A flow cytometer is a medical technology device that counts and analyzes cells one at a time as they move through a narrow fluid stream. It is important in blood testing, cancer diagnosis, immunology, and research because it can measure thousands of cells per second. Instead of looking at cells slowly under a microscope, the instrument turns cell properties into light signals and data.
This lets clinicians and scientists identify different cell types with speed and precision.
Inside the instrument, cells are focused into single file by sheath fluid and pass through a laser beam. Each cell scatters light and may also give off fluorescence if it has been tagged with special markers. Detectors measure these signals, electronics convert them into counts and graphs, and software separates groups of cells by size, structure, or marker expression.
Some flow cytometers can also sort cells by giving droplets an electric charge and deflecting selected cells into different containers.
Key Facts
- Hydrodynamic focusing uses sheath fluid to align cells so they pass the laser one at a time.
- Forward scatter, often called FSC, is mainly related to cell size.
- Side scatter, often called SSC, is mainly related to internal complexity or granularity.
- Fluorescence signal occurs when a tagged molecule absorbs laser light and emits light at a longer wavelength.
- Event rate = number of detected cells / time, such as cells per second.
- Concentration = counted cells / sample volume, if dilution and collection efficiency are known.
Vocabulary
- Flow cytometer
- A device that measures physical and fluorescent signals from cells as they flow past a laser one at a time.
- Sheath fluid
- A clean surrounding fluid that narrows and centers the sample stream so cells line up single-file.
- Forward scatter
- Light scattered mostly in the forward direction, commonly used as an indicator of cell size.
- Fluorescence
- Light emitted by a molecule after it absorbs light energy from a laser or other source.
- Cell sorting
- The process of separating selected cells into different containers based on their measured signals.
Common Mistakes to Avoid
- Assuming the cytometer photographs each cell, which is wrong because most flow cytometers measure light signals rather than making detailed images.
- Confusing forward scatter with side scatter, which is wrong because forward scatter is more closely linked to size while side scatter is more closely linked to internal structure.
- Ignoring sample dilution, which is wrong because cell concentration calculations must account for any dilution before the sample entered the instrument.
- Thinking brighter fluorescence always means more cells, which is wrong because brightness usually describes signal per cell while event count describes how many cells were detected.
Practice Questions
- 1 A flow cytometer detects 48,000 cells in 2.0 minutes. What is the event rate in cells per second?
- 2 A sample is diluted 1:5 before testing. The cytometer reports 2.4 x 10^5 cells/mL in the diluted sample. What was the cell concentration in the original sample?
- 3 A cell population has high forward scatter, high side scatter, and strong green fluorescence. Explain what each signal suggests about the cells and how a sorter could use those signals.