Overview

  • Product name
  • Description
    Rabbit polyclonal to KCC2
  • Host species
    Rabbit
  • Tested applications
    Suitable for: IHC-Fr, WB, ICC/IFmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Chicken, Dog
    Predicted to work with: a wide range of other species
  • Immunogen

    N-terminal His-tag fusion protein corresponding to residues 932-1043 of rat K+/Cl- Cotransporter (KCC2).

  • Positive control
    • Rat brain membrane preparation; Madin Darby Canine Kidney (MDCK) cells

Properties

Applications

Our Abpromise guarantee covers the use of ab49917 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 Use at an assay dependent concentration.
WB 1/1000. Predicted molecular weight: 123 kDa.
ICC/IF 1/200 - 1/500. PubMed: 23440186

Target

  • Function
    Mediates electroneutral potassium-chloride cotransport in mature neurons. Transport occurs under isotonic conditions, but is activated 20-fold by cell swelling. Important for Cl(-) homeostasis in neurons.
  • Tissue specificity
    Brain specific. Detected in neuronal cells.
  • Sequence similarities
    Belongs to the SLC12A transporter family.
  • Cellular localization
    Membrane.
  • Information by UniProt
  • Database links
  • Alternative names
    • Electroneutral potassium chloride cotransporter 2 antibody
    • Electroneutral potassium-chloride cotransporter 2 antibody
    • Erythroid K Cl cotransporter 2 antibody
    • Furosemide sensitive K Cl cotransporter antibody
    • hKCC2 antibody
    • K-Cl cotransporter 2 antibody
    • KCC 2 antibody
    • KCC2 antibody
    • KIAA1176 antibody
    • Neuronal K Cl cotransporter antibody
    • Neuronal K-Cl cotransporter antibody
    • Potassium Chloride Cotransporter antibody
    • Potassium chloride transporter 5 antibody
    • rKCC2 antibody
    • S12A5 antibody
    • S12A5_HUMAN antibody
    • SLC12A5 antibody
    • Solute carrier family 12 (potassium chloride transporter) member 5 antibody
    • Solute carrier family 12 member 5 antibody
    see all

Images

  • All lanes : Anti-KCC2 antibody (ab49917) at 1/2000 dilution

    Lane 1 : Whole tissue lysate prepared from chicken auditory brainstem/tectum.
    Lane 2 : Whole tissue lysate prepared from chicken auditory brainstem/tectum, preabsorped with immunogenic peptide.

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : HRP-conjugated goat anti-rabbit at 1/5000 dilution

    Developed using the ECL technique.

    Predicted band size: 123 kDa


    Exposure time: 10 minutes

    See Abreview

  • ab49917 staining KCC2 in rat spinal cord tissue sections by Immunohistochemistry (frozen sections). Tissue was fixed with paraformaldehyde and permeabilized using 0.1% Triton-X100. Samples were then blocked with 2% serum for 2 hours at 26°C followed by incubation with the primary antibody, at a 1/4000 dilution, for 19 hours at 4°C. The secondary antibody used was a goat anti-rabbit IgG conjugated to Alexa Fluor® 546 (red) used at a 1/4000 dilution.

    See Abreview

References

This product has been referenced in:
  • Miceli S  et al. Reduced Inhibition within Layer IV of Sert Knockout Rat Barrel Cortex is Associated with Faster Sensory Integration. Cereb Cortex 27:933-949 (2017). Read more (PubMed: 28158484) »
  • Hu D  et al. Bumetanide treatment during early development rescues maternal separation-induced susceptibility to stress. Sci Rep 7:11878 (2017). Read more (PubMed: 28928398) »
See all 5 Publications for this product

Customer reviews and Q&As

1-9 of 9 Abreviews or Q&A

Answer



The image on the datasheet is from a abreview that was written by a customer.

It shows that with the blocking peptide that blocks all specific binding both bands disappear. This means that both bands depict the KCC2 protein.

This is a multipass membrane protein, and these proteins have the unfortunate tendency to aggregate during boiling periods with high temperatures and beta- mercaptoethanol due to their hydrophobic nature leading to extra bands that are usually running higher than expected.

Also KCC2 can be glycosylated. Glycosylation will increase the molecular weight of a protein and therefore if the protein is in the cell in both states (glycosylated and un-glycosulated) two band will be shown on the WB.

In summary, we are not concerned that this antibody is binding un-specifically, but are confident that all versions of the KCC2 protein are recognized.

Read More

Answer

Gracias por contactarnos de nuevo.

Siento mucho los problemas ocasionados por estos productos que no han funcionado como se esperaba.

Para poder dar el soporte oportuno de los mismos y entender por qué han dado ese resultado, te rogaría que por favor me ampliaras la siguiente información:

- ¿Cuál es el numero de pedido de los anticuerpos?

- ¿Cuáles son los lotes que funcionaron en el pasado de ab59791 y ab49917?

- ¿En qué condiciones se ha llevado a cabo el lisado de tejidos (buffer, inhibidores, agente reductor, etc...)?

- ¿Se ha usado un control positivo en paralelo con las muestras?

- ¿Se ha comprobado el buen estado del secundario utilizado, así como del sustrato DAB? Es decir, ¿se han usado con otros sistemas obteniendo un resultado positivo?

- Recomendamos llevar a cabo una incubación de anticuerpo de 12-16 horas para asegurar que el anticuerpo alcanza el epítopo.

Por otro lado, quisiera asegurar al cliente que los anticuerpos no han podido afectarse por el hecho de llegarles a 8C. El envío del producto en los bloques azules de hielo es solo una medida de precaución, y ésta no es la primera vez que un cliente nos reporta la llegada de los productos a temperatura ambiente, sin que posteriormente haya sufrido algún problema con el mismo.

En nuestros laboratorios hemos hecho estudios de estabilidad de los anticuerpos y proteínas con la temperatura, demostrando que son estables hasta a 45C durante una semana sin que pueda impactar en su actividad.

El lunes estaré fuera de la oficina y no podre contestar los correos, pero a partir del martes día 2 podre segur adelante con esta reclamación. Espero tu respuesta entonces.

Que pases un buen día.

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Answer

Thank you for the protocol details. I think the protocol is generally good, but I am concerned about the length of the post-fixation, 7 days. The perfusion, followed by 4 hours at most for post-fixation, should be sufficient for preservation of the brain morphology. The fixation for 7 days may interfere with antibody:antigen recognition.

You may find that an antigen retrieval step is necessary, but it can be difficult to apply to frozen sections of this thickness, without damaging them. I am attaching an article that discusses heat-mediated antigen retrieval for frozen tissue.

You may also find that a detergent permeabilization before applying the goat serum, for instance 0.1% Triton X-100 in PBS for 20 minutes, will help develop the stain.

You did not mention the concentration of the antibody ab49917 that you used (unless I overlooked it) but I recommend trying a dilution series: 1/1000, 1/500, and 1/250.

I hope these suggestions help. If not, or if you have any questions, please contact me.

Read More

Answer

Thank you for contacting us.

Are you trying to detect NKCC2 or KCC2?

NKCC2 (SLC12a1) moves one sodium ion, one potassium ion and 2 chloride ions together. KCC2 (SLC12a5) moves two potassiums with 2 chlorides. Because of the differences in electrochemical driving force between K (outward) and Na (inward), KCC2 generally carries solute from inside the cell to outside, whereas NKCC2 generally carries solute into the cell. NKCC2 is basically specific to a single cell type in kidney. KCC2 is found in many cells. You mentioned NKCC2 not KCC2, although some people do use that acronym for KCC2 as well.

The target of antibodies ab49117 and ab134300 is described at the following URL:

http://www.uniprot.org/uniprot/Q63633

Read More

Answer

Thank you very much for your interest in our antibodies. For c-Fos, I would recommend either ab7963 or ab87655.  Both of these antibodies will be guaranteed for use in IHC on mouse samples.  For Aromatase, ab18995 and ab35604 will be guaranteed for your purposes.  To our knowledge, none of our KCC2 antibodies have been tested in IHC on mouse samples. The product that would most likely be suitable is ab49917 which is known to react with rat for use in IHC-Fr and ICC/IF.  If you would like to test ab49917 for use on mouse samples, I can offer you a discount off a future purchase. All you would need to do is buy ab49917 now, test it in mouse and submit feedback to us in the form of an Abreview. It doesn’t matter whether the Abreview is positive or negative, we would just really like to receive your feedback. The discount would be to the value of 1 free primary antibody. If you are interested in this offer, please follow these steps: 1. Reply to this e-mail to let me know that you would like to proceed and test ab49917 in mouse. I will then send a discount code. This code must be issued before purchasing ab49917 so please wait for my reply before ordering. 2. Purchase ab49917 either by phone, fax, or online (www.abcam.com). 3. Test it in mouse. 4. Let us know the results, positive or negative, using our Abreview system (this will take about 10 minutes and images are great if you have them!). To find out how to submit an Abreview, please visit: https://www.abcam.com/abreviews. 5. After the review is submitted to us, the discount code becomes active. Simply place your new order by phone, fax, or on the web and mention the discount code. The discount can be redeemed for any primary antibody ordered and the discount code is valid for 4 months after issue. We are always pleased to obtain feedback about our products and any information is greatly appreciated! Even if ab49917 turns out to be unsuitable for mouse, you will still receive the discount on your next purchase after your Abreview has been submitted. Please let me know if you have any questions about this offer and I would be happy to help you further. The Terms and Conditions of this offer can be found at: www.abcam.com/collaborationdiscount.  

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Chicken Tissue lysate - whole (Auditory brainstem/Tectum)
Loading amount
20 µg
Specification
Auditory brainstem/Tectum
Gel Running Conditions
Reduced Denaturing (7,5% Acrylamid)
Blocking step
The Blocking Solution (Candor Bioscience) as blocking agent for 12 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 4°C

Ms. Stefanie Kurth

Verified customer

Submitted Feb 10 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Frozen sections)
Sample
Rat Tissue sections (spinal cord tissue)
Specification
spinal cord tissue
Fixative
Paraformaldehyde
Permeabilization
Yes - 0.1% Triton-X100
Blocking step
Serum as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 26°C

Abcam user community

Verified customer

Submitted Sep 23 2010

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Chicken Tissue lysate - other (Brain tissue)
Loading amount
40 µg
Specification
Brain tissue
Gel Running Conditions
Reduced Denaturing (10%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5%

Ms. Yuhua Liu

Verified customer

Submitted Feb 09 2010

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Chicken Tissue lysate - other (brain tissue)
Loading amount
40 µg
Specification
brain tissue
Gel Running Conditions
Non-reduced Non-Denaturing (Native) (10%, seperate gel)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5%

Ms. Wei Shi

Verified customer

Submitted Aug 25 2009

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

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