NSF Overexpression Lysate

Western Blot: NSF Overexpression Lysate (Adult Normal) [NBL1-13808] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for NSF.

• Reactivity: Hu
• Applications: WB

NSF Overexpression Lysate Summary

• Description:

  NSF Transient Overexpression Lysate

  
  Expression Host: HEK293T
  
  Plasmid: RC205983
  
  Accession#: NM_006178
  
  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: NSF

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: 82.4 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 NSF Overexpression Lysate

• EC 3.6.4.6
• NEM-sensitive fusion protein
• N-ethylmaleimide-sensitive factor
• N-ethylmaleimide-sensitive factor-like protein
• N-ethylmaleimide-sensitive fusion protein
• SKD2
• vesicle-fusing ATPase
• Vesicular-fusion protein NSF

Background

N ethylmaleimide sensitive factor (NSF) is an ~83 kDa protein (predicted MW) which was initially isolated based on its ability to restore intercisternal Golgi transport to N ethylmaleimide treated membranes. NSF was subsequently discovered to be involved in a variety of membrane fusion events. NSF is an ATPase with two ATP binding domains that are essential for membrane fusion and homo oligomerization. In order to interact with target membranes, NSF requires another set of soluble proteins called Soluble NSF attachment proteins (SNAPs). NSF has been shown to play a prominent role in synaptic vesicle fusion. Together with synaptotagmin and a SNAP, NSF modulates the interaction of SNAP25, VAMP, and syntaxin, in an ATP dependent manner, to form a complex with a sedimentation coefficient of 20 S. This complex may represent an intermediate involved in synaptic vesicle docking and fusion.

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.

Bioinformatics

• Gene Symbol: NSF