NLRP3/NALP3 Overexpression Lysate

Name: NLRP3/NALP3 Overexpression Lysate
Brand: Novus Biologicals
SKU: NBL1-13675

Novus Biologicals, part of Bio-Techne | Catalog # NBL1-13675

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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 NLRP3/NALP3 Overexpression Lysate

NLRP3/NALP3 Transient Overexpression Lysate
Expression Host: HEK293T

Plasmid: RC220952

Accession#: NM_004895

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

118 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 NLRP3/NALP3 Overexpression Lysate

Western Blot: NLRP3/NALP3 Overexpression Lysate [NBL1-13675]

Western Blot: NALP3 Overexpression Lysate (Adult Normal) [NBL1-13675] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for NALP3.

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: NLRP3/NALP3

NACHT, LRR and PYD domains-containing protein 3 (NALP3), Nucleotide-Binding Oligomerization Domain, or Leucine Rich Repeat and Pyrin Domain Containing 3 (NLRP3) (human NLRP3-isoform 2 theoretical molecular weight 118kDa) acts as a cytosolic receptor for stress-signals initiated by a wide variety of pathogen-associated molecular patterns (PAMPS) and danger-associated molecular patterns (DAMPs) (1). NLRP3 belongs to the family of NOD-like receptors, a type of pattern-recognition receptor (PRR) which participates in innate immune responses. The exact mechanisms leading to NLRP3 activation are still not fully resolved. Some proposed mechanisms for NLRP3 inflammasome activation include induced ion channel flux as exemplified by potassium efflux, sodium influx and calcium signaling, generation of mitochondrial reactive oxygen species (ROS), and lysosomal destabilization (2). Activation of NLRP3 induces its association with the adaptor protein Apoptosis-associated Speck-like protein containing CARD (ASC) forming a complex that binds to Caspase-1 (1, 2). The inflammasome complex, comprised of NLRP3, ASC and Caspase 1, induces the processing of pro-IL-1beta and pro-IL-18 and release of the mature inflammatory cytokines IL-1beta and IL-18 and pyroptosis (1-3).

NLRP3 is expressed in a variety of cell types such as monocytes, dendritic cells, lymphocytes and epithelial cells (3). Abnormal NLRP3 activation may occur as the result of inherited mutations and is associated with diseases such as hereditary periodic fevers (HPFs) and familial cold autoinflammatory syndrome (FCAS). Additionally, abnormal NLRP3 activation is associated with a variety of diseases such as gout, obesity, atherosclerosis, Alzheimers, multiple sclerosis and type 2 diabetes (1, 3). NLRP3 inflammasome is regulated by several post-translational modifications (e.g., ubiquitination, phosphorylation and sumoylation) as well as by various interacting proteins (e.g., JNK1, FBXO3, TXNIP, MARK4 and PKD) (4).

References

1. Abderrazak, A., Syrovets, T., Couchie, D., El Hadri, K., Friguet, B., Simmet, T., & Rouis, M. (2015). NLRP3 inflammasome: From a danger signal sensor to a regulatory node of oxidative stress and inflammatory diseases. Redox Biology. https://doi.org/10.1016/j.redox.2015.01.008

2. Yang, Y., Wang, H., Kouadir, M., Song, H., & Shi, F. (2019). Recent advances in the mechanisms of NLRP3 inflammasome activation and its inhibitors. Cell Death and Disease. https://doi.org/10.1038/s41419-019-1413-8

3. Zahid, A., Li, B., Kombe, A. J. K., Jin, T., & Tao, J. (2019). Pharmacological inhibitors of the nlrp3 inflammasome. Frontiers in Immunology. https://doi.org/10.3389/fimmu.2019.02538

4. Kelley, N., Jeltema, D., Duan, Y., & He, Y. (2019). The NLRP3 inflammasome: An overview of mechanisms of activation and regulation. International Journal of Molecular Sciences. https://doi.org/10.3390/ijms20133328

Long Name

NLR Family, Pyrin Domain Containing 2

Alternate Names

AGTAVPRL, AII, CIAS1, Cryopyrin, FCAS, FCU, MWS, NACHT, NALP3, PYPAF1

Gene Symbol

NLRP3

Additional NLRP3/NALP3 Products

Product Documents for NLRP3/NALP3 Overexpression Lysate

Product Datasheets

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COA

SDS

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Product Specific Notices for NLRP3/NALP3 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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