Methods and devices based on capillary monolithic columns, preferably consisting of an underivatized poly(styrene-divinylbenzene) monolith, for separating a mixture of polynucleotides by ion pair-reverse phase-high performance chromatography (IP-RP-HPLC). In various aspects of the method and device the monolith is characterized by one or more of the following: the monolith is contained within a capillary tube; the monolith is immobilized by covalent attachment at the inner wall of the tube; the tube is devoid of retaining frits; the monolith is characterized by having above 10,000 theoretical plates per meter and preferably above 200,000 theoretical plates per meter; the method uses a mobile phase which is devoid of EDTA; the monolith has a surface morphology that is rugulose or brush-like; the chromatographic surfaces of the monolith are non-porous; the monolith has channels sufficiently large for convective flow of the mobile phase; the monolith is formed from a polymerization mixture including underivatized styrene, a crosslinking agent, and a porogen, wherein the porogen includes tetrahydrofuran. The monolith can be incorporated into a miniaturized chromatography system which can be coupled to a mass spectrometer for on-line separation and mass determination of single- or double-stranded polynucleotides.
基于毛细管单体柱的方法和设备,优选由未衍生的聚
苯乙烯-二
乙烯苯单体组成的单体柱,用于通过离子对-反相-高效
液相色谱(IP-RP-HPLC)分离多核苷酸混合物。在方法和设备的各个方面中,单体柱的特征之一或多个如下:单体柱被包含在毛细管管内;单体柱被共价固定在管的内壁上;管内没有保留过滤器;单体柱的特征是每米具有10,000个以上的理论板,最好是每米具有200,000个以上的理论板;该方法使用不含
EDTA的流动相;单体柱具有皱褶状或刷状的表面形态;单体柱的色谱表面是非多孔的;单体柱具有足够大的通道以便流动相的对流流动;单体柱由包括未衍生
苯乙烯、
交联剂和孔原剂的聚合混合物形成,其中孔原剂包括
四氢呋喃。单体柱可纳入微型色谱系统中,可与质谱仪耦合,用于在线分离和质量测定单链或双链多核苷酸。