Calreticulin Antibody - BSA Free

Novus Biologicals | Catalog # NB600-103

Novus Biologicals
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Key Product Details

Validated by

Knockout/Knockdown

Species Reactivity

Validated:

Human, Mouse

Cited:

Human, Mouse

Applications

Validated:

Knockout Validated, Immunohistochemistry, Immunohistochemistry-Paraffin, Immunohistochemistry-Frozen, Western Blot, Immunocytochemistry/ Immunofluorescence, Simple Western, Knockdown Validated

Cited:

Immunohistochemistry-Paraffin, Immunohistochemistry-Frozen, Western Blot

Label

Unconjugated

Antibody Source

Polyclonal Rabbit IgG

Format

BSA Free
View all Calreticulin Primary Antibodies »

Product Specifications

Immunogen

Calreticulin Antibody was developed against a full-length protein to mouse Calreticulin [UniProt# P14211]

Localization

Calreticulin staining is localized to the ER in most cell types and under most conditions.

Marker

Endoplasmic Reticulum Marker

Clonality

Polyclonal

Host

Rabbit

Isotype

IgG

Theoretical MW

48 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Scientific Data Images for Calreticulin Antibody - BSA Free

Knockout Validated: Calreticulin Antibody [NB600-103]

Western Blot: Calreticulin Antibody [NB600-103]

Western Blot: Calreticulin Antibody [NB600-103] - Western blot shows lysates of HeLa human cervical epithelial carcinoma parental cell line and Calreticulin knockout (KO) HeLa cell line. PVDF membrane was probed with 1:2500 of Rabbit Anti-Human Calreticulin Polyclonal Antibody (Catalog # NB600-103) followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog #HAF008). Specific band was detected for Calreticulin at approximately 55 kDa (as indicated) in the parental HeLa cell line, but is not detectable in the knockout HeLa cell line. This experiment was conducted under reducing conditions.
Simple Western: Calreticulin Antibody [NB600-103]

Simple Western: Calreticulin Antibody [NB600-103]

Simple Western: Calreticulin Antibody [NB600-103] - Lane view shows a specific band for Calreticulin in 0.2 mg/ml of HeLa lysate. This experiment was performed under reducing conditions using the 12-230 kDa separation system.
Immunocytochemistry/ Immunofluorescence: Calreticulin Antibody [NB600-103]

Immunocytochemistry/ Immunofluorescence: Calreticulin Antibody [NB600-103]

Immunocytochemistry/Immunofluorescence: Calreticulin Antibody [NB600-103] - IF Confocal analysis of 3T3 cells using Calreticulin antibody (NB600-103, 1:20). An Alexa Fluor 488-conjugated Goat to rabbit IgG was used as secondary antibody (green). Actin filaments were labeled with Alexa Fluor 568 phalloidin (red). DAPI was used to stain the cell nuclei (blue).
Immunohistochemistry-Paraffin: Calreticulin Antibody [NB600-103]

Immunohistochemistry-Paraffin: Calreticulin Antibody [NB600-103]

Immunohistochemistry-Paraffin: Calreticulin Antibody [NB600-103] - Analysis of FFPE human prostate cancer using Calreticulin antibody at 1:20 on a Bond Rx autostainer (Leica Biosystems). The assay involved 20 minutes of heat induced antigen retrieval (HIER) using 10mM sodium citrate buffer (pH 6.0) and endogenous peroxidase quenching with peroxide block. The sections were incubated with primary antibody for 30 minutes and Bond Polymer Refine Detection (Leica Biosystems) with DAB was used for signal development followed by counterstaining with hematoxylin. Whole slide scanning and capturing of representative images was performed using Aperio AT2 (Leica Biosystems). Cytoplasmic staining in glands was observed. Staining was performed by Histowiz.
Western Blot: Calreticulin Antibody [NB600-103]

Western Blot: Calreticulin Antibody [NB600-103]

Western Blot: Calreticulin Antibody [NB600-103] - Detection of calreticulin in human kidney lysate using NB600-103 at 1:1,000. ECL exposure, 10 seconds.
Western Blot: Calreticulin Antibody [NB600-103]

Western Blot: Calreticulin Antibody [NB600-103]

Western Blot: Calreticulin Antibody [NB600-103] - Analysis of HeLa and C6 cell lysate using calreticulin antibody at 1:1000.

Applications for Calreticulin Antibody - BSA Free

Application
Recommended Usage

Immunocytochemistry/ Immunofluorescence

1:20-1:100. Use reported by customer review

Immunohistochemistry

1:20

Immunohistochemistry-Frozen

reported in scientific literature (PMID 26643212)

Immunohistochemistry-Paraffin

1:20

Knockdown Validated

1:2500

Simple Western

1:500

Western Blot

1:1000-1:10000
Application Notes
A band at approx. 55 kDa is seen in Western Blot. Note that Calreticulin has several reported isoforms that range from 48-62 kDa.

In Simple Western only 10 - 15 uL of the recommended dilution is used per data point.
See Simple Western Antibody Database for Simple Western validation: Tested in HeLa lysate 0.2 mg/mL, separated by Size, antibody dilution of 1:500, apparent MW was 45 kDa. Separated by Size-Wes, Sally Sue/Peggy Sue.

Reviewed Applications

Read 3 reviews rated 4.7 using NB600-103 in the following applications:

Formulation, Preparation, and Storage

Purification

Unpurified

Formulation

Whole antisera

Format

BSA Free

Preservative

0.1% Sodium Azide

Concentration

This product is unpurified. The exact concentration of antibody is not quantifiable.

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: Calreticulin

Calreticulin, also called CALR, is an endoplasmic reticulum (ER) protein that functions as a calcium (Ca2+) buffering molecular chaperone (1-3). Together with calnexin and ERp57, calreticulin has a critical role in protein folding and quality control of newly synthesized glycoproteins (1-3). Calreticulin has many diverse functions beyond Ca2+ binding, including cell adhesion, lectin binding, apoptosis, nuclear transport, antigen presentation, and immune response (1-3). Specifically, calreticulin is part of the peptide-loading complex that resides on the ER membrane and is responsible for antigen loading onto MHC class I molecules (1-3). Calreticulin protein has a theoretical molecular weight of 46 kDa and is 417 amino acids (aa) in length (1-3). The protein has three main domains: the N-terminal lectin-like domain, the proline-rich P-domain, and the acidic C-domain which is followed by an ER-retention KDEL signal sequence (1-3).

Given its role in multiple biological processes, it makes sense that calreticulin is implicated in both healthy and disease states. Studies have found that calreticulin mutations were present in patients with myeloproliferative neoplasms (MFN) and essential thrombocythaemia (ET) (4). Calreticulin expression is typically upregulated in most cancer lines, however it is downregulated in some tissues including cervical carcinomas, prostate cancer, and human colon adenocarcinoma (3). High expression of calreticulin on cancer cells is related to tumor cell phagocytosis and is often correlated with, and counteracted by, elevated CD47 expression to prevent cancer cell phagocytosis (3). Calreticulin mutations can serve as a major diagnostic biomarker for MFN and ET and additionally the calreticulin gene may be a potential target for cancer therapeutics (3,4).

References

1. Michalak, M., Groenendyk, J., Szabo, E., Gold, L. I., & Opas, M. (2009). Calreticulin, a multi-process calcium-buffering chaperone of the endoplasmic reticulum. The Biochemical Journal. https://doi.org/10.1042/BJ20081847

2. Fucikova, J., Spisek, R., Kroemer, G., & Galluzzi, L. (2021). Calreticulin and cancer. Cell Research. https://doi.org/10.1038/s41422-020-0383-9

3. Sun, J., Mu, H., Dai, K., & Yi, L. (2017). Calreticulin: a potential anti-cancer therapeutic target. Die Pharmazie. https://doi.org/10.1691/ph.2017.7031

4. Prins, D., Gonzalez Arias, C., Klampfl, T., Grinfeld, J., & Green, A. R. (2020). Mutant Calreticulin in the Myeloproliferative Neoplasms. HemaSphere. https://doi.org/10.1097/HS9.0000000000000333

Alternate Names

Autoantigen Ro, CALR, Calregulin, cC1qR, CRT, SSA, Vasostatin

Entrez Gene IDs

811 (Human)

Gene Symbol

CALR

UniProt

P14211 (Mouse)

Additional Calreticulin Products

Product Documents for Calreticulin Antibody - BSA Free

Certificate of Analysis

To download a Certificate of Analysis, please enter a lot or batch number in the search box below.

Product Specific Notices for Calreticulin Antibody - BSA Free

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Citations for Calreticulin Antibody - BSA Free

Customer Reviews for Calreticulin Antibody - BSA Free (3)

4.7 out of 5
3 Customer Ratings
5 Stars
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4 Stars
33%
3 Stars
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1 Stars
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Customer Images

Showing  1 - 3 of 3 reviews Showing All
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  • Calreticulin Antibody
    Name: Anonymous
    Application: Western Blot
    Sample Tested: Spinal cord
    Species: Mouse
    Verified Customer | Posted 01/07/2019
    Careticulin band can be observed at about 60 kDa.
    Whole spinal cord extracts, 30 ug of protein loaded. Primary antibody: 1/500, one hour secondary antibody: 1/1000, one hour
    Calreticulin Antibody - BSA Free NB600-103
  • Name: Bryan Tinsley
    Application: Immunocytochemistry
    Sample Tested:
    Species: Mouse
    Verified Customer | Posted 06/11/2014
    IF Confocal analysis of 3T3 cells using Calreticulin antibody (NB600-103, 1:20).
    Calreticulin Antibody - BSA Free NB600-103
  • Name: Bryan Tinsley
    Application: Western Blot
    Sample Tested:
    Species: Human
    Verified Customer | Posted 06/11/2014
    Western blot analysis of extracts from HeLa and C6 cells using Calreticulin antibody (NB600-103, 1:1000).
    Calreticulin Antibody - BSA Free NB600-103

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Protocols

View specific protocols for Calreticulin Antibody - BSA Free (NB600-103):


Western Blot Protocol

1. Perform SDS-PAGE (4-12%) on samples to be analyzed, loading 40 ug of total protein per lane.

2. Transfer proteins to Nitrocellulose according to the instructions provided by the manufacturer of the transfer apparatus.

3. Rinse membrane with dH2O and then stain the blot using ponceau S for 1-2 minutes to access the transfer of proteins onto the nitrocellulose membrane. Rinse the blot in water to remove excess stain and mark the lane locations and locations of molecular weight markers using a pencil.

4. Rinse the blot in TBS for approximately 5 minutes.

5. Block the membrane using 5% non-fat dry milk + 1% BSA in TBS for 1 hour at room temperature.

6. Rinse the membrane in dH2O and then wash the membrane in wash buffer [TBS + 0.1% Tween] 3 times for 10 minutes each.

7. Dilute the rabbit anti-calreticulin primary antibody (NB 600-103) in blocking buffer and incubate 1.5 hours at room temperature.

8. Rinse the membrane in dH2O and then wash the membrane in wash buffer [TBS + 0.1% Tween] 3 times for 10 minutes each.

9. Apply the diluted rabbit-IgG HRP-conjugated secondary antibody in blocking buffer (as per manufacturer's instructions) and incubate 1 hour at room temperature.

10. Wash the blot in wash buffer [TBS + 0.1% Tween] 3 times for 10 minutes each (this step can be repeated as required to reduce background).

11. Apply the detection reagent of choice in accordance with the manufacturer's instructions (Pierce's ECL).

Note: Tween-20 can be added to the blocking or antibody dilution buffer at a final concentration of 0.05-0.2%, provided it does not interfere with antibody-antigen binding.

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FAQs

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