Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-Calpain 1 antibody [EPR3319] (ab108400)

Reviews (1)Q&A (1)References (10)
Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
  • Flow Cytometry (Intracellular) - Anti-Calpain 1 antibody [EPR3319] (ab108400)
  • Immunocytochemistry/ Immunofluorescence - Anti-Calpain 1 antibody [EPR3319] (ab108400)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calpain 1 antibody [EPR3319] (ab108400)
  • Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
  • Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
  • Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
  • Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
  • Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calpain 1 antibody [EPR3319] (ab108400)
  • Immunocytochemistry/ Immunofluorescence - Anti-Calpain 1 antibody [EPR3319] (ab108400)
  • Anti-Calpain 1 antibody [EPR3319] (ab108400)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR3319] to Calpain 1
  • Suitable for: WB, IHC-P, ICC/IF, Flow Cyt (Intra)
  • Knockout validated
  • Reacts with: Mouse, Rat, Human

Conjugates logo Related conjugates and formulations

Carrier Free

Overview

  • Product name

    Anti-Calpain 1 antibody [EPR3319]
    See all Calpain 1 primary antibodies

  • Description

    Rabbit monoclonal [EPR3319] to Calpain 1

  • Host species

    Rabbit

  • Specificity

    ab108400 recognises the large subunit of Calpain

  • Tested applications

    Suitable for: WB, IHC-P, ICC/IF, Flow Cyt (Intra)more details

  • Species reactivity

    Reacts with: Mouse, Rat, Human

  • Immunogen

    Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • Human fetal kidney, U937, A431, (D) 293T, and T47D cell lysates; Human kidney tissue; HeLa cells.

  • General notes

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

Properties

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab108400 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. Predicted molecular weight: 82 kDa.

For unpurified use at 1/1000 - 1/10000.
IHC-P
1/200. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

For unpurified use at 1/500 - 1/1000.
ICC/IF
1/50.

For unpurified use at 1/250 - 1/500.
Flow Cyt (Intra)
1/380.

For unpurified use at 1/100 - 1/500.  ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.
Notes
WB
1/1000 - 1/5000. Predicted molecular weight: 82 kDa.

For unpurified use at 1/1000 - 1/10000.
IHC-P
1/200. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

For unpurified use at 1/500 - 1/1000.
ICC/IF
1/50.

For unpurified use at 1/250 - 1/500.
Flow Cyt (Intra)
1/380.

For unpurified use at 1/100 - 1/500.  ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

Target

  • Function

    Calcium-regulated non-lysosomal thiol-protease which catalyze limited proteolysis of substrates involved in cytoskeletal remodeling and signal transduction.

  • Tissue specificity

    Ubiquitous.

  • Sequence similarities

    Belongs to the peptidase C2 family.
    Contains 1 calpain catalytic domain.
    Contains 4 EF-hand domains.

  • Cellular localization

    Cytoplasm. Cell membrane. Translocates to the plasma membrane upon Ca(2+) binding.
  • Information by UniProt

  • Database links

    see all

  • Alternative names

    • Ca2 activated neutral protease antibody
    • Calcium activated neutral proteinase antibody
    • Calcium activated neutral proteinase small subunit antibody
    • Calcium dependent protease small subunit 1 antibody
    • Calcium dependent protease small subunit antibody
    • Calcium-activated neutral proteinase 1 antibody
    • Calpain 1 antibody
    • Calpain 1 large subunit antibody
    • Calpain 1, (mu/I) large subunit antibody
    • Calpain mu type antibody
    • Calpain mu-type antibody
    • Calpain regulatory subunit antibody
    • Calpain small subunit 1 antibody
    • Calpain, large polypeptide L1 antibody
    • Calpain-1 catalytic subunit antibody
    • Calpain-1 large subunit antibody
    • CAN1_HUMAN antibody
    • CANP 1 antibody
    • CANP antibody
    • CANP small subunit antibody
    • CANP1 antibody
    • CANPL 1 antibody
    • CANPL1 antibody
    • CAPN 1 antibody
    • CAPN1 antibody
    • Cell proliferation inducing protein 30 antibody
    • Cell proliferation-inducing gene 30 protein antibody
    • Micromolar Calpain antibody
    • Micromolar-calpain antibody
    • Mu Calpain antibody
    • muCANP antibody
    • muCL antibody
    see all

Images

  • Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    All lanes : Anti-Calpain 1 antibody [EPR3319] (ab108400)

    Lane 1 : Wild type HAP1 whole cell lysate
    Lane 2 : CAPN1 (Calpain 1) knockout HAP1 whole cell lysate
    Lane 3 : A431 whole cell lysate

    Predicted band size: 82 kDa



    Ab108400 was shown to recognize Calpain 1 in wild-type HAP1. A faint cross-reactive band was observed in  CAPN1 (Calpain 1) knockout cells at 82 kDa when over-exposed. Due to the immunogen’s homology with CAPN2 (Calpain 2 Uniprot: P17655), this band could correspond to the Calpain 2 protein. Please note that cross-reactivity with this protein has not been confirmed experimentally.

  • Flow Cytometry (Intracellular) - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Flow Cytometry (Intracellular) - Anti-Calpain 1 antibody [EPR3319] (ab108400)

    Overlay histogram showing Jurkat cells stained with ab108400 (red line) at 1/380 dilution. The cells were fixed with 2% paraformaldehyde. The secondary antibody used was a FITC conjugated goat anti-rabbit IgG at 1/150 dilution. Isotype control antibody (green line) was rabbit monoclonal IgG used under the same conditions.

  • Immunocytochemistry/ Immunofluorescence - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Immunocytochemistry/ Immunofluorescence - Anti-Calpain 1 antibody [EPR3319] (ab108400)

    ab108400 staining Calpain 1 in the T-47D cell line by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with 4% Paraformaldehyde permeabilized with 0.1% Triton X-100. Samples were incubated with primary antibody (1/70). ab150077(1/500) an Alexa Fluor®488-conjugated Goat anti-rabbit IgG was used as the secondary antibody. Nuclei were counterstained with DAPI.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calpain 1 antibody [EPR3319] (ab108400)

    ab108400 staining Calpain 1 in Human transitional cell carcinoma of bladder tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed and paraffin-embedded, antigen retrieval was by heat mediation in Tris/EDTA buffer pH9. Samples were incubated with primary antibody (1/200). An undiluted HRP-conjugated mouse anti-rabbit IgG was used as the secondary antibody. Tissue counterstained with Hematoxylin. PBS was used in the negative control rather than the Primary antibody.

  • Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    All lanes : Anti-Calpain 1 antibody [EPR3319] (ab108400) at 1/1000 dilution

    Lane 1 : Mouse brain lysate
    Lane 2 : Rat brain lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG, (H+L), HRP-conjugated at 1/1000 dilution

    Predicted band size: 82 kDa

  • Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Anti-Calpain 1 antibody [EPR3319] (ab108400) at 1/1000 dilution + C6 Lysate at 20 µg

    Secondary
    Goat Anti-Rabbit IgG, (H+L), HRP- conjugated at 1/1000 dilution

    Predicted band size: 82 kDa

  • Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Anti-Calpain 1 antibody [EPR3319] (ab108400) at 1/5000 dilution + T47-D Lysate at 10 µg

    Secondary
    Goat Anti-Rabbit IgG, (H+L), HRP- conjugated at 1/1000 dilution

    Predicted band size: 82 kDa

  • Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Anti-Calpain 1 antibody [EPR3319] (ab108400) at 1/5000 dilution + Human fetal kidney lysate at 10 µg

    Secondary
    Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

    Predicted band size: 82 kDa

  • Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Western blot - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    All lanes : Anti-Calpain 1 antibody [EPR3319] (ab108400) at 1/1000 dilution (unpurified)

    Lane 1 : Human fetal kidney cell lysate
    Lane 2 : U937 cell lysate
    Lane 3 : A431 cell lysate
    Lane 4 : 293T cell lysate
    Lane 5 : T47D cell lysate

    Lysates/proteins at 10 µg per lane.

    Predicted band size: 82 kDa

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Calpain 1 antibody [EPR3319] (ab108400)

    ab108400, unpurified, at 1/500 dilution staining Calpain in Human kidney by Immunohistochemistry, Paraffin-embedded tissue.

  • Immunocytochemistry/ Immunofluorescence - Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Immunocytochemistry/ Immunofluorescence - Anti-Calpain 1 antibody [EPR3319] (ab108400)

    ab108400, unpurified, at 1/250 dilution staining Calpain in HeLa cells by Immunofluorescence.

  • Anti-Calpain 1 antibody [EPR3319] (ab108400)
    Anti-Calpain 1 antibody [EPR3319] (ab108400)

References (10)

Publishing research using ab108400? Please let us know so that we can cite the reference in this datasheet.

ab108400 has been referenced in 10 publications.

  • Xu J  et al. Self-Assembling Peptide Scaffold Carrying Neural-Cell Adhesion Molecule-Derived Mimetic-Peptide Transplantation Promotes Proliferation and Stimulates Neurite Extension by Modulating Tau Phosphorylation and Calpain/Glycogen Synthase Kinase 3 beta (GSK-3ß) in Neurons. Ann Transplant 25:e924093 (2020). PubMed: 32686658
  • Yu W  et al. Lens-specific conditional knockout of Msx2 in mice leads to ocular anterior segment dysgenesis via activation of a calcium signaling pathway. Lab Invest N/A:N/A (2019). PubMed: 30683901
  • Yu P  et al. CD163?SRCR5 MARC-145 Cells Resist PRRSV-2 Infection via Inhibiting Virus Uncoating, Which Requires the Interaction of CD163 With Calpain 1. Front Microbiol 10:3115 (2019). PubMed: 32038556
  • Xu C  et al. Overexpression of calpain-1 predicts poor outcome in patients with colorectal cancer and promotes tumor cell progression associated with downregulation of FLNA. Oncol Rep 41:3424-3434 (2019). PubMed: 31002357
  • Wang X  et al. A Label-Free Quantitative Proteomic Analysis of Mouse Neutrophil Extracellular Trap Formation Induced by Streptococcus suis or Phorbol Myristate Acetate (PMA). Front Immunol 9:2615 (2018). PubMed: 30555459
  • Yu LM  et al. High calpain-1 expression predicts a poor clinical outcome and contributes to tumor progression in pancreatic cancer patients. Clin Transl Oncol N/A:N/A (2018). PubMed: 30565085
  • Zeng X  et al. Acylated and unacylated ghrelin inhibit atrophy in myotubes co-cultured with colon carcinoma cells. Oncotarget 8:72872-72885 (2017). PubMed: 29069832
  • Lin XY & Chen SZ Calpain inhibitors ameliorate muscle wasting in a cachectic mouse model bearing CT26 colorectal adenocarcinoma. Oncol Rep 37:1601-1610 (2017). PubMed: 28112357
  • Tsai JC  et al. The role of calpain-myosin 9-Rab7b pathway in mediating the expression of Toll-like receptor 4 in platelets: a novel mechanism involved in a-granules trafficking. PLoS One 9:e85833 (2014). WB ; Human . PubMed: 24489676
  • Ozkan G  et al. How does colistin-induced nephropathy develop and can it be treated? Antimicrob Agents Chemother 57:3463-9 (2013). PubMed: 23629704

Customer reviews and Q&As

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1-2 of 2 Abreviews or Q&A

Immunohistochemistry (Frozen sections) abreview for Anti-Calpain 1 antibody [EPR3319]

 Good
Abreviews
abreview image
Application
Immunohistochemistry (Frozen sections)
Sample
Rabbit Tissue sections (Hippocampus (Neurons in the CA1 Region))
Permeabilization
Yes - 0,1% Triton in PBS
Specification
Hippocampus (Neurons in the CA1 Region)
Blocking step
Serum as blocking agent for 45 minute(s) · Concentration: 10% · Temperature: 25°C
Fixative
Paraformaldehyde
Read More

Helena Lichtenfeld

Verified customer

Submitted Aug 18 2020

Question

A customer have a problem with Calpain 1 antibody (ab108400) IHC
staining. Briefly, the antibody staining pattern is unexpected. You can
find the detailed protocol and staining photos in the attached files.

Read More
Answer

Thank you for taking time to complete our questionnaire. I am sorry to hear that this antibody is not providing satisfactory results.

The details provided will enable us to investigate this case and will provide us with vital information for monitoring product quality.

Having reviewed this case, I would like to offer some suggestions to help optimize the results from ab108400.


How long did they perform the antigen retrieval in the pressure cooker? They may have over-retrieved so they can decrease the amount of time.


They should also omit any permeabilization agent like Triton X-100.


I see that the customer only blocked for 5 - 10 mins and this may be too short of a time. What buffer have they tried and at what percentage? They should block with 10% serum for 1 hr at RT.


They may also want to try diluting the antibody more to 1:2000 or 1:5000 and incubate that overnight at 4C instead of 1 hr at RT.


Have they run a no primary control?


Should the suggestions not improve the results, please do let me know.

In the event that a product is not functioning in the species and applications cited on the product datasheet (and the problem has been reported within 6 months of purchase), we would be pleased to provide a free of charge replacement, credit note, or refund.

I hope this information is helpful, and I thank you for your cooperation.

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