title: Photobleaching of red fluorescence in oral biofilms.
abstract: OBJECTIVE
  Many species of oral bacteria can be induced to fluoresce due to the presence of endogenous porphyrins, a phenomenon that can be utilized to visualize and quantify dental plaque in the laboratory or clinical setting. However, an inevitable consequence of fluorescence is photobleaching, and the effects of this on longitudinal, quantitative analysis of dental plaque have yet to be ascertained.
  
  
  METHODS
  Filter membrane biofilms were grown from salivary inocula or single species (Prevotella nigrescens and Prevotella intermedia). The mature biofilms were then examined in a custom-made lighting rig comprising 405 nm light-emitting diodes capable of delivering 220 W/m(2) at the sample, an appropriate filter and a digital camera; a set-up analogous to quantitative light-induced fluorescence digital. Longitudinal sets of images were captured and processed to assess the degradation in red fluorescence over time.
  
  
  RESULTS
  Photobleaching was observed in all instances. The highest rates of photobleaching were observed immediately after initiation of illumination, specifically during the first minute. Relative rates of photobleaching during the first minute of exposure were 19.17, 13.72 and 3.43 arbitrary units/min for P. nigrescens biofilms, microcosm biofilm and P. intermedia biofilms, respectively.
  
  
  CONCLUSIONS
  Photobleaching could be problematic when making quantitative measurements of porphyrin fluorescence in situ. Reducing both light levels and exposure time, in combination with increased camera sensitivity, should be the default approach when undertaking analyses by quantitative light-induced fluorescence digital.
authors.affiliation: School of Dental Sciences, University of Liverpool, Liverpool, UK. chope@liv.ac.uk
authors.forename: C K
authors.initials: CK
authors.lastname: Hope
authors.forename: E
authors.initials: E
authors.lastname: de Josselin de Jong
authors.forename: M R T
authors.initials: MR
authors.lastname: Field
authors.forename: S P
authors.initials: SP
authors.lastname: Valappil
authors.forename: S M
authors.initials: SM
authors.lastname: Higham
mesh_terms: D001431:Bacteriological Techniques; D018441:Biofilms / Q000528:radiation effects*; D003773:Dental Plaque / Q000382:microbiology*; D005374:Filtration / Q000295:instrumentation; D005453:Fluorescence; D006801:Humans; D007091:Image Processing, Computer-Assisted; D008029:Lighting / Q000295:instrumentation; D038761:Photobleaching; D010781:Photography / Q000295:instrumentation; D011166:Porphyrins / Q000528:radiation effects; D018713:Prevotella intermedia / Q000528:radiation effects*; D045242:Prevotella nigrescens / Q000528:radiation effects*; D011829:Radiation Dosage; D012463:Saliva / Q000382:microbiology; D013997:Time Factors
chemical_list: D011166:Porphyrins