製品の詳細

  • 製品名

    Recombinant human GRP78 BiP protein (Active)
    GRP78 BiP タンパク質・ペプチド 製品一覧
  • 生理活性

    ab78432 has ATPase activity at the time of manufacture of 2.3µM phosphate liberated/hr/µg protein in a 200µl reaction at 37°C (pH7.5) in the presence of 20ul of 1mM ATP, using a Malachite Green assay.

  • 精製度

    > 90 % SDS-PAGE.
    Ab78432 is purified by affinity purification.
  • 発現系

    Escherichia coli
  • アクセッション番号

  • タンパク質長

    Full length protein
  • Animal free

    No
  • 由来

    Recombinant
    • 生物種

      Human
    • 配列

      MKLSLVAAML LLLSAARAEE EDKKEDVGTV VGIDLGTTYS CVGVFKNGRV EIIANDQGNR ITPSYVAFTP EGERLIGDAA KNQLTSNPEN TVFDAKRLIG RTWNDPSVQQ DIKFLPFKVV EKKTKPYIQV DIGGGQTKTF APEEISAMVL TKMKETAEAY LGKKVTHAVV TVPAYFNDAQ RQATKDAGTI AGLNVMRIIN EPTAAAIAYG LDKREGEKNI LVFDLGGGTF DVSLLTIDNG VFEVVATNGD THLGGEDFDQ RVMEHFIKLY KKKTGKDVRK DNRAVQKLRR EVEKAKRALS SQHQARIEIE SFYEGEDFSE TLTRAKFEEL NMDLFRSTMK PVQKVLEDSD LKKSDIDEIV LVGGSTRIPK IQQLVKEFFN GKEPSRGINP DEAVAYGAAV QAGVLSGDQD TGDLVLLDVC PLTLGIETVG GVMTKLIPRN TVVPTKKSQI FSTASDNQPT VTIKVYEGER PLTKDNHLLG TFDLTGIPPA PRGVPQIEVT FEIDVNGILR VTAEDKGTGN KNKITITNDQ NRLTPEEIER MVNDAEKFAE EDKKLKERID TRNELESYAY SLKNQIGDKE KLGGKLSSED KETMEKAVEE KIEWLESHQD ADIEDFKAKK KELEEIVQPI ISKLYGSAGP PPTGEEDTAE KDEL
    • 予測される分子量

      78 kDa
    • 配列の追加情報

      his-tagged (Gene ID 3309)(NM_005347)

特性

Our Abpromise guarantee covers the use of ab78432 in the following tested applications.

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

  • アプリケーション

    Other

    Western blot

    ELISA

    Functional Studies

    competitive binding assays

    SDS-PAGE

  • 製品の状態

    Liquid
  • Concentration information loading...

前処理および保存

  • 保存方法および安定性

    Shipped at 4°C. Store at -20°C. Stable for 12 months at -20°C.

    pH: 7.40
    Constituents: 0.61% Tris, 0.87% Sodium chloride, 10% Glycerol

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

関連情報

  • 別名

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

    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.
  • 関連疾患

    Autoantigen in rheumatoid arthritis.
  • 配列類似性

    Belongs to the heat shock protein 70 family.
  • 細胞内局在

    Endoplasmic reticulum lumen. Melanosome. Cytoplasm. Identified by mass spectrometry in melanosome fractions from stage I to stage IV.
  • Information by UniProt

画像

  • SDS-PAGE of 78kDa Grp78 (Bip) protein.
  • Lane 1. Molecular weight ladder

    lane 2. GRP78 BiP protein

参考文献

This product has been referenced in:

  • Hossain Z  et al. Discovery of pancreastatin inhibitor PSTi8 for the treatment of insulin resistance and diabetes: studies in rodent models of diabetes mellitus. Sci Rep 8:8715 (2018). Read more (PubMed: 29880906) »
  • Biswas N  et al. Discovery of a novel target for the dysglycemic chromogranin A fragment pancreastatin: interaction with the chaperone GRP78 to influence metabolism. PLoS One 9:e84132 (2014). Functional Studies . Read more (PubMed: 24465394) »
See all 3 Publications for this product

レビューと Q&A

1-5 of 5 Abreviews or Q&A

Application
ELISA
The following protocol was used to optimize a standard curve in an ELISA assay intended to quantify GRP78 human protein in blood serum samples:

Reagents:
Coating buffer for primary Antobody: Bicarbonate/carbonate coating buffer (100 mM) :
3.03 g Na2CO3,
6.0 g NaHCO3
1000 ml distilled water
Adjust to pH 9.6 (using NaOH or HCL)
 Washing buffer (aka. PBST): 0.05% Tween 20 in PBS (250µLTween 20, 655204, CALBIOCHEM) in 500mL PBS)
 Sample dilution buffer for GRP78 standard (ab78432, Abcam): PBS
 Blocking buffer: 5% Nonfat milk (dissolved in PBS)
 Capture Ab: Rat Ab (ab25192, Abcam), 0.5mg/mL, 200µL of 0.5 mg/mL rat anti-GRP78 mAbis diluted in 10mL PBS, where the final concentration is 10µg/mL (1:50 dilution)
 Detection Ab: Rabbit polyclonal against GRP78 (ab21685): 1:1,000 dilution. For each 96-well plate (High Binding 96 well plate [Immunogen, Thermo, S0611810]), 10 µL of Rabbit anti-GRP78 pAb (ab21685, Abcam) dissolved in 10mL PBS (final dilution is 1:1,000)
 Secondary Ab: Donkey anti rabbit-HRP, sc-2313, Santa Cruz, in PBS (1:8,000 dilution). For each 96-well plate, (1:8,000) 1.25 µL of Donkey anti-rabbit secondary Ab (sc-2313, Santa Cruz) dissolved in 10mL PBS
 Patient’s serum dilution: 1:50 dilution (20 µL sample is diluted in 980 µL PBS)
 GRP78 standard: GRP78 (ab78432, Abcam,50µg at 1000µg/mL): serial dilutions starting at 4000 pg/mL (4000, 2000, 1000, 500, 50, 10, 1 pg/mL)
 TMB solution
 TMB stopping solution (1N H2SO4).

Procedure

1. Add 100 µL of Rat anti-GRP78, 10µg/mL Mab (76-E6) (ab25192, Abcam) into each well in a high binding 96-well plate (Corner Notch, Thermo, 15041).
2. After adding 100 µL anti-GRP78 Mab, seal the plates and incubate overnight at 4°C.
3. Aspirate the anti-GRP 78 Mab antibodies out of the wells.
4. Wash each well with 200 µL of PBST (PBS + 0.05% Tween20) at 5 min x 3 times to remove unbounded anti-GRP78.
5. Add 200 µL 0.1% BSA (0.05g BSA in 50 mL PBS) to each well and incubate for 2 hrs at room temperature, or overnight at 4oC.
6. Remove the BSA and wash each well with 200 µL of PBST (PBS + 0.05% Tween20) at 5 min x 3 times to remove unbounded anti-GRP78.
7. Add samples (100 µL) that have been diluted 1:50 in PBS and GRP78 standard solution (100 µL) into each well in duplicates.
8. Incubate plate at 37 °C for 1.5 hrs on a shake and bake oven.
9. Remove samples via aspiration after incubation and wash each well with 200 µL PBST at 5 min X 3 times
10. Add 100 µL of detecting Ab (rabbit polyclonal against GRP78, ab21685, abcam, 1:1,000), seal plate and incubate for 2 hours 37.
11. Remove the rabbit polyclonal anti-GRP78 (ab21685) via aspiration and wash plate for 5 min X 4 times with PBST.
12. Add 100 µL of Secondary Ab (donkey anti rabbit-IgG-HRP, sc-2313, Santa Cruz, 1:8,000) into each well in the plate.
13. Seal plate and incubate at room temperature for 2hrs.
14. Discard Secondary Ab and wash plate for 5 min x 4 times with PBST.
15. Add the chromagen (100 µL of TMB (substrate) to the plate and incubate for15 min at 37 degrees Celsius (if blue color is too faint, increase incubation time).
16. Stop the reaction by adding 50 µL of 1N H2SO4 into each well and mix gently.
17. Check the absorbance at 450nm with Spectrophotometer.

Results: absorbance was the same across all concentrations of GRP78 protein standard (including background).

Abcam user community

Verified customer

投稿 Jan 08 2014

Answer

Thank you for contacting Abcam.

The purity of ATP used in the kit used to calculate the biological activity of the protein was >99%.

I believe my colleagues have already emailed you with the catalogue number of the Malachite Green assay that was used during the testing.

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

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Answer

Thank you for your reply.

I apologize, however there was a misunderstanding. I did not know that you were asking for the malachite green based assay used for assessing the activity of ab78432.

We do not knowwith which kitthe recombinant GRP78ab78432 activity has been tested. We have now asked the laboratory to receive these details and will update you on this.

The ab65622 is the only malachite green based kit we have in our catalog currently - this is why I cited this product.

Once again, please apologize for the confusion. We will write you again, as soon as we have more information. Thank you for your patience.

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Answer

Thank you for contacting us.

I have found out the the buffer listed on the datasheet is incorrect. The buffer supplied does not have any sodium phosphate included and is instead:



20mM Tris,50mM KCl,5mM MgCl2,1mM DTT, pH7.5.



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!
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Question
Answer

Thank you for contacting us. The His tags for all four proteins you asked about are all 6X tags. I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.  

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