Measuring the purity of low volumes of DNA at 4 °C using the Agilent Cary 60 UV-Vis spectrophotometer with fiber optics microprobe

Significance of the Topic

The precise assessment of DNA purity in minimal volumes at refrigerated temperatures is critical for many biotechnological and molecular biology applications. Traditional spectrophotometric methods often require cuvettes and ambient temperature measurements, introducing handling time and potential sample degradation.

Objectives and Overview of the Study

This study demonstrates how the Agilent Cary 60 UV-Vis spectrophotometer with a fiber optics microprobe enables direct measurement of DNA purity at 4 °C. The goal is to evaluate accuracy, reproducibility, and time efficiency compared to conventional cuvette-based assays.

Methodology and Instrumentation

A Cary 60 UV-Vis instrument equipped with a high-intensity xenon flash lamp and fiber optics accessory was used. Key components included:

• Cary 60 UV-Vis with WinUV software
• Fiber optic microprobe and probe coupler
• Quartz cuvettes for calibration
• Calf thymus DNA standards in MilliQ water

Samples (0–50 µg/mL DNA) were analyzed directly at 4 °C in Eppendorf tubes under ambient fluorescent lighting. Spectral scans from 340 to 240 nm identified the 260 nm absorbance peak, and the 260/280 ratio was calculated automatically using the WinUV software module.

Main Results and Discussion

Measurements of five DNA concentrations were completed in under two minutes, achieving a linear response (Abs = 0.02004×Conc; R² = 0.9976) in accordance with the Beer–Lambert law. Cold-temperature readings eliminated sample warming delays and reduced contamination risk associated with cuvette cleaning. The high-intensity xenon lamp provided stable signals unaffected by ambient light.

Benefits and Practical Applications

This fiber optics approach offers:

• Direct analysis of refrigerated samples without cuvettes
• Significant time savings (>80% reduction in analysis time)
• Minimal sample volumes (as low as 40 µL)
• Broad temperature range (4–110 °C) free from condensation artifacts
• Capability to measure solids, powders, and gels via reflectance probes

Future Trends and Possible Applications

Further integration of fiber optics UV-Vis with automated workflows may enhance real-time quality control in bioprocessing. Advances in probe miniaturization and coupling with microfluidic platforms could extend applications to single-cell analysis and high-throughput screening.

Conclusion

The Agilent Cary 60 UV-Vis spectrophotometer with fiber optics microprobe delivers rapid, accurate, and reproducible DNA purity measurements at 4 °C, streamlining laboratory workflows and reducing costs. Its versatile design supports a wide range of sample types and temperatures, making it an invaluable tool for modern analytical laboratories.

References

1. Warburg O. and Christian W. (1942) Biochem Z. 310:384.
2. Sambrook J., Fritsch E.F., and Maniatis T. (1989) Molecular Cloning: A Laboratory Manual, 2nd Ed., Cold Spring Harbor Laboratory Press.
3. Keighley B. and Fyfe D. (1995) Application Note #91, Agilent Technologies.