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How to obtain a good TRPS measurement of your particles

Come misurare in maniera precisa e affidabile le tue nanoparticelle (vescicole extracellulari, esosomi, ecc...) con la TRPS

Post publication date:

Monday, October 26, 2020

TRPS is the only technology which accurately delivers concentration of particles per unit volume across a defined particle size, accurate size distribution of these particles and simultaneous measurement of particle surface charge. Thus, reagent quality and sample preparation are important for obtaining the high precision data that you need. Reagent quality is vital for stable instrument use and high-quality reagents result in low background noise, stable baseline currents and highly stable and sensitive particle detection. In contrast, out-of-date reagents that have traces of contaminants or precipitates greatly increase pore instability and reduce accuracy of the data measured. To ensure you obtain high quality TRPS measurements, we strongly recommend all users to adopt Izon's best practice recommendations and protocols that detail easy step-by-step instructions. Please click here for more information.

The three most important parameters to look out for when performing a TRPS measurement are:

Baseline current
Rate plot
Calibration recording

Table 1. Resolving a drifting/fluctuating baseline current.

The baseline current reflects the conductivity of the electrolyte and indicates the state of the nanopore. If your particles have been prepared in the same electrolyte, the conductivity (or current in nA) should be similar. Changes in baseline current within 5% between successive recordings is acceptable. Any sudden change in baseline current outside this range suggests a partial blockage or modification of the nanopore and should be attended to before proceeding to the next measurement. A poorly prepared sample can cause issues during measurement particularly in partial blocking of the nanopore.

Should you come across a partial blockage during a measurement run, you can pause the measurement to avoid measurement inaccuracies and follow the flowchart to recover baseline. Presence of air bubbles and/or electrolyte leakage in the upper or lower fluid cell can cause the baseline current to drift or fluctuate. Please refer to Table 1 (image here attached) for recommendations to stabilise the baseline current.

Figure 1. Diagrammatic representation of the particle rate plot obtained during sample or calibration measurements on the qNano

Particle concentration is measured by comparing the gradient of the rate plot of the sample measurement to the calibration measurement. The rate plot must maintain a linear gradient to ensure recording of high-quality data as shown in Figure 1 here below. System stability is crucial for obtaining a linear plot and this can be achieved by ensuring proper sample preparation without introducing any contaminants in the sample. Non-linear rate plots will result in inaccurate size and concentration calculations and recommendations to rectify this issue can be found here.

It is good practice to run a calibration recording after each sample measurement, however provided that the samples do not modify the nanopore, a few samples can be recorded in succession before a calibration is recorded. For example, C1, S1, S2, C2, S3, S4, C3, etc. (C= calibration, S= sample).

Please visit Izon's support portal for more tips and recommendations for qNano and TRPS measurements.