An in vitro comparative study of the physical and imaging enhancement characteristics between two nano-scale ultrasound contrast agents: PFOB lipid particles and C3F8 lipid microbubbles

Abstract

Objective To prepare two nano-scale ultrasound contrast agents,lipid particles and microbubbles,and to compare their physical and imaging enhancement characteristics.Methods Lipid nanoparticles were prepared with phospholipid and liquid perfluorocarbon (PFOB).Lipid microbubbles were prepared with phospholipid and perfluoropropane(C3F8).Physical and chemical properties of the two ultrasound contrast agents were measured,including morphology,size,zeta potential,concentration and stability.Both lipid nanoparticles and microbubbles with/without biotin were imaged using a 12.0 MHz linear-array transducer with/without avidin.The imaging data were analyzed by using Matlab software.Comparative data between the two groups were statistically analyzed by two-sample t test.Repeated measures analysis of variance was used among the observations at multiple time points within the same sample and Bonferroni method was used for post hoc analysis.Comparison of the signal intensity with and without avidin of the same sample was performed by paired t test.Results (1) Microscopically,both PFOB lipid particles and C3F8 microbubbles were in form of spheres.Their particle diameters were (152.30 ± 35.99) nm and (774.59 ± 108.59) nm,respectively (t =-24.327,P ＜ 0.001).Their zeta potential was (-40.90 ±6.51) mV and (-14.80 ± 3.97) mV,respectively (t =-15.308,P ＜ 0.001).(2) Concentration and size of PFOB lipid particles remained stable during the whole observation period (C0h =(2.28 ± 0.64) ×1011/ml,C1w =(2.06 ±0.53) × 1011/ml; D0h =(152.30 ±35.99) nm,D1 w =(178.80 ±63.07) nm).Concentrations of lipid microbubbles were stable within 12 h (C0h =(4.08 ± 0.96) × 1010/ml,C12h =(3.25 ± 1.02) × 1010/ml) ; and then reduced significantly after 24 h,2 d,4 d and 1 week (C24h:(2.28 ±0.73) ×1010/ml; C2d:(1.56±0.54) ×1010/ml; C4d:(1.03±0.37) ×1010/ml; C1w:(0.74 ±0.24) ×1010/ml ; F =78.515,P ＜ 0.01).Diameters of the particles remained stable within 2 d (D0h =(774.59 ±108.59) nm,D2d =(1020.68 ± 223.64) nm) ; and then increased after 4 d and 1 week (D4d:(1391.67 ±268.65) nm; D1w:(1532.41 ± 326.25) nm,respectively,F =50.772,P ＜ 0.01).(3) Neither control nor biotinylated PFOB particles were visible without avidin.With the addition of avidin,significantly enhanced images were generated by biotinylated PFOB particles (2.18 ±0.71,82.19 ± 15.68,t =-15.698,P ＜ 0.001) but not with the control PFOB particles.Both control and biotinylated C3F8 microbubbles showed no significant difference in their signal intensity with/without avidin (29.16 ± 6.84 and 31.34 ±7.03,33.69±8.24 and28.75±7.18,t=0.653,-1.393,bothP＞0.05).Conclusion Compared to C3F8 microbubbles,PFOB particles have smaller size,greater stability,higher SNR ratio and better imaging enhancement characteristics,therefore,more suitable for use as the target contrast agent for ultrasound imaging. Key words: Contrast media; Ultrasonography ; Lipid nanoparticles ; Lipid microbubbles ; Comparative study