Prothrombin Overexpression Lysate

Western Blot: Prothrombin Overexpression Lysate [NBL1-10407] - Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for Prothrombin.

• Reactivity: Hu
• Applications: WB

Prothrombin Overexpression Lysate Summary

• Description:

  Prothrombin Transient Overexpression Lysate

  
  Expression Host: HEK293T
  
  Plasmid: RC208589
  
  Accession#: NM_000506
  
  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: F2

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: 67.5 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 Prothrombin Overexpression Lysate

• coagulation factor II (thrombin)
• Coagulation factor II
• EC 3.4.21
• EC 3.4.21.5
• prothrombin B-chain
• prothrombin
• PT
• serine protease

Background

Hemostasis following tissue injury involves the deployment of essential plasma procoagulants (Prothrombin and Factors X, IX, V and VIII), which mediate a blood coagulation cascade that leads to the formation of insoluble Fibrin clots and the promotion of platelet aggregation. Proteolytic cleavage of Prothrombin (Factor II) at residue 44 leads to formation of Thrombin in the first step of the coagulation cascade. Thrombin cleaves bonds between Arg and Gly and activates Factors V, VII, VIII and XIII in complex with thrombomodulin and Protein C. Thrombin maintains vascular integrity during development and postnatal life and coordinates connective tissue proteins by stimulating fibroblast procollagen production.

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: F2