Certificate of Analysis & Data Sheet

Recombinant ovine leptin, one polypeptide chain containing 146 amino and additional Ala at N-terminus acids and having a molecular mass of ~ 16 kDa. The wild type leptin was mutated at position 4 replacing arginine by cysteine. Recombinant ovine leptin R4C mutant was purified by proprietary chromatographic techniques, see Reicher et al. J. Animal Sciences 90:410-418 (2012).

E. coli

White lyophilized (freeze-dried) powder

The recombinant ovine leptin R4C mutant was lyophilized from a concentrated (1mg/ml) solution with 0.0045mM NaHCO3.

It is recommended to reconstitute the lyophilized recombinant ovine leptin R4C mutant in sterile 0.4% NaHCO3 adjusted, not less than 100µg/ml, which can then be further diluted to other aqueous solutions.

Lyophilized recombinant ovine leptin R4C mutant although stable at room temperature for several weeks, should be stored desiccated below -18C. Upon reconstitution of recombinant ovine leptin at > 0.1 mg/ml and up to 2 mg/ml and filter sterilization recombinant ovine leptin can be stored at 4C or even room temperature for several weeks. At lower concentration of recombinant ovine leptin addition of a carrier protein (0.1% HSA or BSA) is suggested. Please prevent freeze-thaw cycles.

The purity of recombinant ovine leptin R4C mutantis greater than 98.0% as determined by:(a) Gel filtration analysis(b) Analysis by reducing and non-reducing SDS-PAGE gel.

The sequence of the first five N-terminal amino acids of recombinant ovine leptin R4C mutant was determined and was found to be Ala-Val-Pro-Ile-Cys.

The purified recombinant ovine leptin (16K) R4C mutant consists of > 95% monomers as determined by gel-filtration chromatography.

PLR’s recombinant ovine leptin R4C mutant is fully biologically active compared to the wild-type ovine leptin as evidenced by inducing proliferation of BAF/3 cells stably transfected with the long form of human leptin receptor.

Endotoxin content of recombinant ovine leptin is less than 0.05 ng/µg (0.5 IEU/µg) of ovine leptin R4C mutant.

Protein quantitation of recombinant ovine leptin R4C mutant was carried out by UV spectroscopy at 280 nm using the absorbency value of 0.20 as the extinction coefficient for a 0.1% (1mg/ml) solution at pH 8.0. This value is calculated by the PC GENE computer analysis program of protein sequences (IntelliGenetics).

Recombinant ovine leptin R4C is offered by PLR for laboratory research