like to offer a refund or credit note in compensation (providing the items have been purchased in the 6 month guarantee period). I am sorry I notice I do no think we have received the order numbers in this case? To enable us to proceed with this, I would appreciate if you could please confirm the order numbers and date of purchase?

Please note that it is important for us to receive the order number as soon as possible when a complaint is submitted, preferably within the first email. This is vital for our investigation of the case and for our quality monitoring purposes.

Thank you for your continued cooperation. I look forward to hearing from you with the order information and details of how you would like to proceed.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (Dorsal Root Ganglia (DRG) neurons)
Specification
Dorsal Root Ganglia (DRG) neurons
Fixative
Formaldehyde
Permeabilization
Yes - tween 20
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 2% · Temperature: 21°C

Abcam user community

Verified customer

Submitted Nov 05 2012

Question

Dear Technical Team,

Thank you for your mail.

Please find below the reply from the customer on your further queries from him.

Looking forward towards your valued reply on it.

With Best Regards,

Antibody 1 ab72439



1. How has this antibody been stored? -20 degree C after received from vendor

2. Which species is the brain tissue from? rat brain

3. Is the secondary antibody working well with other primary antibodies? What
is the supplier and product number? Yes, supplier Millipore, No:AP307P

4. I can recommend to incubate overnight at4oC. This can often
provide more specific and efficient binding.

5. Has the transfer to the membrane and quality of the sample been assessed
using a loading control? yes



Antibody 2 ab6789

1.
Which species was the sample from? rat brain

2. How has this antibody been stored? -20 degree C after received from vendor

3. Is the primary GAPDH antibody working well with other secondary antibodies? yes, it worked
perfectly with secondary gt anti ms from santacruz biotech.

4. I can recommend to incubate overnight at4oC. This can often
provide more specific and efficient binding.

5. Has the transfer to the membrane and quality of the sample been assessed
using a loading control? yes



Antibody 3 ab77450

1. I can confirm this antibody is not tested in perfusion fixed sections, but
because it has been tested in IHC-P and IHC-Fr which are very similar, I can
provide an exception for the guarantee in this case.

2. I can recommend to try different times for antigen retrieval, such as 3. 5
and10 minutes. This can sometimes require some optimization, and shorter
time can sometimes be better.

3.Triton is a strong detergent and can sometimes disrupt protein
epitopes. I can suggest to try a more gentle detergent such as 0.2% Tween 20.

I will contact again after trying the recommendations. But I think first two antibody need to be replaced. For third one I am changing triton to tween 20 and antigen retrival time. Lets see what happen.

Thanks

Read More
Answer

Thank you for your message and I appreciate the cooperation from the customer. It is regrettable that three antibodies have not worked for them.

I look forward to hearing from you with the results of the suggestions made. I hope these will help improve the results. Please do not hesitate to contact me with further details if there are still any concerns.

Read More

Question

Dear Technical Team,

Hope this e-mail finds you well!

Please find below a complaint from one of our customerto whom we suppliedthese Abcam antibodies.

The informations of technical formare alsomentioned in the trailing mail.

Request you to kindly look into it & adviseus accordingly todo further needful on it.

Looking forward towards your valued reply/ comments on it.

Thanking you in advance.

With Best Regards,


Antibody 1

1) Abcam product code ab72439

2) Abcam order reference number or product batch number : lot:GR3888-7

3) Description of the problem : we have not got any result so far, not even nonspecific or faint band

4) Sample preparation:
Type of sample (whole cell lysates, fraction, recombinant protein…): brain tissue
Lysis buffer : RIPA buffer
Protease inhibitors: protease inhibitor cocktail, sigma
Phosphatase inhibitors: None
Reducing agent : 2 Marcaptoethanol
Boiling for ≥5 min? yes/no: 3 min
Protein loaded ug/lane or cells/lane: 50-100ug/lane
Positive control
Negative control

5) Percentage of gel: 10 and 12%
Type of membrane: PVDF 0.22micron
Protein transfer verified: Ponceau S
Blocking agent and concentration: 3% BSA
Blocking time: 3 Hr to overnight
Blocking temperature: RT for 3 hr, 4 degree C for overnight

6) Primary antibody (If more than one was used, describe in “additional notes”) : Rb anti BDNF
Concentration or dilution: 1:500 to 1:2000
Diluent buffer: 1.5% BSA
Incubation time: overnight or 3 hr.
Incubation temperature: : RT for 3 hr, 4 degree C for overnight

7) Secondary antibody: gt anti rabbit
Species:Goat
Reacts against: Rabbit
Concentration or dilution: 1:5000
Diluent buffer : PBST
Incubation time: 2-3hr
Incubation temperature: RT
Fluorochrome or enzyme conjugate:HRP

8) Washing after primary and secondary antibodies:
Buffer: PBST
Number of washes: 4

9) Detection method: ECL or DAB

10) How many times have you run this staining? 7 times
Do you obtain the same results every time? yes
What steps have you altered to try and optimize the use of this antibody? Altered steps are mentioned as “or” “and”

Document attachment: Attaching images of your blot is strongly recommended and can greatly speed up our investigation of your problem.

Antibody 2


1) Abcam product code ab6789
2) Abcam order reference number or product batch number : lot:GR23382-5

3) Description of the problem : we have not got any result so far, not even nonspecific or faint band

4) Sample preparation:
Type of sample (whole cell lysates, fraction, recombinant protein…): brain tissue
Lysis buffer : RIPA buffer
Protease inhibitors: protease inhibitor cocktail, sigma
Phosphatase inhibitors: None
Reducing agent : 2 Marcaptoethanol
Boiling for ≥5 min? yes/no: 3 min
Protein loaded ug/lane or cells/lane: 50-100ug/lane
Positive control
Negative control

5) Percentage of gel: 10 and 12%
Type of membrane: PVDF 0.22micron
Protein transfer verified: Ponceau S
Blocking agent and concentration: 3% BSA
Blocking time: 3 Hr to overnight
Blocking temperature: RT for 3 hr, 4 degree C for overnight

6) Primary antibody (If more than one was used, describe in “additional notes”) : Ms anti GAPDH
Concentration or dilution: 1:2000
Diluent buffer: 1.5% BSA
Incubation time: overnight or 3 hr.
Incubation temperature: RT for 3 hr, 4 degree C for overnight

7) Secondary antibody: gt anti mouse
Species:Goat
Reacts against: ms
Concentration or dilution: 1:1000, 1:2000 and 1:5000
Diluent buffer : PBST
Incubation time: 2-3hr
Incubation temperature: RT
Fluorochrome or enzyme conjugate:HRP

8) Washing after primary and secondary antibodies:
Buffer: PBST
Number of washes: 4

9) Detection method: ECL or DAB

10) How many times have you run this staining? 5 times
Do you obtain the same results every time? yes
What steps have you altered to try and optimize the use of this antibody? Altered steps are mentioned as “or” “and”

Antibody 3

Order Details
Antibody code * ab77450
Lot number lot:944667
Purchase order number
or preferably Abcam order number


General Information
Antibody storage conditions (temperature/reconstitution etc)
-20 degree C

Description of the problem (high background, low signal, non-specific satining etc.)
No binding at all.

Sample (Species/Tissue/Cell Type/Cell Line etc.) Rat brain section, 30micron


Fixation of sample (Ethanol/Methanol/Acetone/Paraformaldehyde/Other/Duration etc.)
Fixation by 4% paraformaldehyde through transcardial perfusion then kept in 4% paraformaldehyde for 24hr

Antigen retrieval (Enzymatic method, Heat mediated technique etc.)
Heat induced, 95 degree C for 15 min in sodium citrate ph 6.0

Permeabilization step
0.2% triton

Blocking conditions (Buffer/time period, Blocking agent etc.)
5% normal goat serum in PBS with 0.3% triton for 2hr

Primary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step)
Rb anti DCX, 1:200 and 1:500, for 24hr or 48 hr, washed with PBST, abcam

Secondary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step)
Vector elite kit


Detection method:Impact DAB

Positive and negative controls used (please specify)



Optimization attempts (problem solving)
How many times have you tried the IHC? 6times


Have you run a "No Primary" control? yes


Do you obtain the same results every time?yes


What steps have you altered? Alteration mentioned as “or” “and”

Read More
Answer

Thank you for taking the time to complete our questionnaire and contact us. I am sorry to hear you have had difficulty obtaining satisfactory results from this antibody.

The details you have kindly provided will enable us to investigate this case for you and this is also helpful in our records for monitoring of quality.

I would like to reassure you that the antibodies are tested and covered by our 6 month guarantee for the species and applications listed on the datasheets. In the event that a product is not functioning in the applications cited on the product data sheet, we will be pleased to provide a credit note or free of charge replacement.

Reviewing this case, I would like to offer some suggestions to help optimize the results. I would also appreciate if you can confirm some further details:

Antibody 1 ab72439

1. How has this antibody been stored?
2. Which species is the brain tissue from?
3. Is the secondary antibody working well with other primary antibodies? What is the supplier and product number?
4. I can recommend to incubate overnight at4oC. This can often provide more specific and efficient binding.
5. Has the transfer to the membrane and quality of the sample been assessed using a loading control?

Antibody 2 ab6789
1. Which species was the sample from?
2. How has this antibody been stored?
3. Is the primary GAPDH antibody working well with other secondary antibodies?
4. I can recommend to incubate overnight at4oC. This can often provide more specific and efficient binding.
5. Has the transfer to the membrane and quality of the sample been assessed using a loading control?


Antibody 3 ab77450
1. I can confirm this antibody is not tested in perfusion fixed sections, but because it has been tested in IHC-P and IHC-Fr which are very similar, I can provide an exception for the guarantee in this case.
2. I can recommend to try different times for antigen retrieval, such as 3. 5 and10 minutes. This can sometimes require some optimization, and shorter time can sometimes be better.
3.Triton is a strong detergent and can sometimes disrupt protein epitopes. I can suggest to try a more gentle detergent such as 0.2% Tween 20.

I hope this information is helpful, thank you for your cooperation. Should the suggestions not improve the results, please do not hesitate to contact me again with the further requested details.

Read More

Answer

Thank you for contacting Abcam.

I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement.

To check the status of the order please contact our Customer Service team and reference this number.

Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know.

I wish you the best of luck with your research.

Read More

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
Abcam has not validated the combination of species/application used in this Abreview.
Application
Immunohistochemistry (PFA perfusion fixed frozen sections)
Sample
Rat Tissue sections (Brain sections)
Antigen retrieval step
None
Permeabilization
No
Specification
Brain sections
Fixative
Paraformaldehyde

Dr. Sophie Pezet

Verified customer

Submitted May 10 2010

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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