Nanoparticle Tracking Analysis: A powerful tool for characterizing magnetosome preparations

Abstract

Nanoparticle Tracking Analysis (NTA) has been employed to measure the particle concentration and size distribution of magnetosomes extracted and purified from Magnetospirillum gryphiswaldense MSR-1, and then exposed to probe ultrasonication for various times, or 1% (w/v) sodium dodecyl sulphate (SDS) for 1 h. Particle concentration increased 3.7-fold over the first 15 min of ultrasonication (from 2 × 10 8 to >7.3 × 10 8 particles mL -1 ), but fell steeply to ~3.6 × 10 8 particles mL -1 after 20 min. NTA of untreated magnetosome preparation confirmed a wide particle distribution dominated by larger species (D[1,0] = 312 nm; Dn50 = 261 nm; mode = 243 nm) with no particles in the size range of isolated single magnetosomes. After 5 min of ultrasonication the whole particle size distribution shifted to smaller size (D[1,0] = 133 nm; Dn50 = 99 nm; mode = 36 nm, corresponding to individual magnetosomes), but longer treatment times (15 and 20 min) reversed the previous transition; all characteristic numbers of the particle size distributions increased and very few small particles were detected. Side-by-side comparison of NTA and TEM sizing data revealed remarkable similarity at low ultrasonication times, with both showing single magnetosomes accounted for ~30% population after 5 min. Exposure of magnetosomes to SDS resulted in a ~3-fold increase in particle concentration to 5.8 × 10 8 particles mL -1 , narrowing of the size distribution and gross elimination of particles below 60 nm. We conclude that NTA is a rapid cost-effective technique for measuring particle number, size distribution and aggregation state of magnetosomes in solution, but requires further work to improve its resolving power.

Evaluation of cell disruption technologies on magnetosome chain length and aggregation behaviour from Magnetospirillum gryphiswaldense MSR-1

“…Lysozyme had previously been used to study periplasmatic and cytoplasmatic fractions in MTB cells ( Noguchi et al, 1999 ; Suzuki et al, 2007 ), but very few studies have used this enzyme for magnetosome extraction ( Nakamura et al, 1991 ). French press ( Grünberg et al, 2004 ; Curcio et al, 2020 ), high-pressure homogenization ( Tang et al, 2012 ; Amor et al, 2014 ; Fernández-Castané et al, 2021 ) or sonication ( Kobayashi et al, 2006 ; Alphandéry and Chebbi, 2012 ) are more commonly used for disruption of MTB due to their simplicity and cost-effectiveness. The effect of these disruption techniques on magnetosome chains has not been studied in detail yet, but large aggregates have been observed when the magnetosome membrane is removed.…”

“…As a result of chain breakage, short chains which were not present in pre-disrupted MSR-1 cells were detected after cell disruption. HPH treatment seemed to be the technique that generated less damage to the chains, yielding the lowest short-chain frequency (4%) and the highest long-chain frequency (66%) ( Alphandéry et al, 2013 ; Li, 2018 ; Fernández-Castané et al, 2021 ). The results also revealed that EZ treatment most significantly affected the chain integrity as only 34% of the chains were considered long and 17% short.…”

“…It is known that uncoated or partially coated magnetosomes are prone to self-aggregation ( Kobayashi et al, 2006 ). Previous studies revealed extensive chain breakage and magnetosome agglomeration as a direct consequence of magnetosome ultrasonication over an extended period of time ( Alphandéry and Chebbi, 2012 ; Fernández-Castané et al, 2021 ). Nakamura and co-workers compared ultrasonication and lysozyme disruption treatments by studying the particle sizes as well as magnetosome membrane thickness ( Nakamura et al, 1991 ).…”

“…The results suggest the better suitability of nFCM to characterize single magnetosomes than longer magnetosome chains. In addition, the necessity of having to acquire two size ranges (40–200 nm and 200–1,000 nm) and the sample preparation protocol may biase the actual size distributions as well as the particle concentrations of the different magnetosome preparations, which may represent a disadvantage over other similar techniques such as Nanoparticle Tracking Analysis (NTA) ( Fernández-Castané et al, 2021 ). nFCM is a reliable technique to determine nanoparticle concentrations but due to the heterogeneity in the magnetosome chain length and the nature of the technology, this technique is not suited to characterize the size distribution of untreated magnetosome preparations.…”

“…Regardless of the methodology employed to disrupt MSR-1 cells, magnetosomes were recovered following the same purification protocol adapted from Fernandez-Castane et al ( Fernández-Castané et al, 2021 ). The disrupted cell suspensions were subjected to magnetic separation using a NdFeB N45 magnet by performing at least a series of 10 washes.…”