title: Obtaining artificially caries-affected dentin for in vitro studies.
abstract: OBJECTIVE
  This study evaluated and improved a protocol for obtaining standard caries-affected dentin (CAD) by Streptococcus mutans biofilm demineralization process.
  
  
  METHODS
  Forty-eight human molars were divided in six experimental groups, according to: period of cariogenic challenge (7, 14 or 21 days) and type of dentin (erupted or unerupted teeth). After complete cariogenic challenge sound and CAD dentin were evaluated by: visual inspection (VI), digital radiography (DR), optical coherence tomography (OCT) and laser fluorescence (LF).
  
  
  RESULTS
  Visual inspection confirmed the formation of CAD based on tissue yellowing and loss of surface gloss. Digital radiography detected the presence of radiolucent images, suggesting caries. Three calibrated examiners viewed all images obtained by VI and DR and were able to distinguish healthy from CAD. Fisher's exact statistical test (p < 0.05) confirmed no difference between groups by VI (G1/G4: p = 0.6; G2/G5: p = 1; G3/G6: p = 1) or DR (G1/G4: p = 1; G2/G5: p = 1; G3/G6: p = 1). Both LF values and demineralization depth, as determined by OCT, were subjected to ANOVA (p < 0.05). For LF, a statistically significant difference was observed for the type of substrate (p = 0.001). For OCT, no statistically significant differences in the type of substrate (p = 0.163), length of cariogenic challenge (p = 0.512) or interaction between factors (p = 0.148) were observed. Scanning electron micrographs confirmed the presence of CAD; a more uniform demineralization surface was observed in the dentin of unerupted teeth.
  
  
  CONCLUSIONS
  This protocol suggests that standard CAD can be obtained in 7 days of cariogenic challenge using unerupted teeth. Clinical significance: With the new perspective on the clinical treatment of caries lesions, bonding is increasingly performed to demineralize CAD, which is susceptible to remineralization. A useful protocol to standardize the production of CAD, by microbiological cariogenic challenge, would be an important contribution to laboratorial test in the field of operative dentistry.
authors.affiliation: PhD Student, Department of Operative Dentistry, University of São Paulo São Paulo, Brazil.
authors.forename: Cynthia Soares
authors.initials: CS
authors.lastname: de Azevedo
authors.affiliation: BDS, Department of Operative Dentistry, University of São Paulo São Paulo, Brazil.
authors.forename: Bruna Uglik
authors.initials: BU
authors.lastname: Garbui
authors.affiliation: BDS, Department of Operative Dentistry, University of São Paulo São Paulo, Brazil.
authors.forename: Caroline
authors.initials: C
authors.lastname: Martins e Silva
authors.affiliation: Assistant Teacher, Institute of Biomedical Sciences, University of São Paulo, São Paulo, Brazil.
authors.forename: Maria Regina
authors.initials: MR
authors.lastname: Simionato Lorenzetti
authors.affiliation: Researcher and PhD, Department of Lasers and Applications, Nuclear and Energy Research, São Paulo, Brazil.
authors.forename: Anderson Zanardi
authors.initials: AZ
authors.lastname: de Freitas
authors.affiliation: Full Professor, Department of Operative Dentistry, University of São Paulo São Paulo, Brazil.
authors.forename: Adriana Bona
authors.initials: AB
authors.lastname: Matos
mesh_terms: D018441:Biofilms; D019411:Clinical Laboratory Techniques; D003731:Dental Caries / Q000382:microbiology / Q000473:pathology*; D003804:Dentin / Q000382:microbiology / Q000473:pathology*; D005453:Fluorescence; D006801:Humans; D066298:In Vitro Techniques; D007834:Lasers; D008855:Microscopy, Electron, Scanning; D010808:Physical Examination; D019252:Radiography, Dental, Digital / Q000379:methods; D013295:Streptococcus mutans / Q000502:physiology*; D013997:Time Factors; D041623:Tomography, Optical Coherence / Q000379:methods; D017001:Tooth Demineralization / Q000382:microbiology; D014097:Tooth, Unerupted / Q000382:microbiology / Q000473:pathology