Capillary Electrophoresis with Glycerol as an Additive

Two kinds of semi-liquid separation media are frequently used for the separation of DNA by high performance capillary electrophoresis (CE), namely dynamic or entangled free solution polymer sieving matrix, and dilute liquid-gel solutions. Polymer solutions may be defined in three different functional concentrations: dilute, semidilute, and entangled. In dilute solutions, the polymer molecules are physically separate; as the concentration of polymer increases the molecules begin to interact, and at a critical concentration the polymers begin to physically interact to form an entangled physical network (1 –3 ). These media have the ability to act as size selective sieving agents that partition DNA molecules, but since the media are liquid the injection of fresh media and the removal of expended media are facilitated. The media are typically neutral high molecular weight polymers and polymer sugars, which provide a high uniformity of electrophoretic medium if capillary electroosmotic flow (EOF) effects are suppressed. For the purpose of this review, particular attention will be drawn to entangled free solution capillary electrophoresis (ESCE) using Tris-borate-EDTA (TBE) buffers, with glycerol as an additive. For DNA separations, many different ESCE sieving media and dilute liquid-gel solutions have been used (Table 1 ): polysaccharide polymers such as, hydroxypropyl cellulose (HPC) (2 –4 ), hydroxypropyl methyl cellulose (HPMC) (5 –7 ), methyl cellulose (8 ,9 ), hydroxyethyl cellulose (HEC) (3 ,7 ,10 –15 ), dextran (16 ,17 ), TreviSol (18 ), semi-liquid agarose (19 ,20 ), and galactomannans (1 ). Nonsaccharide polymers such as linear polyacrylamides and substituted polyacrylamides (3 ,7 ,10 –26 ), linear and branched poly(ethylene oxides), and poly vinyl polymers (27 ) are also employed for a wide variety of separations purposes.

Assay	Matrix	Buffer	Detector	Comment	Ref.
dsDNA PCR-RFLP	HPMC	TBE/glycerol�ethidium	UV, LIF	Glycerol improves resolution	5,6,46,47,49
dsDNA PCR-RFLP	0.2–1% HEC	TBE	UV	Good resolution	2,4,10,11
STR-PCR PCR-RFLP	0.5% MC	0.5–2.5X TBE TAE	YoPro, LIF, no dye, UV	Better resolution in TBE	8,9,15
STR-PCR PCR-RFLP	1.25% HEC	TBE 7 M urea	no dye, UV	Good resolution of several STR	14
Supercoil plasmid	0.1–0.3% HEC	0.5–2.5X TBE	UV	Good resolution	54
dsDNA LCR	HPMC	TBE/glycerol	LIF	Good resolution 25 and 50 bp	48
dsDNA PCR-RFLP	0.5% PEO	TBE/dyes	LIF	Good resolution	27,55
dsDNA PCR-RFLP	Liquid	TBE		Better resolution in TBE	20,39
	Aragose	TAE
dsDNA PCR-RFLP	Trevisol	TAPS	UV	Good resolution	18
HPA	HPMC	TBE/glycerol�ethidium	UV, LIF	Glycerol improves resolution	50
SSCP	HPMC	TBE/glycerol�ethidium	UV. LIF	Glycerol stablizes confomers	51
SSCP	LPA	TBE�glycerol	UV	Good resolution	17,22,52,57
dsDNA PCR-RFLP	LPA	TBE�ethidium	UV	Good resolution	21,24,44
RNA	HPMC	TBE 7 M urea	UV	Glycerol diminishes resolution	45