Vibrational Responses of Bound and Nonbound Targeted Lipid-Coated Single Microbubbles

Abstract

One of the main challenges for ultrasound molecular imaging is acoustically distinguishing nonbound microbubbles from those bound to their molecular target. In this in vitro study, we compared two types of in-house produced targeted lipid-coated microbubbles, either consisting of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine, C16:0 (DPPC) or 1,2-distearoyl-sn-glycero-3-phosphocholine, C18:0 (DSPC) as the main lipid, using the Brandaris 128 ultrahigh-speed camera to determine vibrational response differences between bound and nonbound biotinylated microbubbles. In contrast to previous studies that studied vibrational differences upon binding, we used a covalently bound model biomarker (i.e., streptavidin) rather than physisorption, to ensure binding of the biomarker to the membrane. The microbubbles were insonified at frequencies between 1 and 4 MHz at pressures of 50 and 150 kPa. This paper shows lower acoustic stability of bound microbubbles, of which DPPC-based microbubbles deflated most. For DPPC microbubbles with diameters between 2 and [Formula: see text] driven at 50 kPa, resonance frequencies of bound microbubbles were all higher than 1.8 MHz, whereas those of nonbound microbubbles were significantly lower. In addition, the relative radial excursions at resonance were also higher for bound DPPC microbubbles. These differences did not persist when the pressure was increased to 150 kPa, except for the acoustic stability which further decreased. No differences in resonance frequencies were observed between bound and nonbound DSPC microbubbles. Nonlinear responses in terms of emissions at the subharmonic and second harmonic frequencies were similar for bound and nonbound microbubbles at both pressures. In conclusion, we identified differences in vibrational responses of bound DPPC microbubbles with diameters between 2 and [Formula: see text] that distinguish them from nonbound ones.

Dispersing and Sonoporating Biofilm-Associated Bacteria with Sonobactericide

“…Our data is in agreement with the current knowledge that clustering happens quickly (ms) [61]. The range of microbubble sizes (3.8-5.4 µm) leading to clustering were both within, and slightly outside, of resonance; however, that was determined at 50 kPa [58]. Increasing pressure is known to (linearly) dampen resonance frequency [62].…”

“…However, other sonobactericide studies used similarly sized microbubbles [50,51,57], and the mean diameter was in range of the clinically approved human albumin-coated microbubbles (Optison, mean diameter (range) 3-4.5 µm). It is of potential importance that this larger size is closer to resonance frequency, at 2 MHz [58], which could translate to a more efficacious sonobactericide treatment, since a larger microbubble population would achieve maximum excursion amplitude. The microbubble concentration used in this study was chosen to maximize sonoporation potential, due to the sheer number of cells (~7000), and was supported by Dong et al [50], who observed that a higher microbubble concentration led to enhanced bacterial PI uptake in biofilms.…”

Sonobactericide: An Emerging Treatment Strategy for Bacterial Infections

“…It has been observed that as the initial stand-off distance increases, biofilm disruption, sonoporation and cytoskeleton disassembly decrease (Goh et al 2015;Wang et al 2018b). Moreover, microbubble dynamics also depend on the distance from and material properties of a surface (Overvelde et al 2011;van Rooij et al 2017).…”

Opening doors with ultrasound and microbubbles: Beating biological barriers to promote drug delivery

“…The direct impact of microbubble cavitation on cellular barriers has been investigated in vitro on various cell lines including cancer cell lines and primary endothelial cells [8,[28][29][30][31][32][33][34][35][36][37]. The latter is the most biologically relevant as microbubbles cannot extravasate from the blood stream after intravenous injection.…”