Bio-Nanotechnology Laboratory, Department of Neuroscience and Pharmacology, University of Copenhagen, 2100 Copenhagen, Denmark.
Langmuir. 2011 Feb 1;27(3):866-9. doi: 10.1021/la1035163. Epub 2011 Jan 4.
The aggregation of casein micelles (CMs) induced by milk-clotting enzymes is a process of fundamental importance in the dairy industry for cheese production; however, it is not well characterized on the nanoscale. Here we enabled the monitoring of the kinetics of aggregation between single CMs (30-600 nm in diameter) by immobilizing them on a glass substrate at low densities and subsequently imaging them with fluorescence microscopy. We validated the new method by a quantitative comparison to ensemble measurements of aggregation. Single-particle statistics allowed us to observe for the first time several heterogeneities in CM aggregation. We observed two types of CM growth: a slow increase in the size of CMs and a stepwise increase attributed to interactions between aggregates preformed in solution. Both types of growth exhibit a lag phase that was very heterogeneous between different CMs, suggesting significant differences in their composition or structure. Detailed size histograms of CMs during aggregation also revealed the presence of two distinct subpopulations with different growth amplitudes and kinetics. The dependence of these distinct nanoscale processes/parameters on aggregation conditions is not accessible to bulk measurements that report only ensemble-average values and may prove important to an in-depth understanding of CM aggregation.
在乳品行业中,用于奶酪生产的凝乳酶诱导的酪蛋白胶束(CM)聚集是一个非常重要的过程;然而,目前对于该过程在纳米尺度上的特性还没有很好的描述。在这里,我们通过将直径为 30-600nm 的单个 CM 固定在低密度的玻璃基底上,并随后用荧光显微镜对其进行成像,从而实现了对单个 CM 之间聚集动力学的监测。我们通过与聚集的集合测量的定量比较来验证了新方法的有效性。单粒子统计允许我们首次观察到 CM 聚集中的几个异质性。我们观察到两种 CM 生长类型:CM 尺寸的缓慢增加和归因于溶液中预先形成的聚集体之间相互作用的逐步增加。这两种类型的生长都表现出滞后阶段,不同 CM 之间的滞后阶段非常不均匀,这表明它们的组成或结构存在显著差异。在聚集过程中对 CM 进行详细的尺寸直方图分析还揭示了存在两种具有不同生长幅度和动力学的不同亚群。这些不同的纳米级过程/参数对聚集条件的依赖性无法通过仅报告集合平均值的体相测量来获得,这对于深入了解 CM 聚集可能非常重要。