Flow cytometry (FC) is an important tool for analyzing complex pathways and responses of single cells, which can track cell phenotypes and functions in multiple dimensions. It is also increasingly applied in drug development. FC not only can detect intracellular and extracellular components, but also detect soluble analytes in serum or plasma samples, such as cytokines, drug complexes, or anti-drug antibodies (ADA).
FC can perform single-cell analysis at a rate of hundreds to tens of thousands of cells per second. Its advantages of high throughput and rapidity are very suitable for large-scale drug development and testing. FC can detect multiple parameters on the cell surface. Coupled with a data analysis system, it can generate a sufficient and complex data stream that excludes false positives in single-parameter tests. In addition, FC can work with a variety of fluorescent dyes and probes, and can have many flexible options. For example, the most advanced mass spectrometry flow cytometer can use a variety of metal element probes.
FC analysis can characterize individual cells in terms of cell surface or internal antigen expression, subcellular components (such as mitochondria), and cell dynamic properties (such as changes in protein expression, calcium flow, membrane potential, and cell proliferation).
Application of flow cytometry in drug development and research
FC has a wide range of applications and participates in all aspects of the entire drug development process, including drug development and target confirmation, non-clinical safety and toxicity evaluation, and clinical research.
FC can simultaneously measure multiple parameters at the single-cell level to gain insight into the complex interaction mechanism (MOA) of the compound and the interaction between sub-cellular populations. In recent years, engineered cell lines (by transfection, siRNA gene knockout, etc.) have been used as models for target verification-related drug development.
These cell lines are designed to stably express recombinant targets for detection. For example, monoclonal antibody drugs that bind to cell target antigens can be used for drug efficacy and PK analysis or to detect neutralizing antibodies that inhibit drug binding.
Cultured cells can also be used for complement-mediated cytotoxicity (CDC) and antibody-dependent cytotoxicity (ADCC) and other tests.
In non-clinical research, FC can be used to evaluate the safety of drugs in vivo and in vitro. In toxicological assessment, flow cytometry provides higher accuracy and lower variability, reducing time, reagents, and animal costs. Flow cytometry has been used to assess drug-induced vascular damage (cell death and circulating endothelial cell test), and to measure changes in T/DC cell activation or protein expression levels on specific cell subsets. In addition to determining toxicity, FC can be used to perform target regulation and functional determination to understand the MOA during drug toxicity evaluation.
Cell functions, such as cytokine production and changes, intracellular signal transduction, and cell proliferation can be measured by FC. Functional assays are also used as potency biomarkers for cancer vaccine potency evaluation.
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