Confluency refers to the percentage of the surface area in a cell culture vessel that is covered by adherent cells. In other words, it is a measure of how crowded or dense the cells are within the culture vessel. Confluency is a crucial parameter in cell culture, as it influences cell growth, morphology, behavior, and response to various treatments or experimental conditions.

When cells are initially seeded in a culture vessel, they attach to the surface and begin to proliferate, gradually occupying more of the available surface area. As the cells divide and spread, they eventually reach a point where they cover the entire surface and are in close contact with neighboring cells. This stage is known as 100% confluency or complete confluency.

Depending on the cell type and experimental requirements, researchers may choose to passage or subculture the cells at different confluency levels. For example, some cell types may require passaging at 70-80% confluency to maintain optimal growth and health, while others may tolerate higher confluency levels before being passaged.

Maintaining appropriate confluency levels is important for several reasons:

1. Cell growth and proliferation: As adherent cells become more confluent, they receive inhibitory signals from neighboring cells, leading to contact inhibition and a decrease in their growth rate. Overly confluent cultures may become growth-arrested or experience changes in morphology and function.
2. Nutrient availability: Cells in a highly confluent culture may deplete nutrients and produce waste products more quickly, leading to changes in the culture conditions and potentially impacting cell health.
3. Experimental consistency: Maintaining consistent confluency levels across different samples or time points can help ensure reproducibility and comparability of experimental results.
4. Cellular behavior and response: Cells at different confluency levels may exhibit varying behaviors or responses to treatments, such as differences in drug sensitivity or gene expression patterns.

To assess confluency, researchers typically use an inverted microscope to visually inspect the cells and estimate the percentage of the surface area covered by the cells. Alternatively, automated cell counting systems or image analysis software can be employed to quantify confluency more accurately.