A cryogenic optical waveguide spectrometer for the measurement of low-temperature absorption spectra of dilute biological samples.
A cryogenic optical waveguide spectrometer that uses a Teflon-AF 2400 liquid core waveguide is described. In comparison to standard low-temperature absorption techniques, the liquid core waveguide approach not only affords the use of microliter samples but also provides significant improvements in sensitivity. Here we show low-temperature absorption spectra of various flavoproteins, including DNA photolyase, measured using this new technique. The technique has high reproducibility and can afford the detection of 15 ng of flavoprotein. In addition, the technique requires several hundredfold less protein than standard low-temperature techniques for the same sensitivity. The performance of the spectrometer in the ultraviolet (UV) region is investigated experimentally and compared with standard UV absorption techniques. Results indicate that, below 300 nm, the observed absorbances deviate from the Beer-Lambert law.
Siddiqui
M Salim U
MS
Department of Biochemistry and Department of Chemistry, Temple University, Philadelphia, PA 19122, USA.
Stanley
Robert J
RJ
D003080:Cold Temperature; D004255:Deoxyribodipyrimidine Photo-Lyase / Q000032:analysis; D004867:Equipment Design; D029968:Escherichia coli Proteins / Q000032:analysis; D005336:Fiber Optic Technology; D005420:Flavoproteins / Q000032:analysis; D011138:Polytetrafluoroethylene; D015203:Reproducibility of Results; D013053:Spectrophotometry / Q000295:instrumentation*
D029968:Escherichia coli Proteins; D005420:Flavoproteins; D011138:Polytetrafluoroethylene; D004255:Deoxyribodipyrimidine Photo-Lyase