Overview

  • Product name
  • Description
    Rabbit polyclonal to Calpain 5
  • Host species
    Rabbit
  • Specificity
    Does not cross react with the other calpain family members (Calpain 1, Calpain 2, Calpain 3, ncl 3, etc.).
  • Tested applications
    Suitable for: WB, ICC/IFmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide corresponding to Human Calpain 5 aa 1-30 (N terminal).
    (Peptide available as ab41310)

  • General notes
    Diluted antibody can be reused several times if not contaminated; store at 4°C with 0.05% Sodium Azide as preservative.


    ab28280 does not recognize amino-processed Calpain 5, and can be used to discriminate between latent and amino-processed forms. ab28280 will also bind to the non-reduced protein, and thus may have application for immunoprecipitation, immunohistochemistry, and ELISA.

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
  • Storage buffer
    Preservative: 0.05% Sodium azide
    Constituent: 50% Glycerol
  • Concentration information loading...
  • Purity
    Immunogen affinity purified
  • Purification notes
    The antibody has been peptide-affinity purified.
  • Primary antibody notes
    ab28280 does not recognize amino-processed Calpain 5, and can be used to discriminate between latent and amino-processed forms. ab28280 will also bind to the non-reduced protein, and thus may have application for immunoprecipitation, immunohistochemistry, and ELISA.
  • Clonality
    Polyclonal
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab28280 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 1/1000 - 1/5000. Detects a band of approximately 73, 68 kDa.

1/1000 (colorimetric) - 1/5000 (chemiluminescent). A series of further cleaved forms seen when used against the reduced protein. Dilution optimised using Chromogenic detection.

ICC/IF Use a concentration of 1 - 5 µg/ml.

Target

Images

  • ICC/IF image of ab28280 stained HepG2 cells. The cells were 100% methanol fixed (5 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 (ab28280, 1µg/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) at a concentration of 1.43µM.

References

This product has been referenced in:
  • Schaefer KA  et al. Calpain-5 Expression in the Retina Localizes to Photoreceptor Synapses. Invest Ophthalmol Vis Sci 57:2509-21 (2016). Read more (PubMed: 27152965) »
  • Singh R  et al. Calpain 5 is highly expressed in the central nervous system (CNS), carries dual nuclear localization signals, and is associated with nuclear promyelocytic leukemia protein bodies. J Biol Chem 289:19383-94 (2014). WB ; Human, Rat . Read more (PubMed: 24838245) »
See all 2 Publications for this product

Customer reviews and Q&As

1-2 of 2 Abreviews or Q&A

Answer

Thank you for your reply.

As this is a rabbit polyclonal antibody, we would expect some lot to lot variation in optimal antibody concentrations. What are the conditions of your current WB protocol? I would like to see if there are any suggestions that can be made to improve your experience with this antibody.

Currently, we only have the lot you are already using, so a replacement from a different lot is not possible. If I am unable to make any protocol suggestions that improve your results with this antibody, I would be happy to offer another vial of this antibody from the current lot or a credit.

I look forward to your reply so that I may assist you further. Please do not hesitate to contact us if you have any additional questions.

Read More

Answer

Thank you for contacting Abcam and I am very sorry about the delay in getting you this information.
Answers to your questions:
1 - We used lysates from A2058 cells, and from RD cells in Western blot to look for Calpain-5 protein. We don't see strong signals in most cell lines, as there is little calpain-5 there.
2 - Tissue lysates from colon, kidney, liver and trachea would make good positive control samples. I would recommend either ab29889 (human) or ab7265 (mouse) as good positive control samples if you wish to purchase one.
3 - I think the band you are seeing in your western blot is fine and it is running at the correct size (anything within about 5kDa of the predicted molecular weight is OK).
4 - You are correct that this antibody does not recognize amino-processed Calpain 5 and so you should not be seeing a band at 63kDa, and since you are not seeing this bad, I do not think it is a problem. I will have our data sheets team look at this and see if the reference to the 63kDa band should be removed from the website.
Once again please accept my apologies for the long delay in responding to you, this was due to us waiting on a response from the lab.
If there is anything else I can help you with, please let me know.

Read More

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

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