What is the basis for the Coulter Principle of electronic particle counting?
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Wallace H Coulter discovered the Coulter Principle in the late 1940s (though a patent was not awarded until October 20, 1953). Coulter was influenced by the atomic bombs dropped on Hiroshima and Nagasaki. These events motivated Coulter to simplify and improve blood cell analysis so that large populations could be screened rapidly, as would be necessary in the event of a nuclear war. Partial funding of the project came from a grant award from the Office of Naval Research.[4][5] Show "Coulter Principle" refers to the use of an electric field for counting and sizing dilute suspensions of particles in conducting liquids. Wallace H. Coulter was awarded US Patent #2,656,508, Means for Counting Particles Suspended in a Fluid. The Coulter Principle is most commonly employed in a Coulter counter, which is an analytical instrument designed for a specific task such as counting cells. However, there are numerous other ways to implement the Coulter Principle. Several of these have been attempted, some with commercial success, and some purely for academic research. To date, the most commercially successful application of the Coulter Principle is in hematology, where it is used to obtain information about patients’ blood cells. The Coulter Principle relies on the fact that particles moving in an electric field cause measurable disturbances in that field. The magnitudes of these disturbances are proportional to the size of the particles in the field. Coulter identified several requirements necessary for practical application of this phenomenon. First, the particles should be suspended in a conducting liquid. Second, the electrical field should be physically constricted so that the movement of particles in the field causes detectable changes in the current. Finally, the particles should be dilute enough so that only one at a time passes through the physical constriction, preventing an artifact known as coincidence. While the Coulter Principle can be implemented in a variety of designs, there are two that have become the most commercially relevant. These include an aperture format and a flow cell format. Anomalous electrical pulses was generated because the concentration of samples were so high that multiple particles entered the aperture simultaneously. This situation is known as coincidence. This condition occured because there was almost no way to ensure that a single large pulse is the result of a single large particle or multiple small particles entering the aperture at once. To prevent this situation, samples must be fairly dilute, but we didn’t dilute our samples because we needed an accurate results. The shape of the generated electrical pulse varied with the particle path through the aperture. Shoulders and other artifacts occured because the electric field density varied across the diameter of the aperture. This variance is a result of both the physical constriction of the electric field and also the fact that the liquid velocity varied as a function of radial location in the aperture. In the flow cell format, this effect was minimized since sheath flow ensured each particle traveled an almost identical path through the flow cell. In the aperture format, we used the signal processing algorithms to correct the artifacts resulting from particle path. Conductive particles are often common concern for considering the Coulter Principle. Although, there are interesting scientific questions, on this procedure, it rarely affects the results of an experiment. This is because the conductivity difference between most conductive materials and ions in liquid (referred to as the discharge potential) is so great that most conductive materials act as insulators in a Coulter counter. The voltage we used to break down this potential barrier is referred to as the breakdown voltage. For those highly conductive materials that presented problems, the voltage we used during our Coulter experiment reduced below the breakdown potential (which was determined empirically). We use the Coulter principle to measured the volume of our samples, since the disturbance in the electric field is proportional to the volume of electrolyte displaced from the aperture. This medium has led to confusion amongst researchers who are use to optical measurements from microscopes or other systems that only view two dimensions and also show the boundaries of a sample. The Coulter Principle, on the other hand measured three dimensions and the volume displaced by a sample. Direct current vs alternating current Direct current has been used in the Coulter counters found in most research and cell laboratories. Direct current measurements are useful for an array of particles and allow for simplified data acquisition and processing. Base on this, we used direct current to simplified and processed data acquisition. Alternating current measurements are sometimes used in clinical hematology instruments, due to the special nature of cell membranes. At low frequencies (below 500 kHz), alternating and direct current measurements behave essentially the same way. At intermediate frequencies (500 kHz - 6 MHz), the plasma membrane of cells can become polarized, leading to a decreased capacitance of the measurement systems. However, at high frequencies (6-20 MHz), the cell membrane loses its polarization, and the electrical pulses provide information about the cell cytoplasm. The most successful and important application of the Coulter Principle is in the characterization of blood cells. The technique has been used to diagnose a variety of diseases, and is the standard method for obtaining red blood cell counts (RBCs) and white blood cell counts (WBCs) as well as several other common parameters. When combined with other technologies such as fluorescence tagging and light scattering, the Coulter Principle can help produce a detailed profile of patients’ blood cells. In addition to clinical counting of blood cells (cell diameters of ~6-10 micrometres, typically), the Coulter principle has established itself as the most reliable laboratory method for counting a wide variety of cells, ranging from bacteria (< 1 micrometre in size), fat cells (~400 micrometre), plant cell aggregates (>~1200 micrometre), and stem cell embryoid bodies (~900 micrometre). The technique has become so standardized that ASTM International has published a procedure on the topic: ASTMF2149-01(2007) Standard Test Method for Automated Analyses of Cells-the Electrical Sensing Zone Method of Enumerating and Sizing Single Cell Suspensions. Particle characterization The Coulter Method has proved useful for applications well beyond cellular studies. The fact that it individually measures particles, is independent of any optical properties, is extremely sensitive, and is very reproducible has appeal to a wide variety of fields. Consequently, the Coulter Principle has been adapted to the nanoscale to produce a novel nanoparticle characterization technique called Tunable Resistive Pulse Sensing, or TRPS. TRPS enables high-fidelity analysis of a diverse set of nanoparticles, including (but not limited to): functionalized drug delivery nanoparticles, Virus-like particles (VLPs), liposomes, exosomes, polymeric nanoparticles, microbubbles. Benefits of Coulter Method - Increase productivity with consistently reliable results. - With its versatile closed tube sampling system, the coulter save time and enhance safety for laboratorians. - Flexible specially-formulated reagents, fully automated QC and calibration platforms provide consistently reliable results.
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