Recombinant Ovine Leptin (Anti-Obesity Protein)

 {rOLeptin (AOBP)}

Certificate of Analysis and Data Sheet

 

       ×         Description:

Recombinant Ovine Leptin produced in E.Coli is a single, non-glycosylated, polypeptide chain containing 146 amino acids and having a molecular mass of 16,130 Dalton.

          The rOLeptin is purified by proprietary chromatographic techniques.

 

       ×         Source:

Escherichia Coli.

 

       ×         Physical Appearance:

Sterile Filtered White lyophilized (freeze-dried) powder.

 

       ×         Formulation & Packaging:

The protein was lyophilized from a concentrated (1mg/ml) solution with 0.0045mM NaHCO3.

 

       ×         Solubility:

The lyophilized rOLeptin is very soluble in water and most aqueous buffers below and above the isoelectric point.

 

       ×         Stability:

Lyophilized rOLeptin although stable at room temperature, should be stored desiccated below 0°C. Reconstituted rOLeptin is best stored refrigerated at 4°C.

 

       ×         Purity:

Greater than 95.0% as determined by:

 (a) Analysis by RP-HPLC.

           (b) Anion-exchange FPLC.

         (c) Analysis by reducing and non-reducing SDS-PAGE Silver Stained.

 

(Limit of acceptance:³98.0%. No more than 2% total impurities; no single impurity greater than 1%)

 

       ×         Amino Acid Composition:

In total agreement with the expected amino acid composition of native Ovine Leptin.

 

       ×         Amino acid sequence:

The sequence of the first five N-terminal amino acids was determined and was found to be Ala-Val-Pro-Ile-Arg, which agrees with the sequence of native Ovine Leptin.

 

       ×         Dimers and aggregates:

Less than 1% as determined by silver-stained SDS-PAGE gel analysis.

 

       ×         Biological Activity:

Our rOLeptin is fully biologically active when compared to standards.

  The ED50,  calculated by the leptin-dependant stimulation of  Human OB-R transfected murine BaF3 indicator cells  is 0.33-0.05 ng/ml.

 

       ×         Endotoxin:

Less than 0.1 ng/µg (IEU/µg) of rOLeptin.

 

       ×         Protein content:

          Protein quantitation was carried out by two independent methods:

 

1.      UV spectroscopy at 280 nm using the absorbency value of 0.2 as the extinction

               coefficient for a 0.1% (1mg/ml) solution. This value is calculated by the PC GENE

     computer  analysis program of protein sequences (IntelliGenetics).

          2.  Analysis by RP-HPLC,using  calibrated solution of Ovine Leptin as a Reference Standard.

 

      ×        Usage:

This material is offered for research, laboratory or further manufacturing purposes.