Detection of total DNA with single-stranded DNA binding protein conjugates.
We have developed a rapid and sensitive method for total DNA measurement using single-stranded DNA binding protein from E coli conjugated with horseradish peroxidase or urease. To detect DNA, the sample is heated or alkali treated to denature the DNA and then filtered through nylon or nitrocellulose membranes. After the single-stranded DNA is bound to the membrane, single-stranded DNA binding protein enzyme-conjugate is incubated with the membrane. Next, the unbound conjugate is washed off the membrane and the bound conjugate detected colorimetrically. The assay can detect 10 pg of DNA in less than 3 hr. This method can be applied to the detection of DNA contamination in therapeutic proteins produced by recombinant DNA or hybridoma techniques.
Sheldon
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Molecular Devices Corporation, Menlo Park, California 94025.
Nagainis
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PA
Kung
V T
VT
D003124:Colorimetry / Q000379:methods; D004247:DNA / Q000032:analysis* / Q000378:metabolism; D004269:DNA, Bacterial / Q000032:analysis; D004268:DNA-Binding Proteins / Q000378:metabolism; D004926:Escherichia coli / Q000302:isolation & purification / Q000378:metabolism; D014510:Urease / Q000378:metabolism; D014871:Water Microbiology
D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D004247:DNA; D014510:Urease