iNOS Overexpression Lysate

Western Blot: iNOS Overexpression Lysate (Adult Normal) [NBL1-13721] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for iNOS.

• Reactivity: Hu, Rt
• Applications: WB

iNOS Overexpression Lysate Summary

• Description:

  iNOS Transient Overexpression Lysate

  
  Expression Host: HEK293T
  
  Plasmid: RC211819
  
  Accession#: NM_000625
  
  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.
• Gene: NOS2

Applications/Dilutions

• Dilutions:
  • Western Blot
• 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.
• Theoretical MW: 131 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.

Packaging, Storage & Formulations

• Storage: Store at -80C. Avoid freeze-thaw cycles.
• Buffer: RIPA buffer

Lysate Details for Array

• Type: Overexpression

Notes

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.

Alternate Names for iNOS Overexpression Lysate

• EC 1.14.13.39
• Hepatocyte NOS
• HEP-NOS
• Inducible NO synthase
• Inducible NOS
• iNOS
• nitric oxide synthase 2, inducible
• nitric oxide synthase 2A (inducible, hepatocytes)
• nitric oxide synthase, inducible
• nitric oxide synthase, macrophage
• NOS type II
• NOS
• NOS2
• NOS2A
• Peptidyl-cysteine S-nitrosylase NOS2

Background

Nitric oxide (NO) is a colorless, free radical gas that carries a variety of messages between cells. Vasorelaxation, neurotransmission and cytotoxicity can all be potentiated through cellular response to NO. NO production is mediated by members of the nitric oxide synthase (NOS) family including the two constitutive isoforms: brain, bNOS, or neuronal NOS, nNOS (type I) and endothelial cell NOS, eNOS (type III); along with the inducible isoform, iNOS (type II). NOS catalyzes the oxidization of L-arginine to produce L-citrulline and NO, requiring the cofactors calmodulin, nicotinamide adenine dinucleotide phosphate (NADPH), flavin adenine dinucleotide (FAD), and flavin mononucleotide (FMN), heme, and tetrahydrobiopterin (1).

The 131 kDa enzyme, iNOS, is found in a variety of cell types including macrophages, hepatocytes, synoviocytes, and smooth muscle cells. While constitutively expressed in kidneys, in other tissues iNOS is induced by bacterial lipopolysaccharides (LPS), growth factors, and cytokines such as IFN-gamma, TNF, IL-1 and IL-2. iNOS is not regulated by the level of intracellular Ca2+ and is constantly active as a dimer when expressed. iNOS activity is elevated in a variety of diseases including atherosclerosis, heart failure, sepsis, solid tumors, and type 2 diabetes. Acting as a critical mediator of inflammation and apoptosis, iNOS inhibitors have been shown to alleviate obesity and stress inducted insulin resistance in mouse models (2,3).

References

1. Forstermann U, and Sessa WC. (2012) Nitric oxide synthases: regulation and function. Eur Heart J. 33(7): 829-837. PMID: 21890489

2. Aktan F. (2004) iNOS-mediated nitric oxide production and its regulation. Life Sci. 75(6):639-53. PMID: 15172174

3. Cinelli MA, Do HT, Miley GP, Silverman RB. (2020) Inducible nitric oxide synthase: Regulation, structure, and inhibition. Med Res Rev. 40(1):158-189. PMID: 31192483

Limitations

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.

Publications for iNOS Lysate (NBL1-13721)(1)

Publication using NBL1-13721

• Sayed WM, Elzainy A Impact of platelet-rich plasma versus selenium in ameliorating induced toxicity in rat testis: histological, immunohistochemical, and molecular study Cell and tissue research 2021-04-01 [PMID: 33791879] (WB, Rat)

FAQs for iNOS Lysate (NBL1-13721). (Showing 1 - 1 of 1 FAQ).

1. We need to evaluate the level of iNOS in Rat sciatic nerve by Western Blot. I see that you have few antibodies suitable for that, could you please recommend the best what you have for our purposes?
   • We have 4 iNOS antibodies that are suitable for your experiment. The best choice for you will depend on your specific preferences, in this case the main difference will be the immunogen location. If the immunogen is not important to you, then I would recommend choosing NB300-605 as it gives you the best value and we are able to provide the exact immunogen.

Bioinformatics

• Gene Symbol: NOS2