Roselle Christopher, Whitehouse Dana, Follmer Thy, Ansbro Fran, Bouaraphan Silikhone, Guan Liming, Wang Sha-Ke, Shank-Retzlaff Mary, Verch Thorsten
Merck & Co. Inc., Kenilworth, NJ.
J Immunol Methods. 2017 Mar;442:20-28. doi: 10.1016/j.jim.2016.12.003. Epub 2016 Dec 27.
Dilutions are a common source of analytical error, both in terms of accuracy and precision, and a common source of analyst mistakes. When serial dilutions are used, errors compound, even when employing laboratory automation. Direct point dilutions instead of serial dilutions can reduce error but is often impractical as they require either large diluent volumes or very small sample volumes when performed with traditional liquid handling equipment. We evaluated preparation of dilution curves using a picoliter digital dispenser, the HP, Inc. / TECAN D300 which is capable of accurately delivering picoliter volumes directly into sample wells filled with assay diluent. Dilution linearity and variability of the direct dilutions were similar to or less than those generated with a traditional liquid handler as measured using a fluorophore assay and an ELISA used to measure vaccine potency. Minimum concentrations for detergent in the dispensed sample were identified but no correlation with detergent characteristics was observed. The tolerance to protein in the sample was evaluated as well with up to 5% BSA having no impact on dispense linearity and precision. We found the digital dispenser to reduce automation complexity while maintaining or improving assay performance in addition to facilitating complex plate lay-outs.
稀释是分析误差的常见来源，无论是在准确性还是精密度方面，也是分析人员出错的常见原因。当使用系列稀释时，即使采用实验室自动化，误差也会累积。直接点稀释而非系列稀释可以减少误差，但通常不切实际，因为使用传统液体处理设备进行时，它们要么需要大量稀释剂体积，要么需要非常小的样品体积。我们评估了使用皮升数字分配器（惠普公司/帝肯D300）制备稀释曲线的情况，该分配器能够将皮升体积的液体准确地直接输送到装有分析稀释剂的样品孔中。使用荧光团测定法和用于测量疫苗效力的酶联免疫吸附测定法（ELISA）测量时，直接稀释的稀释线性和变异性与传统液体处理仪产生的相似或更低。确定了分配样品中洗涤剂的最低浓度，但未观察到与洗涤剂特性的相关性。还评估了样品中蛋白质的耐受性，高达5%的牛血清白蛋白（BSA）对分配线性和精密度没有影响。我们发现数字分配器除了便于进行复杂的板布局外，还能降低自动化复杂性，同时保持或提高分析性能。
