Rule G S, Montagna R A, Durst R A
Analytical Chemistry Laboratories, Cornell University, Geneva, New York 14456-0462, USA.
Anal Biochem. 1997 Jan 15;244(2):260-69. doi: 10.1006/abio.1996.9875.
Liposomes that have been labeled externally with a DNA oligomer are used in a capillary-migration, sandwich-hybridization assay for specific DNA target sequences. The liposomes are used in a DNA detection scheme that produces visually observable results in 10 min. The preparation and covalent attachment of a thiol-activated 22-base oligomer to the external surface of dye-containing liposomes is described, and the specificity of the assay toward perfectly complementary target DNA is demonstrated. Several characteristics of DNA-tagged liposomes that allow the use of increased stringency during hybridization are evaluated. These include the effect of temperature, formamide, and salt concentration on both the sandwich-hybridization assay and the liposomes themselves. The effects of several components of a common hybridization solution are determined with regard to both assay performance and liposome stability. Using a solution of 0.02% sodium dodecyl sulfate in 3X standard saline citrate, a visual detection limit of 200 amol of target DNA was obtained.
用DNA寡聚物进行外部标记的脂质体用于针对特定DNA靶序列的毛细管迁移夹心杂交分析。脂质体用于一种DNA检测方案，该方案可在10分钟内产生肉眼可见的结果。描述了硫醇活化的22碱基寡聚物在含染料脂质体外部表面的制备和共价连接，并证明了该分析对完全互补靶DNA的特异性。评估了DNA标记脂质体的几个特性，这些特性允许在杂交过程中提高严谨性。其中包括温度、甲酰胺和盐浓度对夹心杂交分析和脂质体本身的影响。就分析性能和脂质体稳定性而言，确定了常见杂交溶液中几种成分的影响。使用在3X标准柠檬酸盐缓冲液中的0.02%十二烷基硫酸钠溶液，获得了200 amol靶DNA的视觉检测限。
