Cytokines & Growth Factors - Quality
What does Cytokine Quality mean for your research?
To ensure maximum performance of our cytokines we abide by rigorous quality control standards:
- Most products are free of tags or additional amino acids, and many are produced using a mammalian expression system to maximize natural conformation and post-translational modifications.
- ED50 or 50% of maximum response is determined by relevant and standard cell based assays for every lot.
- Most cytokines are greater than 98% pure as demonstrated by SDS-PAGE.
- Several lots are tested side by side to ensure consistent bioactivity and purity.
- Endotoxin levels are tested by the LAL assay and are less than 0.01 ng/µg cytokine.
- Reduced protein is run on SDS-PAGE to determine purity, and non-reduced protein is run to determine presence of cystine-linked dimers.
- Bioactivity and purity data are shown on each product webpage and datasheet.
Purity
The purity of recombinant Human Vascular Endothelial Growth Factor-121 (hVEGF121) #8908 was determined by SDS-PAGE of 6 µg reduced (+) and non-reduced (-) recombinant hVEGF121 and staining overnight with Coomassie Blue. Heterogeneity in SDS-PAGE is due to glycosylation. The non-reduced cystine-linked homodimer migrates as a 30–36 kDa protein.
Bioactivity
The proliferation of HUVEC treated with increasing concentrations of Human Vascular Endothelial Growth Factor-121 (hVEGF121) #8908 was assessed. After 72-hour treatment with hVEGF121 cells were incubated with a tetrazolium salt and the OD450–OD650 was determined.
Western Blot
Western blot analysis of extracts from HUVEC untreated or treated with Human Vascular Endothelial Growth Factor-121 (hVEGF121) #8908 for 15 minutes, using Phospho-p38 MAPK (Thr180/Tyr182) (3D7) Rabbit mAb #9215 (upper) and p38 MAPK Antibody #9212 (lower).
Human Tumor Necrosis Factor-α (hTNF-α)
HeLa cells were serum-starved and then treated with increasing doses of Human Tumor Necrosis Factor-α (hTNF-α) #8902 for 20 minutes. IκBα (L35A5) Mouse mAb (Amino-terminal Antigen) #4814 and Phospho-NF-κB p65 (Ser536) (93H1) Rabbit mAb #3033 were used to assess NF-κB activation. With increasing concentrations of hTNF-α, a decrease in total IκBα (3-fold) and increase in phospho-NF-κB (Ser536) protein (5-fold) was observed, consistent with TNF-α induced degradation of IκBα resulting in activation of NF-κB. The signal for each antibody was analyzed using an Acumen® Explorer and images were acquired with Cellomics ArrayScan® VTI.
Inset: Signal obtained for IκBα, untreated or treated for 20 minutes with 100 ng/ml TNF-α.
The purity of recombinant Human Tumor Necrosis Factor-α (hTNF-α) #8902 was determined by SDS-PAGE of 6 µg reduced (+) and non-reduced (-) recombinant hTNF-α and staining overnight with Coomassie Blue.
The viability of L-929 cells treated with increasing amounts of Human Tumor Necrosis Factor-α (hTNF-α) #8902 in the presence of 2 ng/ml actinomycin D was determined. Cells were stained with crystal violet at the end of treatment and the OD595 was determined.
