Ranibizumab Bioassay Kit

• Cat.No.: Kit-1612
• Product Overview: These assays utilize Enzyme Fragment Complementation (EFC) technology, where the b-galactosidase (b-gal) enzyme is split into two fragments: ProLink (PK) and Enzyme Acceptor (EA). Independently these fragments have no b-gal activity; however, when forced to complement through protein-protein interactions, they form an active b-gal enzyme.
  The Ranibizumab Bioassay can be used to detect ligand-induced dimerization of two subunits of a receptor-dimer pair. This assay detects VEGF-Ainduced homodimeriziaton of the kinase insert domain (KDR), also known as VEGFR2 receptor. The cells have been engineered to co-express KDR fused to PK, and another KDR construct fused to EA. Activation of the KDR receptor through VEGF-A leads to receptor dimerization, which is an essential event in the receptor's signaling cascade. Receptor dimerization forces the two enzyme fragments to complement, resulting in the formation of a functional b-gal enzyme. The enzyme hydrolyzes a substrate to generate a chemiluminescent signal. Ranibizumab binds to and inactivates the VEGF-A protein, preventing VEGF from activating the VEGFR2 receptors. This leads to an inhibition of the signaling event and therefore a reduction in signal.

Specification

• Storage: HEK 293 KDR/KDR Bioassay Cells:
  Cells are shipped on dry ice and should arrive in a frozen state. To ensure maximum cell viability, thaw the vials as soon as possible upon receipt.
  If continued storage of the frozen vials is necessary, store as follows: ƒ
  Short term (24 hours or less): Store vials at -80°C immediately upon arrival. (DO NOT store at -80°C for more than 24 hours). ƒ
  Long term (greater than 24 hours): Vials should ONLY be stored in the vapor phase of liquid nitrogen (LN2).
  
  Bioassay Detection Kit
  Store at -20°C. Once thawed, the detection reagents can be kept at 4°C for up to 4 days. For longer storage (up to the expiration date listed on the kit certificate of analysis), the reagent should be aliquoted and stored at -20°C until needed. Avoid multiple freeze-thaw cycles.
  To make aliquots suitable for testing one assay plate each, 1mL of Detection Reagent 1 per aliquot can be dispensed and frozen down. 4mL of Detection Reagent 2 per aliquot can be dispensed and frozen down separately. Do not mix the two reagents during aliquoting.
  
  AssayComplete Cell Plating Reagent 0
  Once thawed, the Cell Plating Reagent can be stored at 4°C for up to 4 weeks. For longer storage (up to the expiration date listed on the kit certificate of analysis), the reagent should be aliquoted and stored at -20°C until needed. Avoid multiple freeze-thaw cycles. To make aliquots suitable for testing one assay plate each, 30mL of reagent per aliquot can be dispensed and frozen down.
  
  Protein Dilution Buffer
  Once thawed, the Protein Dilution Buffer can be stored at 4°C for up to 4 weeks. For longer storage (up to the expiration date listed on the kit certificate of analysis), the reagent should be aliquoted and stored at -20°C until needed. Avoid multiple freeze-thaw cycles. To make aliquots suitable for testing one assay plate each, 10 mL of reagent per aliquot should be dispensed and frozen down. This amount may vary depending on your stock sample concentrations and should be adjusted accordingly.
  
  Recombinant Human VEGF165 Control Agonist
  Store at -20°C until ready to use (up to the expiration date listed on the kit certificate of analysis). Centrifuge the vial prior to opening to maximize recovery. Reconstitute to a concentration of 100 μg/mL by adding 100 μL of Protein Dilution Buffer. Reconstituted ligand is stable for 12 months at -20 to -80°C, or 1 week at 2-8 °C.
  
  96-well Tissue Culture Treated Plates
  Store at room temperature.