Pulsed Field Gel Electrophoresis: Past, present, and future

Pulsed Field Gel Electrophoresis (PFGE) has been considered for many years the 'gold-standard' for characterizing many pathogenic organisms as well as for subtyping bacterial species causing infection outbreaks. This article reviews the basic principles of PFGE and it includes the main advantages and limitations of the different electrode configurations that have been used in PFGE equipment and their influence on the DNA electrophoretic separation. Remarkably, we summarize here the most relevant theoretical and practical aspects that we have learned for more than 20 years developing and using the miniaturized PFGE systems. We also discussed the theoretical aspects related to DNA migration in PFGE agarose gels. It served as the basis for simulating the DNA electrophoretic patterns in CHEF mini gels and mini-chambers during experimental design and optimization. A critical comparison between standard and miniaturized PFGE systems, as well as the enzymatic and non-enzymatic methods for intact immobilized DNA preparation, is provided throughout the review. The PFGE current applications, advantages, limitations and future challenges of the methodology are also discussed.