Native fluorescence detection of nucleic acids and DNA restriction fragments in capillary electrophoresis

Journal Article · Fri Jan 15 00:00:00 EST 1993 · Analytical Chemistry (Washington); (United States)

DOI:https://doi.org/10.1021/ac00050a011· OSTI ID:5782935

Milofsky, R E; Yeung, E S ^[1]

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1. Ames Laboratory, IA (United States)

A sensitive laser-induced fluorescence (LIF) detection scheme for native nucleic acids and DNA restriction fragments separated by capillary electrophoresis (CE) has been developed. The 275.4-nm line from an argon ion laser or the 248-nm line from a waveguide KrF laser is used to excite native fluorescence. Detection limits for guanosine and adenosine monophosphate (1.5 [times] 10[sup [minus]8] and 5 [times] 10[sup [minus]8] M, respectively) are up to 3 orders of magnitude lower than UV detection. Sensitivity for native fluorescence of DNA restriction fragments in gel-filled capillaries rivals that of UV absorption. The decrease in performance in gel-filled separations using LIF detection is caused by the high background associated with gel fluorescence, as well as gel quenching of the fluorescence emission. The development of gels exhibiting lower background fluorescence or off-column coupling should lead to significant improvements in sensitivity over UV detection. This novel and practical system enables, for the first time, the sensitive detection of nucleic-acid-containing compounds without the need for fluorescence labeling. 48 refs., 4 figs., 1 tab.

• OSTI ID: 5782935
• Journal Information: Analytical Chemistry (Washington); (United States), Vol. 65:2; ISSN 0003-2700
• Country of Publication: United States
• Language: English

Related Subjects

37 INORGANIC
ORGANIC
PHYSICAL AND ANALYTICAL CHEMISTRY
59 BASIC BIOLOGICAL SCIENCES
DNA
FLUORESCENCE SPECTROSCOPY
NUCLEIC ACIDS
CAPILLARIES
ELECTROPHORESIS
LASER SPECTROSCOPY
QUENCHING
SENSITIVITY
BLOOD VESSELS
BODY
CARDIOVASCULAR SYSTEM
EMISSION SPECTROSCOPY
ORGANIC COMPOUNDS
ORGANS
SPECTROSCOPY
400102* - Chemical & Spectral Procedures
550200 - Biochemistry