Sgouropoulou V, Makedou K, Seitanidou M, Trachana M, Karagiozaki V, Logothetidis S
Pediatric Immunology and Rheumatology Referral Center, 1st Department of Pediatrics, Aristotle University of Thessaloniki, Hippokration General Hospital, Thessaloniki, Greece.
Laboratory of Biological Chemistry, School of Medicine, Faculty of Health Sciences, Aristotle University of Thessaloniki, Greece.
Clin Biochem. 2015 Jun;48(9):628-30. doi: 10.1016/j.clinbiochem.2015.03.010. Epub 2015 Mar 27.
To investigate the impact of freezing in -80°C on the structure of isolated low density lipoproteins (LDLs), using nanotechnology, such as Atomic Force Microscopy (AFM).
Blood EDTA plasma was obtained from healthy subject and used immediately to isolate LDL by sequential ultracentrifugation at 10°C in 55,000 rpm for 3h, using a Beckmann XL-90 ultracentrifuge (75Ti rotor), in the presence of KBr in PBS. LDLs were then diluted with PBS until final concentrations of 5 and 15 mg LDL/dl. After initial observation, samples were frozen in -80°C for two weeks and observed again after thawing. Experiments were performed in triplicate on two smooth and clean substrates of different hydrophobicity, glass (HOPG) and Si (c-Si). Statistical significance was set at 0.05.
Macroscopically, LDL particles formed aggregations in a dendroid layout. There were no differences between images taken from both substrates (HOPG and c-Si). Frozen samples presented significantly smaller LDL particles, than fresh ones. In specific, mean diameter of LDL particle in the fresh LDL sample was 19.77 nm, ranging from 13.34 to 28.76 nm. The frozen LDL sample had a mean diameter of 5.2 nm, ranging from 2.0 to 8.0 nm, which was significantly different from the unfrozen.
Atomic Force Microscopy showed that freezing of LDL causes alterations in their size.
使用纳米技术，如原子力显微镜（AFM），研究在-80°C下冷冻对分离的低密度脂蛋白（LDL）结构的影响。
从健康受试者获取血液乙二胺四乙酸（EDTA）血浆，并立即用于通过在10°C下以55,000转/分钟的转速进行连续超速离心3小时来分离LDL，使用贝克曼XL-90超速离心机（75Ti转子），在PBS中存在溴化钾（KBr）的情况下进行。然后用PBS将LDL稀释至最终浓度为5和15 mg LDL/dl。初步观察后，将样品在-80°C下冷冻两周，解冻后再次观察。在两种不同疏水性的光滑干净底物，玻璃（高定向热解石墨，HOPG）和硅（c-Si）上重复进行三次实验。统计学显著性设定为0.05。
宏观上，LDL颗粒形成树状排列的聚集体。从两种底物（HOPG和c-Si）拍摄的图像之间没有差异。冷冻样品中的LDL颗粒明显比新鲜样品中的小。具体而言，新鲜LDL样品中LDL颗粒的平均直径为19.77 nm，范围为13.34至28.76 nm。冷冻LDL样品的平均直径为5.2 nm，范围为2.0至8.0 nm，与未冷冻的样品有显著差异。
原子力显微镜显示，LDL的冷冻会导致其大小发生改变。
