Hung K C, Rashidzadeh H, Wang Y, Guo B
Department of Chemistry, Cleveland State University, Ohio 44115, USA.
Anal Chem. 1998 Jul 15;70(14):3088-93. doi: 10.1021/ac980090e.
Poor detection limits and strong salt effects are two of the main problems encountered in the matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometric analysis of DNA. This work demonstrates that a probe tip with a paraffin wax film (Parafilm) surface improves the MALDI performance in DNA analysis compared to the commonly used metal surface. First, the use of Parafilm increases the detection sensitivity. It was found that the detection limit achieved with Parafilm was 5 times better than that obtained using stainless steel for a 85mer. More importantly, the Parafilm method could improve detection of larger DNA components in the presence of a large excess of a smaller DNA component or in a DNA mixture. This feature is important to analyses of PCR and sequencing products. Second, we found that the use of Parafilm increased the salt tolerance limits for the 17-, 41-, and 85mers studied in this work and that the salt effect was less sensitive to the DNA size. Third, this method offers other analytical benefits, including producing a more homogeneous coverage of matrix/DNA, adding no extra cost and time to sample preparation, and eliminating the commonly required step for cleaning the probe after analysis. In this paper, we will also present our perspectives on why the use of Parafilm can improve the MALDI-TOF performance in DNA analysis.
检测限不佳和强烈的盐效应是DNA的基质辅助激光解吸/电离飞行时间（MALDI-TOF）质谱分析中遇到的两个主要问题。这项工作表明，与常用的金属表面相比，具有石蜡膜（Parafilm）表面的探针尖端可改善DNA分析中的MALDI性能。首先，使用Parafilm可提高检测灵敏度。结果发现，对于一个85聚体，使用Parafilm实现的检测限比使用不锈钢时好5倍。更重要的是，在存在大量较小DNA组分过量或在DNA混合物中时，Parafilm方法可改善对较大DNA组分的检测。这一特性对PCR产物和测序产物的分析很重要。其次，我们发现使用Parafilm提高了本研究中所研究的17聚体、41聚体和85聚体的耐盐极限，并且盐效应对DNA大小不太敏感。第三，该方法还具有其他分析优势，包括使基质/DNA覆盖更均匀，在样品制备过程中不增加额外成本和时间，以及省去分析后清洁探针这一通常需要的步骤。在本文中，我们还将阐述关于为何使用Parafilm可改善DNA分析中MALDI-TOF性能的观点。
