Abstract

Sensing biochemicals with optical fibers has been of interest in clinical analysis for some time. Most efforts to date, however, have been confined to passive, multimode ‘optrodes,’ which merely conduct an optical signal to and from the sensing region.1 The use of the evanescent field of the optical fiber for sensing presents definitive advantages, although the inherent problem of the evanescent field is to have the external information captured efficiently, whether it be fluorescent energy or a transmission modulation. Multimode fibers have limitations in absorption-based systems,2 as they do not adhere to Beer’s law, and their fluorescent energy capture rates are generally low. To overcome these problems, we have used a tapered single-mode optical fiber. An evanescent field sensor for measuring penicillin based on the immobilization of Bacillus Cereus penicillinase (Type 1, E.C. 3.5.2.6) on the surface of a tapered single mode optical fiber has been developed. York Technology SM-450 fiber, with a cutoff wavelength of 450 nm, was tapered adiabatically over an oxybutane flame.3 Tapers of waist diameter less than 2 μM with transmission losses of no more than 0.1 dB were used. Penicillinase labelled with fluorescein isothiocyanate (FITC) was adsorbed to the tapered fiber. The immobilized fluorophores at the core/cladding interface serve to optimize the total fluorescence captured.4 In a comparative quantification of samples of FITC bound to penicillinase and FITC in bulk solution using a fluorimeter, approximately 6 FITC molecules were found to be bound to each penicillinase. FITC- labelled penicillinase activity was verified using the iodometric enzyme assay of penicillinase5 with unmodified penicillinase as the control. The FITC/penicillinase was then attached to the tapered single-mode fiber by incubation overnight in a 0.2 M sodium phosphate buffer pH 5.9, and the unbound solution removed through repeated washings with buffer solution.

© 1994 Optical Society of America

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References

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