Impedance flow cytometry is a method for analyzing individual cells suspended in a fluid using electrical measurements. This technique counts and assesses the physical state of cells by analyzing their intrinsic electrical properties, providing a direct way to quantify and characterize cell populations.
The Core Principle of Operation
The core concept of impedance flow cytometry is the Coulter principle. This principle states that when a particle, like a biological cell, in a conductive solution passes through a small aperture, it displaces some of the solution. This displacement causes a measurable change in electrical impedance, and the magnitude of this change is directly proportional to the particle’s volume.
Modern instruments use microfluidic channels to force cells into a single-file line through a detection zone. Microelectrodes on either side of this channel generate a stable electric current. As each cell passes between the electrodes, it disrupts the current and creates an impedance signal that the system’s electronics detect and record.
The process is analogous to how a large stone displaces more water than a small pebble. A larger cell displaces more conductive fluid, causing a greater increase in impedance compared to a smaller cell. This allows the instrument to determine the size of each cell, and the technology is sensitive enough to differentiate between cell types based on their characteristic volumes.
This measurement is captured rapidly, with thousands of cells per second flowing through the microfluidic chip. The high-throughput analysis generates a detailed size distribution profile of the entire cell population, providing a snapshot of the sample’s physical composition.
Cellular Properties Measured by Impedance
The signals from impedance flow cytometry can reveal more than just cell size. By using electrical currents of different frequencies, the technology probes distinct biophysical properties. The interaction between the cell and the electric field changes depending on the frequency, providing insights into the cell’s structural integrity and internal composition.
At low frequencies, in the kilohertz (kHz) range, the alternating current (AC) cannot breach the cell’s insulating lipid membrane and is forced to flow around it. In this state, the primary factor influencing the impedance signal is the volume of conductive medium displaced. This makes low-frequency measurements a direct method for determining cell size.
Conversely, a high-frequency current in the megahertz (MHz) range has enough energy to penetrate the cell membrane. The resulting impedance signal is then influenced by the electrical properties of the cell’s interior, including its cytoplasm conductivity and membrane capacitance. A healthy cell with an intact membrane produces a different signal than a dead cell with a permeable membrane.
This dual-frequency approach allows the technique to distinguish between live and dead cells. Live cells maintain a controlled internal environment and an intact membrane, leading to a characteristic high-frequency impedance signature. Damaged or dying cells lose this control, altering their internal conductivity and membrane capacitance, which is immediately detectable and assesses the population’s viability.
Comparison to Optical Flow Cytometry
Impedance and optical flow cytometry are both single-cell analysis techniques, but they differ in their measurement approach. The primary distinction is that impedance flow cytometry is a label-free technology. It analyzes cells based on inherent biophysical properties like size and membrane integrity.
In contrast, optical flow cytometry relies on extrinsic fluorescent labels. Researchers use fluorescent dyes or antibodies to mark specific cellular components or surface proteins. The instrument then uses lasers to excite these labels and detectors to measure the emitted light, providing information on the presence and quantity of specific molecules. This makes it ideal for identifying specific cell subpopulations.
These different approaches mean the two methods answer different biological questions. Impedance cytometry provides data on physical characteristics for applications like cell counting, size distribution analysis, and viability assessment. Optical cytometry excels at identifying cells based on specific molecular markers for applications like immunophenotyping or analyzing protein expression levels.
These operational differences extend to the instrumentation. Impedance-based systems do not require complex optical components like lasers, specialized filters, and light detectors. As a result, impedance flow cytometers are simpler in design, more compact, and more cost-effective to manufacture and maintain. This simplicity makes the technology more accessible for use outside specialized labs.
Key Applications in Research and Diagnostics
A primary application of impedance flow cytometry is in hematology. Automated hematology analyzers use this technology to perform the complete blood count (CBC), a routine diagnostic test. The instruments rapidly count red blood cells, white blood cells, and platelets, differentiating them based on their distinct size profiles.
The technique is widely used for cell viability and apoptosis analysis. In pharmaceutical research, it monitors cell culture health during drug screening to determine if a compound is toxic by detecting changes in membrane permeability. In bioprocessing, it ensures the quality of cells used to produce therapeutic proteins or other biologics.
The technology also has applications in microbiology for the detection and quantification of microorganisms. It is used in the food industry to test for bacterial or yeast contamination, providing results in seconds without time-consuming culture methods. This rapid analysis helps ensure food safety and monitor sanitation processes. The method is also valuable for monitoring fermentation processes in industries like brewing or biofuel production.