Advanced Flow Cytometry Solutions for Targeted Drug Discovery
Harness the power of MACSQuant® X Flow Cytometry at 2bind to gain deep insights into cellular characteristics and interactions critical for your drug discovery research. Our team of scientists specializes in flow cytometry applications, particularly in lead selection and profiling. Enhanced screening methods for candidate binding to target cells facilitate the identification of promising drug candidates, while titrations provide a means to rank these candidates effectively. By analyzing binding profiles of candidates within diverse cell populations, we can discover drug candidates with unique specificities. Additionally, functional cell-based assays are applied to determine the potency and efficacy of drug molecules.
Our customized solutions and expert data interpretation, paired with cutting-edge instrumentation like the MACSQuant® X Flow Cytometer, ensures high-quality data and actionable results. Choose 2bind for faster turnaround times, superior sensitivity, and unparalleled support at every step.
Flow cytometry is a cutting-edge laser-based technology that swiftly analyzes individual cells in a fluid stream. Each cell undergoes scrutiny for various parameters, including size, granularity, and fluorescence detection. This method is utilized to examine individual cells for specific antigens by staining them with fluorescently labeled antibodies. Moreover, flow cytometry serves as a tool to evaluate antibody drug candidates for their binding capabilities to target cells. Our preferred instrument, the MACSQuant® X, boasts multi-parametric cell analysis and revolutionizes drug discovery research. Its high-speed and automated measurement mode, coupled with advanced laser and filter components, enables extensive screenings and multiplexed applications to pinpoint cell binders with unique specificities.
MACSQuant® X allows to:
Mean fluorescent intensity is calculated for each cell population after staining with one or more antibodies. In addition to the MFIs, histograms of the stained cells are provided and give insights into the heterogeneity of the stained population.
Flow cytometry after staining of cells with one or more target-specific antibodies allows the identification of subpopulations within a cell preparation that express the target of interest or a combination of cell surface receptors.
Determine potency and efficacy in functional assays according to the drug’s mechanism of action.
Dose-response data. Analyte concentration that gives half-maximal response in cell binding assays. The EC50 is dependent on the analyte’s affinity. For a more precise affinity measurement including kinetic information, please consider the RT-IC technology.
2bind is your ideal partner for flow cytometry-based research. Our expertise in cell-based assays, coupled with our commitment to personalized service and rigorous quality control, ensures that your projects are in expert hands. We go beyond data generation; we help you interpret results and translate them into actionable insights, accelerating your path to drug discovery success.
Flow cytometry with the MACSQuant® X is a versatile tool, adaptable to a wide range of drug discovery applications:
Biotech: Streamline early-stage drug candidate screening and optimize lead compound selection.
Pharmaceutical: Validate drug targets, assess drug efficacy, and gain mechanistic insights into drug action.
Academic Research: Explore cellular heterogeneity in disease models, characterize immune responses, and delve into fundamental cell biology.
Flow cytometry is a powerful analytical technique used to measure the physical and chemical characteristics of individual cells or particles as they pass through a laser beam. The basic principles of flow cytometry include the following key components:
Fluidics
The fluidics system transports cells in a stream so that they pass one by one through a laser beam. Cells are typically suspended in a sheath fluid, which hydrodynamically focuses the cells into a narrow stream. This ensures that each cell passes through the interrogation point (laser beam) individually.
Optics
The optics system includes lasers and detectors. Lasers provide the light source that excites the fluorophores bound to the cells. Different lasers (e.g., violet, blue, red) are used to excite different fluorophores.
As each cell passes through the laser beam, it scatters light and emits fluorescence. The scattered light and emitted fluorescence are collected by a series of detectors.
Detectors
There are two main types of light detected in flow cytometry:
Fluorescence Detectors: These detectors capture the emitted fluorescence from fluorophores tagged to specific cell components. Multiple detectors can be used to measure different wavelengths of fluorescence, allowing for the analysis of multiple parameters simultaneously.
Fluorescence
Fluorophores are fluorescent molecules that can be conjugated to antibodies or other molecules that bind specifically to cellular components (e.g., proteins, nucleic acids). When these fluorophores are excited by the laser, they emit light at a longer wavelength.
The emitted light is collected by the fluorescence detectors and filtered to isolate the specific wavelengths corresponding to each fluorophore. This allows for the identification and quantification of various cellular components.
Electronics
The signals from the detectors are converted into electronic signals. These signals are then processed by a computer to generate data. The data can be displayed as histograms or dot plots, where each point represents an individual cell or particle.
The data analysis software allows for the gating and analysis of different cell populations based on their size, complexity, and fluorescence characteristics.
Data Analysis
The data collected from flow cytometry can be analyzed to determine various characteristics of the cells, such as cell size, granularity, and the presence of specific markers.
Complex analyses can be performed to study cell populations, identify subsets of cells, and measure functional properties (e.g., cell cycle analysis, apoptosis, cytokine production).
In summary, flow cytometry allows for the rapid and quantitative analysis of multiple parameters on a single-cell basis. It is widely used in research and clinical laboratories for applications such as immunophenotyping, cell sorting, apoptosis detection, and intracellular signaling studies.
Flow cytometry can be used with a wide variety of cell types, depending on the specific application and the goals of the analysis. Here are some common types of cells that are often used in flow cytometry:
Peripheral Blood Mononuclear Cells (PBMCs)
These are blood cells with a single round nucleus, including lymphocytes (T cells, B cells, NK cells), monocytes, and dendritic cells.
Commonly used in immunology, infectious disease studies, and cancer research.
Cultured Cell Lines
These are cells that have been adapted to grow continuously in culture. Examples include HeLa cells, HEK293 cells, Jurkat cells, and CHO cells.
Used extensively in drug discovery, cell biology, and cancer research.
Stem Cells
Includes embryonic stem cells, induced pluripotent stem cells (iPSCs), and various types of adult stem cells (e.g., hematopoietic stem cells).
Important for regenerative medicine, developmental biology, and tissue engineering studies.
Primary Cells
These are cells directly isolated from tissues, such as primary hepatocytes, neurons, or fibroblasts.
Used to study cell behavior in a more physiologically relevant context compared to immortalized cell lines.
Tumor Cells
Includes cells isolated from solid tumors or blood cancers (e.g., leukemia, lymphoma).
Used for cancer research, including studying tumor immunology, cell signaling, and drug responses.
Immune Cells
Specific populations of immune cells, such as T cells, B cells, NK cells, macrophages, and dendritic cells.
Crucial for studies on immune responses, vaccine development, and autoimmune diseases.
Bacterial Cells
Certain types of bacteria can be analyzed using flow cytometry, often for research in microbiology, infection, and antibiotic resistance.
Yeast Cells
Used in research involving genetics, cell cycle studies, and protein expression.
Plant Cells
Flow cytometry can be used to study plant cell populations, particularly in research involving genetics and plant development.
Sperm Cells
Used in reproductive biology and for assessing sperm viability and motility.
Platelets: Although not nucleated, platelets can be analyzed for studies related to coagulation, thrombosis, and cardiovascular diseases.
Each type of cell may require specific preparation and staining protocols to ensure optimal results in flow cytometry analysis. The choice of cell type and preparation method will depend on the specific research question and the characteristics of the cells being studied.
2bind’s MACSQuant®X is equipped with three lasers and five filters to excite fluorophores at 405, 488 or 638 nm and to detect the emitted fluorescence at 450, 525, 585, 655 and 750 nm. Combinations of lasers and filters results in eight detection channels that cover most of the commonly used commercially available fluorophores. Staining with combinations of fluorophores are possible enabling combinatorial analysis or target expression and sophisticated gating strategies to visualize binding of antibodies to certain subpopulations within the cell preparation.
Fluorophores Excited by a 405 nm Laser (Violet)
Pacific Blue
Brilliant Violet 421 (BV421)
Alexa Fluor 405
DAPI (used for DNA staining)
Fluorophores Excited by a 488 nm Laser (Blue)
FITC (Fluorescein Isothiocyanate)
PE (Phycoerythrin)
PerCP (Peridinin Chlorophyll Protein Complex)
PerCP-Cy5.5
Alexa Fluor 488
GFP (Green Fluorescent Protein)
Fluorophores Excited by a 638 nm Laser (Red)
APC (Allophycocyanin)
Alexa Fluor 647
APC-Cy7
APC-H7
Testing for cell-binding is a crucial part of the therapeutic antibody discovery process, when the target is a membrane protein. Antibodies are typically generated by immunizing with recombinant extracellular domains of the target protein. These antibodies are often first identified through ELISA studies using the same extracellular domains. It is essential to confirm binding to the native protein on target cells to ensure that the antibodies do not recognize artificial structures present in the recombinant antigen.
Binding to target cells and the lack of binding to non-target cells are critical criteria thoroughly studied before selecting a lead drug candidate. Typically, flow cytometry is used to screen candidates against a panel of cells. Beyond binding tests at a single concentration, analytes can be titrated to determine EC50 values for the binding interaction with each cell line.
In addition to cell-binding, flow cytometry can be utilized for more advanced cell-based assays to study the functional effects of drug candidates on cells. The most common example is apoptosis assays, where Annexin V is used as a marker for apoptotic cells. However, potential mechanisms of action for therapeutic drugs are virtually unlimited, and in many cases, flow cytometry is used to determine the potency and efficacy of drug candidates. For example, drugs that act on GPCRs are frequently screened in calcium flux assays to detect agonists that stimulate a pathway or antagonists that block the signal cascade. Other drug candidates affect the cell cycle or cell differentiation, which is another application of flow cytometry-based assays.
