EXTRACTION OF MEMBRANE-LIPIDS DURING FIXATION, DEHYDRATION AND EMBEDDING OF ACHOLEPLASMA-LAIDLAWII-CELLS FOR ELECTRON-MICROSCOPY
EXTRACTION OF MEMBRANE-LIPIDS DURING FIXATION, DEHYDRATION AND EMBEDDING OF ACHOLEPLASMA-LAIDLAWII-CELLS FOR ELECTRON-MICROSCOPY
复制标题
DOI：
10.1111/j.1365-2818.1983.tb04174.x
复制
发表时间：
1983-01-01
影响因子：
2
通讯作者：
CARLEMALM, E
中科院分区：
文献类型：
--
作者：
WEIBULL, C;CHRISTIANSSON, A;CARLEMALM, E
研究了在高温或低温下脱水和包埋过程中从生物膜中提取脂质的程度。本研究以A. laidlawii的细胞作为实验材料，因为这种细菌的脂质可以很容易地进行放射性标记，而不需要标记细胞的其余部分，而且脂质几乎全部位于细胞质膜上。细胞在277度固定。K用戊二醛，按顺序用该试剂和四氧化锇，或按顺序用戊二醛，四氧化锇和醋酸铀酰。在这些过程中脂质损失是可以忽略不计的。用戊二醛和四氧化二锇固定细胞后，室温下用乙醇脱水，333度包埋Epon。K，即接受常规治疗，约为。提取细胞90%的脂质含量。损失减少到约1000英镑。用醋酸铀酰处理20%，用非极性甲基丙烯酸酯树脂Lowicryl HM20代替Epon。用戊二醛和四氧化二锇固定细胞后，用238度乙醇脱水。室温下包埋在Lowicryl HM20中，几乎没有脂质提取。极性甲基丙烯酸酯-丙烯酸酯树脂Lowicryl K4M取代Lowicryl HM20导致细胞脂质含量减少一半。用乙二醇代替乙醇作为脱水剂对提取效果没有影响。仅用戊二醛固定的细胞，即使在238度脱水和包埋时，也损失了大约一半的脂质含量。K.从戊二醛固定细胞中提取的脂质物质所含的饱和脂肪酸略多于细胞中剩余的饱和脂肪酸。四氧化锇固定电池的情况正好相反。对于脂质种类，提取通常相当不特异性。
The extent to which lipids are extracted from biological membranes during dehydration and embedding procedures carried out at high or low temperatures was studied. Cells of A. laidlawii were used as experimental material, since the lipids of this bacterium can be easily radioactively labeled without labeling the rest of the cell, and the lipids are almost entirely located in the cytoplasmic membrane. The cells were fixed at 277.degree. K with glutaraldehyde, sequentially with this reagent and osmium tetroxide, or with glutaraldehyde, osmium tetroxide and uranyl acetate in that order. Loss of lipid during these procedures was negligible. When cells fixed with glutaraldehyde and osmium tetroxide were dehydrated with ethanol at room temperature and embedded in Epon at 333.degree. K, i.e., subjected to a conventional treatment, .apprx. 90% of the lipid content of the cells was extracted. The loss was reduced to .apprx. 20% when treatment with uranyl acetate was included in the procedure and the non-polar methacrylate resin Lowicryl HM20 was substituted for Epon. When cells fixed with glutaraldehyde and osmium tetroxide were dehydrated with ethanol at 238.degree. K and embedded in Lowicryl HM20 at room temperature, practically no lipid was extracted. Substitution of the polar methacrylate-acrylate resin Lowicryl K4M for Lowicryl HM20 resulted in the loss of half of the lipid content of the cells. The use of ethanediol as dehydrating agent instead of ethanol did not diminish the extraction. Cells fixed solely with glutaraldehyde lost about half of their lipid content, even when both dehydration and embedding were performed at 238.degree. K. The lipid material extracted from glutaraldehyde-fixed cells contained slightly more saturated fatty acids than that remaining in the cells. The reverse was true for osmium tetroxide-fixed cells. With respect to lipid species, the extractions were generally rather unspecific.