Overview

  • Product name
    Anti-Nestin antibody [Rat 401]
    See all Nestin primary antibodies
  • Description
    Mouse monoclonal [Rat 401] to Nestin
  • Host species
    Mouse
  • Specificity
    The clone number has been updated from (2Q178) to (Rat 401) both clone numbers name the same antibody clone.
  • Tested applications
    Suitable for: ICC/IF, Flow Cyt, IHC-FoFr, IHC-P, IHC-Fr, WBmore details
  • Species reactivity
    Reacts with: Mouse, Rat
    Does not react with: Sheep, Cat, Human, Monkey
  • Immunogen

    Full length native protein (purified) corresponding to Rat Nestin.

  • Positive control
    • Mouse or rat embryonic brain tissue. ICC/IF: Mouse embryonic stem cells.

Properties

Applications

Our Abpromise guarantee covers the use of ab6142 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
ICC/IF 1/20 - 1/200. Cells fixed with 4% paraformaldehyde buffered with 50mM sodium borate at pH 9.5.
Flow Cyt Use at an assay dependent concentration. PubMed: 21092738

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

IHC-FoFr Use at an assay dependent concentration. PubMed: 18562299
IHC-P Use a concentration of 0.1 - 1 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. Tissue fixed with 4% paraformaldehyde at pH 7.4 for light microscopy.
IHC-Fr Use a concentration of 0.1 - 1 µg/ml. Tissue fixed with 4% paraformaldehyde at pH 7.4 for light microscopy.
WB Use at an assay dependent concentration. Predicted molecular weight: 200 kDa. Block with milk or BSA but do not dilute primary antibody in buffer containing milk.

Target

  • Function
    Required for brain and eye development. Promotes the disassembly of phosphorylated vimentin intermediate filaments (IF) during mitosis and may play a role in the trafficking and distribution of IF proteins and other cellular factors to daughter cells during progenitor cell division. Required for survival, renewal and mitogen-stimulated proliferation of neural progenitor cells.
  • Tissue specificity
    CNS stem cells.
  • Sequence similarities
    Belongs to the intermediate filament family.
  • Developmental stage
    Upon terminal neural differentiation, nestin is down-regulated and replaced by neurofilaments.
  • Post-translational
    modifications
    Constitutively phosphorylated. This increases during mitosis when the cytoplasmic intermediate filament network is reorganized.
  • Information by UniProt
  • Database links
  • Alternative names
    • ESTM 46 antibody
    • FLJ 21841 antibody
    • FLJ21841 antibody
    • Intermediate filament protein antibody
    • Nbla00170 antibody
    • nes antibody
    • NEST_HUMAN antibody
    • Nestin antibody
    see all

Images

  • Mesencephalic neural stem cells were washed three times in PBS, fixed in 4% paraformaldehyde for 10 min at room temperature (RT), and permeabilized with PBS plus 0.25% Triton X-100 (PBST) for 10 min at RT (or 15 min at RT if the target antigen was a nucleoprotein). After three washes in PBS, the cells were blocked with 1% BSA for 30 min at RT and incubated with ab6142 staining Nestin (Red) at 1/1000 dilution.

    Immunolabeling was visualized by incubation (1 h at RT) with rhodamine-conjugated Affinipure goat anti-mouse or anti-rabbit IgG (1∶100). After three washes in PBS and two in deionized water, immunolabeled cells were counterstained with 50 µl DAPI at 37°C for 10 min and washed in sequence with methanol and ethanol. Cells were observed under a fluorescence microscope.

    Fluorescence photomicrographs show that all neurospheres were immunoreactive for the NSCs marker nestin. The neurospheres of peGFPN1-GDNF-transfected mNSCs (termed GDNF-mNSCs) were strong immunopositive for GDNF compare to mNSCs transfected with peGFPN1. Green: FITC, Red: Nestin. Scale bar = 50 µm.

  • ab6142 at 1/500 dilution staining Nestin in rat brain tissue (green) by immunohistochemistry (frozen sections). Sections were methanol fixed, permeabilized in 0.1% Triton X-100 prior to blocking in 2.5% BSA for 16 hours at 4°C and then incubated with ab6142, for 1 hour at 37°C. Alexa fluor® 680 goat polyclonal to mouse Ig, diluted 1/1000 was used as the secondary antibody.

    See Abreview

  • ab6142 staining mouse embryonic stem cells by immunocytochemistry/ immunofluorescence. Cells were PFA fixed and permeabilized in 0.4% Triton X-100 prior to blocking in 5% serum for 1 hour at 25°C. The primary antibody was diluted 1/500 and incubated with the sample for 20 hours at 4°C. Alexa fluor® 568 goat polyclonal to mouse Ig, diluted 1/400, was used as the secondary antibody.

    See Abreview

  • ab6142 staining Nestin in mouse brain tissue sections by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with acetone, permeabilized with Triton X-100 in PBS and blocked with 5% BSA for 60 minutes at room temperature. Samples were incubated with primary antibody (1/1000 in PBST) for 12 hours at 4°C. An Alexa Fluor® 488-conjugated goat anti-mouse IgG monoclonal was used as the secondary antibody at a dilution of 1/200.

    See Abreview

  • ab6142 at 1/200 dilution staining Nestin in mouse Cor1 neural stem cells by Immunocytochemistry/ Immunofluorescence. Cells were fixed using 4%PFA for 15mins at RT. After x3 PBS rinses over 15mins, cells were blocked/permeabilised using primary Antibody dilution buffer for 15mins befpre application of primary Ab diluted in same buffer. Secondary conjugate was diluted in same buffer, with addition of 1/10K Hoechst, to stain nuclei. After 60mins cells were rinsed x3 over 15mins before coverslipping using Mowiol containing DABCO and azide.

    See Abreview

  • ab6142 staining adult mouse brain tissue section by Immunohistochemistry (Formalin/PFA-fixed, paraffin embedded sections). Tissue underwent fixation in paraformaldehyde, heat mediated antigen retrieval in Sodium Citrate, permeabilization in 1% Triton buffer and blocking in 10% serum for 1 hour at 25°C. The primary antibody, diluted 1/200 (PBS, 2% Donkey serum, 0.2% Triton) for 16 hours at 4°C.  An Alexa Fluor® 488 conjugated donkey polyclonal to mouse Ig, diluted 1/500 was used as the secondary.

    See Abreview

References

This product has been referenced in:
  • Gholinejad M  et al. Adenosine decreases oxidative stress and protects H2O2-treated neural stem cells against apoptosis through decreasing Mst1 expression. Biomed Rep 8:439-446 (2018). Read more (PubMed: 29732147) »
  • Lin GQ  et al. Transplanted human neural precursor cells integrate into the host neural circuit and ameliorate neurological deficits in a mouse model of traumatic brain injury. Neurosci Lett 674:11-17 (2018). IHC . Read more (PubMed: 29501684) »
See all 83 Publications for this product

Customer reviews and Q&As

1-10 of 34 Abreviews or Q&A

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Frozen sections)
Sample
Rat Tissue sections (Brain)
Permeabilization
Yes - Triton-X 100
Specification
Brain
Blocking step
Donkey serum as blocking agent for 10 minute(s) · Concentration: 2% · Temperature: 23°C
Fixative
Paraformaldehyde

Alyssa Cortella

Verified customer

Submitted Aug 24 2018

Application
Immunohistochemistry (PFA perfusion fixed frozen sections)
Sample
Mouse Tissue sections (Brain)
Antigen retrieval step
None
Permeabilization
Yes - TritonX-100
Specification
Brain
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C
Fixative
Paraformaldehyde

Kyungjoo Seong

Verified customer

Submitted Nov 02 2016

Application
Immunocytochemistry
Sample
Mouse Cell (Mouse neural stem cell)
Permeabilization
Yes - tritonx100
Specification
Mouse neural stem cell
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C
Fixative
Paraformaldehyde

Kyungjoo Seong

Verified customer

Submitted Nov 02 2016

Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (brain)
Permeabilization
Yes - Triton
Specification
brain
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C
Fixative
Acetone

Abcam user community

Verified customer

Submitted Feb 09 2016

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Frozen sections)
Sample
Rat Tissue sections (adult rat heart tissue)
Permeabilization
Yes - 0.1% Triton
Specification
adult rat heart tissue
Blocking step
Serum as blocking agent for 20 minute(s) · Concentration: 2% · Temperature: 20°C
Fixative
Paraformaldehyde

Abcam user community

Verified customer

Submitted Jan 28 2016

Application
Western blot
Loading amount
400000 cells
Gel Running Conditions
Reduced Denaturing (12%)
Sample
Mouse Cell lysate - whole cell (Mouse Neuroblastoma (Neuro2A))
Specification
Mouse Neuroblastoma (Neuro2A)
Treatment
Serum deprivation (0.1%) for 24 hrs
Blocking step
BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 28°C

Abcam user community

Verified customer

Submitted Mar 25 2015

Answer

Thank you for contacting us.
I can confirm the isotype controls suggested are suitable for the antibodies requested.
The difference in price between ab91353 and ab81197 lies in the volume difference (the volume of ab91353 doubles ab81197’s volume) and the manufacturing and purification process often make price vary between what seems to be similar products.
I hope this helps. Please do not hesitate to contact us if you require further assistance.

Read More
Abcam has not validated the combination of species/application used in this Abreview.
Application
Western blot
Sample
Apteronotus leptorhynchus Tissue lysate - whole (Brain)
Gel Running Conditions
Reduced Denaturing (4-15%)
Loading amount
50 µg
Specification
Brain
Blocking step
Milk as blocking agent for 14 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C

Abcam user community

Verified customer

Submitted Nov 27 2012

Answer

Vielen Dank für Ihre Anfrage.

Ich stimme Ihnen zu, dass man Mausantikörper auf Mausgeweben generell nicht empfiehlt, da der sekundaere Antikörper auch die endogenes Ig erkennt und somit ein starker Hintergrund auftritt.

Leider kann ich Ihnen nicht sagen,ob die Kunden, die die Abreviews eingeschickt haben, irgendwelche Hilfsmittel eingesetzt haben.

Allerdings sind die IHC Experimente mit Gehirnproben erfolgt (und ich nehme an mit Perfusionstechnik, was alles Blut ausschwämmen würde)und im Gehirn direktsollten keine endogenen Ig vorhanden sein.

Falls Sie generell einmal einen Mausantikörper auf Mausproben verwenden wollen oder müssen, kann ich Ihnen unser Mouse on Mouse (M.O.M) Polymer IHC Kit(ab127055) empfehlen:

https://www.abcam.com/index.html?datasheet=127055 (or use the following: https://www.abcam.com/index.html?datasheet=127055).

Ich hoffe diese Information ist hilfreich und wünsche Ihnen ein schönes Wochenende.

Read More

Answer

Gracias por contactar con nosotros.

Lamento mucho las molestias ocasionadas.

Sin embargo te rogaría que por favor me confirmes si habéis centrifugado el vial antes de abrirlo, y comprobado que el líquido no se quedó adherido a las paredes y la tapa del mismo. Este anticuerpo está muy concentrado, y la cantidad recibida es muy pequeña, por lo que estas comprobaciones previas a la apertura del vial son necesarias.

En ocasiones es cierto que por error se mandan viales con menos volumen del indicado, pero es muy extraño que el vial estuviera completamente vacío.

Agradezco mucho tu cooperación en este asunto, y quedo a la espera de tu respuesta para ayudaros a solucionar este inconveniente.

Read More

1-10 of 34 Abreviews or Q&A

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