Department of Bioengineering, University of Louisville, Louisville, Kentucky, USA.
Department of Surgery, University of Louisville, Louisville, Kentucky, USA.
Ultrasound Med Biol. 2023 Jan;49(1):90-105. doi: 10.1016/j.ultrasmedbio.2022.08.005. Epub 2022 Oct 12.
Continuous-flow acoustofluidic technologies can potentially improve processing of T lymphocytes for cell therapies by addressing the limitations with viral and non-viral delivery methods. The objective of this study was to assess the intracellular delivery efficiency with acoustofluidic treatment compared with that of static ultrasound treatment. Optimization of parameters in acoustofluidic and static configurations was performed by assessing intracellular delivery of a fluorescent compound (calcein) in viable human Jurkat T lymphocytes. Ultrasound pressure and the concentration of cationic phospholipid-coated microbubbles influenced calcein delivery in both systems. In the static system, a treatment time of 45 s increased molecular delivery compared with 0-30 s (p < 0.01). Refined parameters were used to assess molecular delivery of small and large compounds (0.6-kDa calcein and 150-kDa fluorescein isothiocyanate-dextran, respectively) after ultrasound treatment with the acoustofluidic or static systems. Molecular delivery was similar with refined parameters for acoustofluidic treatment and static treatment (p > 0.05), even though acoustofluidic treatment had lower microbubble concentration (24 μg/mL vs. 94 μg/mL) and shorter treatment time (∼2-3 s vs. 45 s). This study indicates that the acoustofluidic system can significantly enhance intracellular molecular delivery, which could potentially enable acoustofluidic cell transfection during continuous flow processing for manufacture of cell therapies or other applications.
连续流声流控技术可以通过解决病毒和非病毒传递方法的局限性，潜在地改善 T 淋巴细胞的细胞治疗处理。本研究的目的是评估与静态超声处理相比，声流控处理的细胞内传递效率。通过评估荧光化合物（钙黄绿素）在活的人 Jurkat T 淋巴细胞中的细胞内传递，对声流控和静态配置中的参数进行了优化。在两种系统中，超声压和阳离子磷脂包被的微泡浓度都影响钙黄绿素的传递。在静态系统中，与 0-30 s 相比，45 s 的处理时间增加了分子传递（p < 0.01）。使用改进的参数来评估小和大化合物（0.6 kDa 钙黄绿素和 150 kDa 异硫氰酸荧光素-葡聚糖）的分子传递，分别在声流控或静态系统进行超声处理后。即使声流控处理的微泡浓度（24 μg/mL 对 94 μg/mL）和处理时间（~2-3 s 对 45 s）较低，声流控处理的改进参数仍与静态处理的分子传递相似（p > 0.05）。这项研究表明，声流控系统可以显著增强细胞内分子传递，这可能使声流控细胞转染能够在连续流处理过程中用于制造细胞疗法或其他应用。
