Certificate of Analysis & Data Sheet

Recombinant human leptin is one polypeptide chain containing 146 amino and additional Ala at N-terminus acids and having a molecular mass of ~ 16 kDa. Recombinant human leptin was mutated, resulting in L39A/D40A/F41A mutant that was purified by proprietary chromatographic techniques. Preparation of leptin antagonists was recently published in Biochemical Journal (Niv-Spector et al, Biochem. J 291;221-230 (2005).

Escherichia coli.

White lyophilized (freeze-dried) powder.

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

It is recommended to reconstitute the lyophilized recombinant human leptin antagonist in sterile 0.4% NaHCO3 adjusted, not less than 100µg/ml, which can then be further diluted to other aqueous solutions preferably in presence of carrier protein.

Lyophilized recombinant human leptin antagonist although stable at room temperature for several weeks, should be stored desiccated below -18C. Upon reconstitution at of recombinant human leptin antagonist > 0.1mg/ml and up to 2 mg/ml and filter sterilization human recombinant leptin antagonist can be stored at +4C or even room temperature for several weeks making it suitable for long term infusion studies using osmotic pumps. At lower concentration of recombinant human leptin antagonist addition of a carrier protein (0.1% HSA or BSA) is suggested. Please prevent freeze-thaw cycles.

The purity of recombinant human leptin antagonist is 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 human leptin antagonist was determined and was found to be Ala-Val-Pro-Ile-Gln

The purified recombinant human leptin antagonist (16K) consists of > 97% monomers as determined by gel-filtration chromatography.

PLR’s recombinant human leptin antagonist is capable of inhibiting leptin-induced proliferation of BAF/3 cells stably transfected with the long form of human leptin receptor. It also inhibits various leptin effects in several in vitro bioassays.

Less than 0.1 ng/µg (IEU/µg) of recombinant human leptin antagonist.

Protein quantitation of recombinant human leptin antagonist was carried out by UV spectroscopy at 280 nm using the absorbency value of 0.88 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).

PLR's recombinant human leptin antagonist is furnished for LABORATORY RESEARCH USE ONLY. The product may not be used as drugs, agricultural or pesticidal products, food additives, local application or household chemicals. It is for Research Purposes Only – Commercial Use has been Licensed Exclusively to a Third Party.