Cell Counting

Access cell viability

Cell viability can be assessed by examining parameters such as cell membrane integrity or the activity of cellular enzymes. On a microplate readers, these parameters can be detected using fluorescent reagents. For example, a red cell-impermeant, DNA-binding dye only stains dead or dying cells whose membranes are compromised, while a green live-cell dye only fluoresces when metabolic enzymes are active.

• Normalize HTRF cytokine assays to cell viability
• Measuring cell viability and cytotoxicity with the EarlyTox Cell Integrity Kit

Alternatives to DAPI staining

DAPI (4’, 6-diamidino-2-phenylindole) is a fluorescent dye often used to stain nuclear DNA. It is employed in imaging experiments such as fluorescent microscopy, chromosome spreads, FACS, and cell-based assays. However, excitation with UV results in a photoconversion of DAPI that leads to detection of DAPI fluorescence in the FITC/GFP channel of an imaging system, causing errors in interpretation of results. DAPI also requires cells to be fixed for maximal staining. Here we show some alternatives to DAPI staining, including StainFree technology, which requires no staining at all and can be used with live cells.

• Alternatives to DAPI staining: imaging and counting live cells
• Count Cells without Cell Staining
• Wells to Westerns: Investigating the cellular heat shock response
• StainFree technology for cellular image analysis without fluorescent dyes

Calculate transfection efficiency

Measuring transfection efficiency provides researchers with an early indication of their gene editing yield before using the modified cells for targeted assays. High quality images can be acquired with both the transmitted light and with a fluorescence channel indicating transfection of the cell. Image analysis software identifies and counts every cell plus scores them as GFP-positive or not. Afterwards, a transfection efficiency may be calculated from the ratio.

• Optimizing GFP transfection with the SpectraMax i3 Multi-Mode Detection Platform
• Validate CRISPR-Edited Cells using Imaging and Western Blot Detection on a Microplate Reader

Cell cycle analysis

Transition of a cell through the cell cycle can be monitored using fluorescently-labeled cell cycle proteins that are cyclically expressed and degraded. FUCCI technology is based on the overexpression the cell cycle-dependent proteins geminin and Cdt1, respectively fused to a green fluorophore and a red fluorophore. Cdt1 levels peak in G1 phase, so cells in G1 appear green; geminin levels rise in late S, G2 and M phase, so cells in these phases appear red.

• Evaluating cell cycle inhibitors using a live cell assay

Cell migration

Cell migration, the movement of cells from one location to another, is a critical component of both normal and abnormal biological processes. The migration of unlabeled or fluorescently labeled cells from one area of a microplate well to another can be monitored using a cellular imaging system. Automated analysis software calculates the amount of cell migration in each well.

• Measure cell migration using a simple scratch assay with timelapse live-cell imaging
• Measure cell migration using discontinuous time-lapse imaging of live cells

Count cells with fluorescent markers

Image and analyze compound effects based on cells staining positive for different fluorescent markers. The consistent and statistically relevant quantitative analysis allows for the testing of multiple compounds at differing concentrations. Read the application note to learn more about the difference between cell counting using fluorescent or transmitted light imaging.

• Count cells with or without fluorescent labels using automated imaging

Counting cells with transmitted light

Transmitted light segmentation (analysis) algorithms increase the accuracy of counting diverse cell types and are typically used to image unstained live or fixed cells. In addition to the standard cell counting methods for detecting fluorescent nuclei or cell bodies. Watch the video to learn how you can set up parameters to run transmitted light cell scoring.

• Transmitted light image analysis for cell count and assessment of cytotoxicity effects
• Improved cell counting method using transmitted light on the SpectraMax MiniMax cytometer
• Transmitted light cell scoring on the ImageXpress Pico
• Counting cells with the Transmitted Light Analysis Module in MetaXpress

Detect cytotoxicity in cells

Cytotoxicity is often measured in response to an experimental treatment or potential drug. Having a way to easily screen cytotoxicity in treated cells is critical to identifying new therapeutic treatments or understanding cellular signaling pathways that affect cell health. Common indicators of cytotoxicity include the ATP level in a cell population and integrity of cellular membranes, both of which can be measured using a variety of microplate-based assays.

• Measuring cell health on the SpectraMax iD3 reader with cell viability assays
• Multiplexed high-content hepatotoxicity assays using iPSC-derived hepatocytes
• Transmitted light image analysis for cell count and assessment of cytotoxicity effects

eBook: Count cells like a pro

Improve cell counts by adding cellular imaging assays in a small footprint and eliminating cell staining and confluence measurements with StainFree™ Cell Detection Technology.

Download eBook 

Image cytometry

Image cytometry is a cell counting technique that combines low magnification microscopy with imaging and analysis tools. Image cytometry is useful for providing information on a broad range of cellular characteristics on a large population of cells in a single assay. Image cytometry can be used on unlabeled or fluorescently labeled cells and can be performed multiple times on a population to provide information about cellular dynamics over time.