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December 29, 2020 – Using positive controls to ensure particle size and concentration accuracy

Let’s face it, most particle analyzers are pretty much “black boxes”: you put the sample into it, and sometime later you get a result, whether it be a true particle size distribution or just an average size as in the case of Dynamic Light Scattering (DLS). Sure, the “calibration” of any of these systems can be checked by running NIST-traceable polystyrene (or silica) beads mixed at a known concentration. However, the performance on any given instrument for these beads (in water or PBS) may not be representative of how the same instrument responds to your actual samples. This is especially true for optically based systems, where the refractive index (and perhaps viscosity or other factors) of both the sample particles and suspending medium can affect the instrument response.

One of the many advantages of Spectradyne’s Microfluidic Resistive Pulse Sensing (MRPS) is that there is absolutely no dependence of the measurements on particle material and optical properties such as viscosity or refractive index. In fact, MRPS directly measures particle volume: this means that a particle of volume X made of solid gold will give the same measurement as a hollow particle of equal volume. So, in the case of an MRPS system, the performance when measuring calibration beads (of any material) is directly indicative of the performance of the instrument on any other sample particle, which is not true for optical systems.

Another advantage is small sample size: MRPS technology uses microfluidic cartridges that require very small volumes of sample, just 3 uL in the case of Spectradyne’s nCS1, small enough that samples of tick saliva can be reliably measured!

A far larger advantage of MRPS, though, is that each particle is measured individually, one at a time. This means that the measurement of any given particle is completely independent of the measurements of other particles in the sample. This is not true in Nanoparticle Tracking Analysis (NTA) systems, where it can be shown that these instruments have a Limit of Detection (LOD) that varies based on sample composition and can cause spurious results.

What this means is that for measurements in an MRPS system, you can actually add calibrated spheres at a known concentration directly into your sample as a positive control. By doing this, you get verification in the sample that the instrument is accurate in both particle sizing and concentration.


EV sample measured twice in Spectradyne’s nCS1TM MRPS system: Once by itself and then with 150nm beads spiked in. The graph overlay visually shows repeatability of the two, while the spiked calibration beads in the second sample quantitatively proves measurement accuracy.

The figure above shows an example of how this works in practice. An extracellular vesicle (EV) sample was run twice in Spectradyne’s nCS1TM MRPS system: The first time by itself, and the second time spiked with 150 nm NIST-traceable polystyrene nanospheres diluted to be 2.0 × 109 particles/mL in concentration. As can be seen, both runs of the sample overlay perfectly, while the spiked version shows a clear peak at 150 nm with a measured concentration of 2.04 × 109 particles/mL. This type of in-measurement control cannot be done with optically based systems because the sixth-order dependence of light scattering on diameter causes the spiked-in control beads to completely “mask” the instrument’s response on the sample particles!

By using in-measurement, positive controls in an MRPS system, one can be assured of each measurement’s accuracy for both particle sizing and concentration. This can be especially critical in regulated environments, where proving measurement traceability is greatly simplified by having in-measurement controls.

Can your particle analyzer do this?

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