Cholate and pH Reduce Interference by Sodium Dodecyl Sulfate in the Determination of DNA with Hoechst

✍ Scribed by M.J. Bester; H.C. Potgieter; W.J.H. Vermaak

• Book ID: 102967319
• Publisher: Elsevier Science
• Year: 1994
• Tongue: English
• Weight: 554 KB
• Volume: 223
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1006/abio.1994.1588

✦ Synopsis

The use of the fluorescent dye 33258 Hoechst (Hoe) to quantitatively determine DNA in cell culture in the presence of lysing agents like sodium dodecyl sulfate (SDS) is limited by the masking effect of high levels of nonspecific fluorescence, caused by the binding of Hoe to micelles. The masking effect can be reduced substantially by increasing the concentration of the counterion, the addition of cholate, or the (\mathrm{pH}) of the buffer. An optimized method was developed, combining the antimasking effects of sodium chloride, cholate, and (\mathrm{pH}) to accurately determine DNA concentrations as low as (15 \mathrm{ng} /) (\mathrm{ml}) in the presence of up to (6.9 \mathrm{~mm}(0.2 %)) SDS. The effectiveness of SDS in cell dissolution can now be combined with the specificity and sensitivity of Hoe to determine cellular DNA. 1994 Academic Press, Inc.