GRP78/HSPA5 Overexpression Lysate

Name: GRP78/HSPA5 Overexpression Lysate
Brand: Novus Biologicals
SKU: NBP2-10471

Novus Biologicals, part of Bio-Techne | Catalog # NBP2-10471

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Product Summary
Product Specifications
Scientific Data
Preparation & Storage
Background
Product Documents
Specific Notices

Key Product Details

Species

Human

Applications

Western Blot

Product Summary for GRP78/HSPA5 Overexpression Lysate

GRP78/HSPA5 Transient Overexpression Lysate
Expression Host: HEK293T

Plasmid: RC205859

Accession#: NM_005347

Protein Tag: C-MYC/DDK

You will receive 1 vial of lysate (100ug), 1 vial of empty vector negative control (100ug), and 1 vial of 2xSDS sample buffer (250ul). Each vial of cell lysate contains 100ug of total protein (at 1 mg/ml). The 2xSDS Sample Buffer consists of 4% SDS, 125mM Tris-HCl pH6.8, 10% Glycerol, 0.002% Bromophenol blue, 100mM DTT.

Product Specifications

Application Notes

This product is intended for use as a positive control in Western Blot. Overexpression of the target protein was confirmed using an antibody to DDK (FLAG) epitope tag (NBP1-71705) present on the protein construct.

Each vial of cell lysate contains 100ug of total protein which should be sufficient for 20-50 reactions. Depending on over-expression level, antibody affinity and detection system, some lysates can go as low as 0.1 ug per load. We recommend starting with 5ug of cell lysate. Add an equal amount of cell lysate and 2X SDS Sample buffer and boil the SDS samples for 10 minutes before loading.

TMW

72.2 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.

Type

Overexpression

Scientific Data Images for GRP78/HSPA5 Overexpression Lysate

Western Blot: GRP78/HSPA5 Overexpression Lysate [NBP2-10471]

Western Blot: GRP78 Overexpression Lysate (Adult Normal) [NBP2-10471] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for GRP78.

Formulation, Preparation, and Storage

Formulation

RIPA buffer

Concentration

The exact concentration of the protein of interest cannot be determined for overexpression lysates. Please contact technical support for more information.

Shipping

The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below.

Storage

Store at -80C. Avoid freeze-thaw cycles.

Background: GRP78/HSPA5

Within the ER, misfolded proteins are detected by Bip (GPR78). In the presence of misfolded proteins, GPR78 disscociates from PERK, allowing it to homodimerize and autophosphoyate. In its dimerized phosphorylated form, PERK phosphorylates eIF2. The phosphorylation of eIF2 blocks the majority of translation to prevent the continued accumulation of protein in the ER. GRP78 also binds to IRE1 and ATF6 under normal ER conditions, but dissociates upon accumulation of misfolded proteins. Unbound ATF6 is translocated to the gogli where it is converted into a cleaved, active form. The cleaved form of ATF6 is then able to up regulate transcription of UPR genes including XBP1. Activated IRE1 acts as an endoribonuclease to an intron from XBP1 transcripts. The XBP1 splice variant codes for an active transcription factor which activates transcription of P58IPK. P58IPK is a HSP40 protein which binds to and inhibits PERK. Thus, if the accumulated misfolded proteins have been removed, P58IPK acts to shut down the UPR by removing the block to translation.

Long Name

75 KD Glucose Regulated Protein

Alternate Names

BIP, HSP70-5, HSPA5

Gene Symbol

HSPA5

Additional GRP78/HSPA5 Products

Product Documents for GRP78/HSPA5 Overexpression Lysate

Product Datasheets

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COA

SDS

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Product Specific Notices for GRP78/HSPA5 Overexpression Lysate

HEK293T cells in 10-cm dishes were transiently transfected with a non-lipid polymer transfection reagent specially designed and manufactured for large volume DNA transfection. Transfected cells were cultured for 48hrs before collection. The cells were lysed in modified RIPA buffer (25mM Tris-HCl pH7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 1xProteinase inhibitor cocktail mix, 1mM PMSF and 1mM Na3VO4, and then centrifuged to clarify the lysate. Protein concentration was measured by BCA protein assay kit.

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Lysates are guaranteed for 6 months from date of receipt.

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