Overview

Description

  • Nature
    Recombinant
  • Source
    Escherichia coli
  • Amino Acid Sequence
    • Accession
    • Species
      Human

Specifications

Our Abpromise guarantee covers the use of ab52101 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

  • Biological activity

    SPECIFIC ACTIVITY: 300,000-400,000 units/mg UNIT DEFINITION: One unit of the recombinant caspase-3 is the enzyme activity that cleaves 1 nmol of the caspase substrate DEVD-pNA (pNA: pnitroanaline) per hour at 37oC in a reaction solution containing 50 mM Hepes, pH 7.2, 50 mM NaCl, 0.1% Chaps, 10 mM EDTA, 5% Glycerol, and 10 mM DTT.

  • Applications

    Functional Studies

  • Purity
    > 95 % SDS-PAGE.

  • Form
    Lyophilised
  • Additional notes
    The lyophilized caspase-3 is stable for 1 year at –70oC The recombinant active human Caspase 3 spontaneously undergoes autoprocessing to yield subunits characteristic of the native enzyme.


    Active Caspase 3 is useful in studying enzyme regulation, determining target substrates, screening caspase inhibitors, or as a positive control in caspase activity assays. The active Caspase 3 preferentially cleaves Caspase 3 substrates e.g. DEVD-AFC or DEVD-pNA.

  • Concentration information loading...

Preparation and Storage

  • Stability and Storage

    Shipped at 4°C. Upon delivery aliquot. Store at -80°C. Avoid freeze / thaw cycle.

    Constituents: PBS, 15% Glycerol

    This product is an active protein and may elicit a biological response in vivo, handle with caution.

  • Reconstitution
    Reconstitute to 1 unit per µl in water.

General Info

  • Alternative names
    • Apopain
    • CASP 3
    • CASP-3
    • CASP3
    • CASP3_HUMAN
    • Caspase 3
    • Caspase-3 subunit p12
    • cleaved caspase 3
    • CPP 32
    • CPP-32
    • CPP32B
    • Cysteine protease CPP32
    • PARP cleavage protease
    • Protein Yama
    • SCA-1
    • SCA1
    • SREBP cleavage activity 1
    • Yama
    see all
  • Function
    Involved in the activation cascade of caspases responsible for apoptosis execution. At the onset of apoptosis it proteolytically cleaves poly(ADP-ribose) polymerase (PARP) at a '216-Asp-
    -Gly-217' bond. Cleaves and activates sterol regulatory element binding proteins (SREBPs) between the basic helix-loop-helix leucine zipper domain and the membrane attachment domain. Cleaves and activates caspase-6, -7 and -9. Involved in the cleavage of huntingtin. Triggers cell adhesion in sympathetic neurons through RET cleavage.
  • Tissue specificity
    Highly expressed in lung, spleen, heart, liver and kidney. Moderate levels in brain and skeletal muscle, and low in testis. Also found in many cell lines, highest expression in cells of the immune system.
  • Sequence similarities
    Belongs to the peptidase C14A family.
  • Post-translational
    modifications
    Cleavage by granzyme B, caspase-6, caspase-8 and caspase-10 generates the two active subunits. Additional processing of the propeptides is likely due to the autocatalytic activity of the activated protease. Active heterodimers between the small subunit of caspase-7 protease and the large subunit of caspase-3 also occur and vice versa.
    S-nitrosylated on its catalytic site cysteine in unstimulated human cell lines and denitrosylated upon activation of the Fas apoptotic pathway, associated with an increase in intracellular caspase activity. Fas therefore activates caspase-3 not only by inducing the cleavage of the caspase zymogen to its active subunits, but also by stimulating the denitrosylation of its active site thiol.
  • Cellular localization
    Cytoplasm.
  • Information by UniProt

References

This product has been referenced in:
  • Yozwiak CE  et al. Toward a Microparticle-Based System for Pooled Assays of Small Molecules in Cellular Contexts. ACS Chem Biol 13:761-771 (2018). Read more (PubMed: 29365249) »
  • Cao L  et al. Pseudo-phosphorylation at AT8 epitopes regulates the tau truncation at aspartate 421. Exp Cell Res 370:103-115 (2018). Read more (PubMed: 29908160) »
See all 2 Publications for this product

Customer reviews and Q&As

1-10 of 11 Abreviews or Q&A

Question
Answer

The active Caspase 3 protein (Active), ab52101, is provided at the equivalent of 100 units. There are between 300,000-400,000 units/mg. Therefore a total of between 0.25-0.33 ug of protein is provide per vial.

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Question
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 our Anti-Caspase 3 antibody [4H334] (ab17819) and active Caspase 3 protein (Active) (ab52101) .

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 ab17819 is tested and covered by our 6 month guarantee for use in WB, ICC, IP and human samples. 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 clarify the results from ab52101 and ab17819. I would also appreciate if you can confirm some further details:

1. Could you please confirm the order details and date of purchase.

2. Could you confirm if active Caspase 3 protein (Active) (ab52101) was constituted in PBS containing 15% glycerol as stated on the datasheet. Could you also confirm how the protein was stored after constitution please?

3. We would recommend to load a sample of the constituted protein onto a SDS Gel and perform a comassie staining. That would confirm the sample contains the protein and also if there is some degradation of the protein. We would recommend to load an amount of 10 to 20 ng for the SDS PAGE. Furthermore we would recommend the same amount of protein for the WB.

4. Could you please confirm what dilutions were used for the primary antibody and what was used as a diluent. Is the primary antibody working with other samples? It would be beneficial to include another sample control for the WB, if not already done.

5. Could you please confirm what secondary antibody was used for the WB and weather it is working well with other primary antibodies? A control for the secondary antibody is beneficial to make sure it is working well.

6. Could you confirm what washing and blocking buffers were used and how the washing was performed. (What reagents? How many wash steps? How long were the steps?)

7. It would be beneficial to confirm the transfer of proteins to the membrane for the WB, if that has not already been done.

8. Could you please give a description about the functional assay that was performed with the active Caspase 3 protein (Active) (ab52101)

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.

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Question
Answer

Thank you again for your call.

Unfortunately, the total concentration of this protein is proprietary information. We sell these by the number of units of enzyme activity totally present in each vial. Therefore in case of ab52101, there would be 100 units of the active human Caspase-3.

Hope this information is still helpful for you.

Read More

Question
Answer

Thank you for contacting us.

I would suggest to use ab17819 Anti-Caspase 3 antibody [4H334]

https://www.abcam.com/index.html?datasheet=17819 (or use the following: https://www.abcam.com/index.html?datasheet=17819).

Even if the blot we have on the website is not very clear, the references we have for this antibody clearly show that this antibody detects both p32 and p17.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

Use our products? Submit an Abreview. Earn rewards!
https://www.abcam.com/abreviews

Read More

Answer

Thank you for providing that information and sorry for the delay in getting back to you.
The information provided has been very useful and I will look into whether these antibodies would be likely to detect the protein and what may be contributing to the problems observed.
In order to understand the problems encountered further, would you be able to give me an outline of the dilutions of the antibodies and proteins used, as well as any blocking steps and explain a little about how a positive signal is deduced (i.e. is a secondary antibody with a conjugate used)?
Also, which antibody combinations were used and achieved a good signal with the positive control and negative control canine samples?
I am sorry but I will be away from the office on Friday but I will get back to you with further information on Monday.
I look forward to receiving your reply.

Read More

Answer

Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products.

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 goodwill free of charge replacement with the order number 1234567.

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.

Please always store our products as indicated on the datasheet, as otherwise we cannot guaranty its stability and the outcome of your experiments.

I wish you the best of luck with your research.

Read More

Answer

You have received the recombinant full length protein: processed form of Caspase 3 consists of large (17kDa) and small (12kDa) subunits which associate to form an active enzyme.

Read More

Answer

Thank you for calling Abcam earlier today.

I am sorry about the issues you have been having with the Caspase 3 protein. As we discussed, I have requested that you be sent (via Cedarlane) a free of charge replacement for the protein.

If there is anything else I can help you with, please let me know.

Read More

Answer

Thank you for contacting Abcam.

I have emailed the lab about your enquiry and below is their response:

This product is not sold by concentration. It is sold by Units. The recombinant active human caspase-3 expressed in E coli spontaneously undergoes autoprocessing to yield subunits characteristic of the native enzyme (Full length gene Accession No. NP_004337) . The active caspase-3 preferentially cleaves caspase-3 substrates (e.g., DEVD-AFC or DEVD-pNA) and is routinely tested for its ability to enzymatically cleave these two substrates Ac-DEVD-pNA or Ac-DEVD-AFC .

From this data it is aliquoted and sold by Units.

Please let me know if ther eis anything else I can help you with.

Read More

Answer

Thank you for contacting Abcam.

For ab39901, Caspase 1, we recommend using 1 unit/assay for analyzing caspase activity and this protein has a biological activity : >5000 units/mg and it is supplied lyophilized and it should be reconstituted to 1 unit per µl in PBS containing 15% glycerol (and as it supplied as 25 units you would end up with 25ul of solution).

For ab52101, Caspase 3, one unit of the recombinant caspase-3 is the enzyme activity that cleaves 1 nmol of the caspase substrate DEVD-pNA (pNA: pnitroanaline) per hour at 37oC in a reaction solution containing 50 mM Hepes, pH 7.2, 50 mM NaCl, 0.1% Chaps, 10 mM EDTA, 5% Glycerol, and 10 mM DTT. You are supplied with 100 units and again it is lyophilized on arrival an you should reconstitute to 1 unit per µl in PBS containing 15% glycerol (therefore you would have 100ul of solution).

The above are recommended starting dilutions, however the optimal dilutions/concentrations should be determined by the end user.

Please let me know if there is anything else I can help you with.

Read More

1-10 of 11 Abreviews or Q&A

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

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