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  • Title: Detection of isozymes of deoxyribonucleases I and II on electrophoresed gels with picogram sensitivity using SYBR Green I.
    Author: Iida R, Yasuda T, Tsubota E, Nakashima Y, Sawazaki K, Aoyama M, Matsuki T, Kishi K.
    Journal: Electrophoresis; 1998 Oct; 19(14):2416-8. PubMed ID: 9820960.
    Abstract:
    A highly sensitive method for detecting deoxyribonucleases (DNases) I and II on an electrophoresed gel is described. A dried agarose film sheet containing DNA as a substrate and a buffer reagent was placed in contact with the gel surface after electrophoresis (DAFO method, Yasuda et al., Anal. Biochem. 1989, 183, 84-88). After an appropriate incubation period, the film sheet was peeled off and stained with SYBR-Green I (SG), and then the DNase isozyme bands were detected using a fluorescence image analyzer. We could detect pg levels of the DNases (DNase I, 2 pg; DNase II, 2pg), which represents a 32- to 128-fold increase in sensitivity compared with the original DAFO method using ethidium bromide (EB) as the fluorescent dye. A combination of this new detection method and isoelectric focusing electrophoresis in polyacrylamide gel allowed accurate DNase I typing from 1 microL human serum. This new technique has been named SG-DAFO, after its original dried agarose film overlay method using EB (EB-DAFO).
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