Recombinant Human Tumor Necrosis Factor-beta
Certificate of Analysis and Data Sheet
Description: Recombinant Human TNF-b produced in E.Coli is a single, non-glycosylated, polypeptide chain containing 171 amino acids and having a molecular mass of 18645 Dalton. The rHuTNF-b is purified by standard chromatographic techniques.
Source: Escherichia Coli.
Physical Appearance: Sterile Filtered White lyophilized (freeze-dried) powder.
Formulation & packaging: Lyophilized protein with no additives.
Solubility: The lyophilized rHuTNF-b is very soluble in water and most aqueous buffers below and above the isoelectric point (pI=7.16).
Stability: Lyophilized rHuTNF-b, although stable at room temperature, should be stored desiccated below 0°C. Reconstituted rHuTNF-b is best stored refrigerated at 4°C.
Purity: Greater than 99.0% 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
human TNF-b.
Amino acid sequence: The sequence of the first five N-terminal amino acids was determined and was found to be Met-Val-Arg-Ser-Ser, conforming the sequence of native human TNF-b.
Dimers and aggregates: Less than 1% as determined by silver-stained SDS-PAGE gel analysis.
Biological Activity: Our rHuTNF-b is fully biologically active when compared to standard. The ED50 as determined by the cytolysis of murine L929 cells in the presence of Actinomycin D is < 0.05ng/ml, corresponding to a Specific Activity of 2.2x107 IU/mg.
Endotoxin: Less than 0.1 ng/µg (IEU/µg) of rHuTNF- b.
Protein content: Protein quantitation was carried out by two independent methods:
1. UV spectroscopy at 280 nm using the absorbency value of 1.08 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 a calibrated solution of TNF-b as a Reference Standard.
Usage: This material is offered for research, laboratory or further manufacturing purposes.
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