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    grp78-bip-antibody-epr40402-ab108613.pdf

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RecombinantRabMAb

Recombinant Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)

  • Datasheet
Submit a review Q&A (6)References (34)

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Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
  • Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
  • Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
  • Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
  • Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
  • Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
  • Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR4040(2)] to GRP78 BiP
  • Suitable for: WB, IHC-P
  • Reacts with: Mouse, Rat, Human

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Overview

  • Product name

    Anti-GRP78 BiP antibody [EPR4040(2)]
    See all GRP78 BiP primary antibodies
  • Description

    Rabbit monoclonal [EPR4040(2)] to GRP78 BiP
  • Host species

    Rabbit
  • Specificity

    The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.

    Stimulation may be required to allow detection of the target protein due to low levels of endogenous expression in some samples.  Please see images below for recommended treatment conditions and positive controls.

  • Tested Applications & Species

    Application Species
    IHC-P
    Human
    WB
    Mouse
    Rat
    Human
    See all applications and species data
  • Immunogen

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

  • Positive control

    • WB: HeLa, LnCaP, ECV-304, C6, NIH/3T3, PC-12, C6 (Rat glial tumor glial cell), RAW264.7, MCF7, 293T, U-87 MG, THP-1, MDA-MB-231 and HepG2 cell lysates; Human liver lysates; Mouse heart, lung, testis and liver lysates; Rat liver lysates. IHC-P: Human brain, ovarian cancer and testis tissue.
  • 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

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
  • Storage buffer

    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.5% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EPR4040(2)
  • Isotype

    IgG
  • Research areas

    • Tags & Cell Markers
    • Subcellular Markers
    • Organelles
    • ER

Associated products

  • Alternative Versions

    • Alexa Fluor® 647 Anti-GRP78 BiP antibody [EPR4040(2)] (ab225082)
    • Alexa Fluor® 488 Anti-GRP78 BiP antibody [EPR4040(2)] (ab225083)
    • Anti-GRP78 BiP antibody [EPR4040(2)] - BSA and Azide free (ab226042)
  • Compatible Secondaries

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab97051)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • Recombinant Protein

    • Recombinant human GRP78 BiP protein (Active) (ab78432)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab108613 in the following tested applications.

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

Guaranteed

Tested applications are guaranteed to work and covered by our Abpromise guarantee.

Predicted

Predicted to work for this combination of applications and species but not guaranteed.

Incompatible

Does not work for this combination of applications and species.

Application Species
IHC-P
Human
WB
Mouse
Rat
Human
Application Abreviews Notes
WB
1/1000 - 1/10000. Detects a band of approximately 78 kDa (predicted molecular weight: 72 kDa).
IHC-P
1/250 - 1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

See IHC antigen retrieval protocols. (Heat to 98oC, allow to cool for 10-20 minutes) 

The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.

Notes
WB
1/1000 - 1/10000. Detects a band of approximately 78 kDa (predicted molecular weight: 72 kDa).
IHC-P
1/250 - 1/500. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

See IHC antigen retrieval protocols. (Heat to 98oC, allow to cool for 10-20 minutes) 

The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.

Target

  • Function

    Probably plays a role in facilitating the assembly of multimeric protein complexes inside the endoplasmic reticulum. Involved in the correct folding of proteins and degradation of misfolded proteins via its interaction with DNAJC10, probably to facilitate the release of DNAJC10 from its substrate.
  • Involvement in disease

    Autoantigen in rheumatoid arthritis.
  • Sequence similarities

    Belongs to the heat shock protein 70 family.
  • Cellular localization

    Endoplasmic reticulum lumen. Melanosome. Cytoplasm. Identified by mass spectrometry in melanosome fractions from stage I to stage IV.
  • Target information above from: UniProt accession P11021 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 3309 Human
    • Entrez Gene: 14828 Mouse
    • Entrez Gene: 25617 Rat
    • Omim: 138120 Human
    • SwissProt: P11021 Human
    • SwissProt: P20029 Mouse
    • SwissProt: P06761 Rat
    • Unigene: 743241 Human
    • Unigene: 330160 Mouse
    • Unigene: 470180 Mouse
    • Unigene: 474909 Mouse
    • Unigene: 11088 Rat
    see all
  • Alternative names

    • 78 kDa glucose regulated protein antibody
    • 78 kDa glucose-regulated protein antibody
    • AL022860 antibody
    • AU019543 antibody
    • BIP antibody
    • D2Wsu141e antibody
    • D2Wsu17e antibody
    • Endoplasmic reticulum lumenal Ca(2+)-binding protein grp78 antibody
    • Endoplasmic reticulum lumenal Ca2+ binding protein grp78 antibody
    • Epididymis secretory sperm binding protein Li 89n antibody
    • FLJ26106 antibody
    • Glucose Regulated Protein 78 antibody
    • Glucose Regulated Protein 78kDa antibody
    • GRP 78 antibody
    • GRP-78 antibody
    • GRP78 antibody
    • GRP78_HUMAN antibody
    • Heat shock 70 kDa protein 5 antibody
    • Heat Shock 70kDa Protein 5 antibody
    • Heat shock protein family A (Hsp70) member 5 antibody
    • HEL S 89n antibody
    • Hsce70 antibody
    • HSPA 5 antibody
    • HSPA5 antibody
    • Immunoglobulin Heavy Chain Binding Protein antibody
    • Immunoglobulin heavy chain-binding protein antibody
    • mBiP antibody
    • MIF2 antibody
    • Sez7 antibody
    see all

Images

  • Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    All lanes : Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613) at 1/1000 dilution

    Lane 1 : MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysates
    Lane 2 : HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysates
    Lane 3 : 293T (Human embryonic kidney epithelial cell) whole cell lysates
    Lane 4 : U-87 MG (Human glioblastoma-astrocytoma epithelial cell) whole cell lysates
    Lane 5 : THP-1 (Human monocytic leukemia monocyte) whole cell lysates
    Lane 6 : MDA-MB-231 (Human breast adenocarcinoma epithelial cell) whole cell lysates
    Lane 7 : NIH/3T3 (Mouse embryonic fibroblast) whole cell lysates
    Lane 8 : Human uterus lysates
    Lane 9 : Human liver lysates
    Lane 10 : Mouse heart lysates
    Lane 11 : Mouse lung lysates
    Lane 12 : Mouse testis lysates
    Lane 13 : Mouse liver lysates
    Lane 14 : Rat liver lysates

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)

    Predicted band size: 72 kDa
    Observed band size: 78 kDa
    why is the actual band size different from the predicted?


    Exposure time: 37 seconds
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human ovarian cancer tissue sections labeling GRP78 BiP with Purified ab108613 at 1:500 dilution (1.17 µg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylinwas used as a counterstain.

  • Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    All lanes : Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613) at 1/1000 dilution

    Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates
    Lane 2 : HeLa treated with 2.5 µg/ml tunicamycin for 24h whole cell lysates
    Lane 3 : HUVEC (Human umbilical vein endothelial cell) whole cell lysates
    Lane 4 : HUVEC treated with 10 µg/ml tunicamycin for 48h whole cell lysates
    Lane 5 : Raw 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysates
    Lane 6 : Raw 264.7 treated with 5 µg/ml tunicamycin for 18h whole cell lysates

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)

    Predicted band size: 72 kDa
    Observed band size: 78 kDa why is the actual band size different from the predicted?


    Exposure time: 3 seconds
  • Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613) at 0.1 µg/ml (purified) + C6 (Rat glial tumor glial cell) whole cell lysates at 15 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 72 kDa



    Blocking and diluting buffer: 5% NFDM/TBST.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)

    Immunohistochemical analysis of paraffin-embedded Human testis tissue using unpurified ab108613 at 1/250

  • Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    All lanes : Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613) at 1/5000 dilution (purified)

    Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates
    Lane 2 : HeLa (Human cervix adenocarcinoma epithelial cell) treated with 10ug/ml
    tunicamycin for 12 hours whole cell lysates
    Lane 3 : MCF-7 (Human breast adenocarcinoma epithelial cell) whole cell lysates
    Lane 4 : MCF-7 (Human breast adenocarcinoma epithelial cell) treated with 10ug/ml
    tunicamycin for 48 hours whole cell lysates
    Lane 5 : MEF (Mouse embryonic fibroblast) whole cell lysates
    Lane 6 : MEF (Mouse embryonic fibroblast) treated with 10ug/ml tunicamycin for 48 hours whole cell lysates

    Lysates/proteins at 15 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 72 kDa



    Blocking and diluting buffer: 5% NFDM/TBST.

    Tunicamycin induces the expression of GRP78 BiP reported by PMID 22859938

    Exposure time:
    Lanes 1 and 2: 3 seconds 
    Lanes 3 and 4: 30 seconds
    Lanes 5 and 6: 5 seconds

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)

    Immunohistochemical analysis of paraffin-embedded Human brain tissue using unpurified ab108613 at 1/250

  • Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    All lanes : Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613) at 1/1000 dilution (Unpurified)

    Lane 1 : HeLa cell lysate
    Lane 2 : LnCaP cell lysate
    Lane 3 : ECV-304 cell lysate
    Lane 4 : HepG2 cell lysate

    Lysates/proteins at 10 µg per lane.

    Predicted band size: 72 kDa
    Observed band size: 78 kDa why is the actual band size different from the predicted?

  • Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    Western blot - Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    All lanes : Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613) at 1/1000 dilution (Unpurified)

    Lane 1 : C6 cell lysate
    Lane 2 : NIH/3T3 cell lysate
    Lane 3 : PC-12 cell lysate
    Lane 4 : RAW264.7 cell lysate

    Lysates/proteins at 10 µg per lane.

    Predicted band size: 72 kDa
    Observed band size: 78 kDa why is the actual band size different from the predicted?

  • Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)
    Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613)

Protocols

  • Western blot protocols
  • Immunohistochemistry protocols

Click here to view the general protocols

Datasheets and documents

    • Datasheet
  • References (34)

    Publishing research using ab108613? Please let us know so that we can cite the reference in this datasheet.

    ab108613 has been referenced in 34 publications.

    • Wu Q  et al. Anti-Nogo-A antibody promotes brain function recovery after cardiopulmonary resuscitation in rats by reducing apoptosis. Mol Med Rep 21:77-88 (2020). PubMed: 31746353
    • Gao G  et al. Rapamycin regulates the balance between cardiomyocyte apoptosis and autophagy in chronic heart failure by inhibiting mTOR signaling. Int J Mol Med 45:195-209 (2020). PubMed: 31746373
    • Chen Y  et al. Iridoid glycosides from Radix Scrophulariae attenuates focal cerebral ischemia-reperfusion injury via inhibiting endoplasmic reticulum stress-mediated neuronal apoptosis in rats. Mol Med Rep 21:131-140 (2020). PubMed: 31746404
    • Wang WP  et al. Progerin in muscle leads to thermogenic and metabolic defects via impaired calcium homeostasis. Aging Cell 19:e13090 (2020). PubMed: 31833196
    • Li T  et al. Combination of Dichloroacetate and Atorvastatin Regulates Excessive Proliferation and Oxidative Stress in Pulmonary Arterial Hypertension Development via p38 Signaling. Oxid Med Cell Longev 2020:6973636 (2020). PubMed: 32617141
    View all Publications for this product

    Customer reviews and Q&As

    Show All Reviews Q&A
    Submit a review Submit a question

    1-6 of 6 Abreviews or Q&A

    Question

    What is the IHC-P protocol for ab108613?

    Read More

    Abcam community

    Verified customer

    Asked on Mar 15 2012

    Answer

    Thank you for contacting us. Below is the protocol used to validate ab108613 in IHC-P. Please let me know if you need any additional information or assistance.

    1. Solutions and reagents
    1.1. Xylene
    1.2. Ethanol, anhydrous denatured, histological grade (100%, 95%, 70%)
    1.3. Washing buffer: TBST washing buffer: 1XTBS/0.1% Tween-20
    To prepare stock solution of 10X TBS: add 24.2 g Trizma base and 80 g sodium chloride to 1L of dH2O.
    Adjust pH to 7.6.
    Working solution. 1XTBST/0.1% Tween-20: add 100ml 10XTBS to 900 ml dH2O. Add 1 ml Tween-20 and
    mix well.
    1.4. Distilled water (dH2O)
    1.5. Antigen Retrieval Solution: 0.01M Sodium Citrate Buffer, pH 6.0
    To prepare stock solutions:
    Solution A. 0.1 M citric acid solution: dissolve 21.0 g of citric acid, monohydrate (C6H8O7.H2O) in 100 ml
    of dH2O.
    Solution B. 0.1M sodium citrate solution: dissolve 29.4 g trisodium citrate dihydrate (C6H5Na3O7.2H2O)
    in 100 ml of dH2O.
    Working solution: Add 9 ml of Stock solution A and 41 ml of stock solution B to 450 ml of dH2O. Adjust
    pH to 6.0.
    1.6. 3% Hydrogene Peroxide
    1.7. Blocking buffer: PBS + 10% serum (serum origin depends on the host of the secondary antibody)
    1.8. Hematoxylin QS.
    1.9. Permanent Mounting medium.
    2. Protocol
    2.1. Deparaffinization/Rehydration
    2.1.1. Heat slides in an oven at 65 °C for 1 hour.
    2.1.2. De-paraffinize/hydrate using the following series of washes: two Xylene washes (5 min each), followed by two
    100% ethanol rinses (5 min each), followed by 95% ethanol, 70% ethanol, 50% ethanol, 30% ethanol, followed by H
    2
    O
    and a TBST wash for 5 min on a shaker.
    2.2. Antigen Retrieval
    2.2.1 Immerse slides into staining dish containing Antigen Retrieval Solution.
    2.2.2 Place covered staining dish into the rice cooker. Add 120 mL dH
    2
    O and press “cook”.
    2.2.3 When “cook” is turned to “warm” (about 20–30 min), unplug the cooker and remove the staining dish to the bench
    top.
    2.2.4 Allow to cool down, without cover, for 20 min.
    2.3. Staining
    2.3.1. Wash slides with TBST for 5 min on a shaker.
    2.3.2. Inactivate endogenous peroxidase by covering tissue with 3% hydrogen peroxide for 10 min.
    2.3.3. Wash slides three times with TBST (3 min each on a shaker).
    2.3.4. Block slides with the blocking solution for 1 hour.
    2.3.5. Dilute primary antibody in the blocking buffer per recommendation on the data sheet.
    2.3.6. Apply primary antibody to each section and incubate overnight in the humidified chamber (4 ºC).
    2.3.7. Wash slides three times with TBST (3 min each on a shaker).
    2.3.8. Apply to each section secondary HRP-conjugated anti-rabbit antibody diluted in the blocking solution per
    manufacturer’s recommendation; incubate for 1 hour at room temperature.
    2.3.9. Wash slides three times with TBST (3 min each on a shaker).
    2.3.10. Add freshly prepared DAB substrate to the sections.
    2.3.11. Incubate tissue sections with the substrate at room temperature until suitable staining develops (generally 2–5
    min).
    2.3.12. Rinse sections with water.
    2.3.13. Counterstain with Hematoxylin.
    2.3.14. Rinse sections with water.
    2.3.15. Dehydrate samples using two rinses with 100% Ethanol (20 dips per rinse) followed by two rinses with Xylene (30
    dips per rinse).
    2.3.16. Mount coverslips on slides using Permount medium.

    Read More

    Abcam Scientific Support

    Answered on Mar 15 2012

    Question




    Thanks for helping out with this inquiry. I attached an annotated power point of the data and the page from my lab notebook that contains the blotting map. If you need anything else from me, let me know.

    Read More

    Abcam community

    Verified customer

    Asked on Jul 20 2012

    Answer



    "We only saw a faint band at 50 kDa using mouse and rat heart, which looks non-specific on our blots. I have not found the identity of this band in the literature. The blocking reagent used was 5% milk. We used our standard WB protocol (see attachment)."

    Let me know if you have anymore questions.

    Read More

    Abcam Scientific Support

    Answered on Jul 20 2012

    Question


    Thank you for your reply. I am so sorry to tell you that we cantest the GRP94 protein which has the similar function and position withGRP78 by western blotting.We extracted the GRP94 protein from SD brain
    tissue using our lysis buffer.The attachment contains the result of GRP94.

    Looking forward your reply!

    Read More

    Abcam community

    Verified customer

    Asked on May 30 2012

    Answer

    Thank you for your reply.
    If you were able to detect a different ER protein using the same samples then it doesn't seem like it was a problem with the sample preparation.
    I would like to offer you a free of charge replacement of ab108613 (although we only have the same lot in stock as the one you tried), a different GRP78 BiP antibody, or a credit note/refund. Please let me know which you would prefer.
    I look forward to your reply.

    Read More

    Abcam Scientific Support

    Answered on May 30 2012

    Question

    We can not get the positive result based on the protocol. Thanks!

    Read More

    Abcam community

    Verified customer

    Asked on May 24 2012

    Answer

    Thank you for taking time to send us your protocol. 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 ab108613.
    We would recommend changing your lysis buffer to RIPA buffer which includes SDS. Since this is an ER protein, you're going to need a pretty strong lysis buffer to disrupt the structure and make the protein accessible to the antibody.
    RIPA buffer (Radio Immuno Precipitation Assay buffer)
    150 mM sodium chloride
    1.0% NP-40 or Triton X-100
    0.5% sodium deoxycholate
    0.1% SDS (sodium dodecyl sulphate)
    50 mM Tris, pH 8.0
    Also, you do an extra blocking step after the primary antibody incubation but this may not be necessary. Here is the protocol used with this antibody:
    Western Blotting

    1. Block nitrocellulose for 1 hour at room temperature or overnight at 4°C using 5% BSA.

    2.Incubate nitrocellulose with appropriate dilutions of primary antibody in 5% BSA overnight at 4°C or for 2 hours at room temperature
    3. Wash nitrocellulose with three 5-min washes using TBST.

    4.Incubate nitrocellulose with goat anti-rabbit HRP antibody, diluted to 1:500 to 100,000 in 5% BSA or 5% NFDM, for 1 hour at room temperature.

    5. Wash nitrocellulose in 3 washes of TBST, then rinse in TBS prior to addition of chemiluminesence reagents.

    6. Remove excess chemiluminescence reagent and sandwich nitrocellulose blot in any type of transparent plastic wrap (plastic cling wrap, transparent sheet protector, etc.).

    7. Acquire image using darkroom development techniques.
    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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    Abcam Scientific Support

    Answered on May 24 2012

    Question

    Phone call to confirm to replace ab2902 with ab108613 and to accept testing discount for ab108613 in ICC/IF.

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    Abcam community

    Verified customer

    Asked on Mar 27 2012

    Answer


    Merci de votre appel. Je suis ravie que vous souhaitiez participer a notre offre et tester ab108613 en immunofluorescence sur des cellules (ICC/IF). Le code de réduction correspondant est xx. Ce code de réduction est utilisable sous 120 jours et pour l’achat d’un autre anticorps primaire.

    Comme convenu au téléphone, nous avons aussi validé la commande pour le ab108613.

    Vous devez dans un premier temps soumettre une Abreview avec vos résultats en ICC/IF en notant ce code dans la section « Additional notes ». Ce code sera ainsi activé et vous pourrez par la suite contacter notre service clientèle par téléphone pour passer votre prochaine commande, en vous munissant des informations suivantes : code de réduction, numéro de commande de ab108613.
    Veuillez consulter le site www.abcam.com/abreviews pour plus d'information sur les abreviews.

    Nous restons à votre disposition pour plus d’informations concernant cette offre promotionnelle. Dans l’attente de recevoir votre Abreview nous vous souhaitons bonne chance dans vos recherches.

    Termes et Conditions : www.abcam.com/collaborationdiscount.

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    Abcam Scientific Support

    Answered on Mar 27 2012

    Question

    Our customer makes a request about Anti-GRP78 BiP antibody [EPR4040(2)] (ab108613). She has ordered the ab108613 recently. Would you please offer the COC for her?

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    Abcam community

    Verified customer

    Asked on Mar 15 2012

    Answer

    Thank you for contacting us, please find the requested CoC attached. Please let me know if you need any other information!

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    Abcam Scientific Support

    Answered on Mar 15 2012

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