Tumor necrosis factor alpha (TNFα) is a potent cytokine involved in immune surveillance and function. Dysregulated TNFα levels have been implicated in chronic diseases such as rheumatoid arthritis, Crohn’s disease, and several neurological diseases.
A number of recombinant anti-TNFα biologicals have been developed and licensed for management of human disease. Simple and predictive bioassays for potency and dose standardization, though, are still sought by the biopharma industry. Automatable, homogeneous bioassays are desirable as they can help streamline this aevaluative process. Implementing predictive automation-friendly bioassays as demonstrated in this tutorial allows effective assessment of anti-TNFα and likely other biologicals for quality control.
Predictability and Productivity
Two predictive bioassays from Promega (www.promega.com) are highlighted in this article. The CellTiter-Glo® Luminescent Cell Viability Assay provides a homogeneous bioluminescent assay for quantitation of ATP, an indicator of metabolically active cells, present inside cells in culture. The assay is designed for use in multiwell plate formats, making it ideal for automated cell viability, cell proliferation, and cytotoxicity studies.
The ONE-Glo™ Luciferase Assay provides a homogeneous bioluminescent assay for quantitation of luciferase expression in mammalian cells containing genes for firefly luciferase. The assay contains a new luciferase substrate, resulting in a reagent that is more stable, more tolerant to sample components, and has less odor than standard luciferase assay reagents, which makes it suitable for high-throughput applications.
Both assays are rapid, simple, and sensitive. The protocols involve adding the reagent directly to cells cultured in growth medium (without washing or preconditioning), mixing, and reading the glowing luminescent signal on a luminometer. For the CellTiter-Glo Assay, the luminescent signal is proportional to the amount of ATP present in the sample, which is directly proportional to the number of cells present. For the ONE-Glo Assay, luminescence is proportional to the upregulation of a specific genetic element and subsequent expression of firefly luciferase.