title: Optimization of the measurement procedure during multiphoton tomography of human skin in vivo.
abstract: BACKGROUND
  The in vivo multiphoton tomography has evolved into a useful tool for the non-invasive investigation of morphological and biophysical characteristics of human skin. Until now, changes of skin have been evaluated mainly by clinical and histological techniques. The current study addresses the effects of a changed acquisition time for single scans in a Z-stack on the directly related qualitative and quantitative interpretability of the data.
  
  
  METHODS
  A test area of the skin was used for scanning 12 Z-stacks of 10 volunteers aged between 25 and 34 years. The stacks were taken up to a depth of 220 μm at increments of 10 μm at four different times, 1, 3, 7, 13 s, per scan. Subsequently, the second harmonic generation (SHG)-to-autofluorescence aging index of dermis (SAAID) was evaluated at three different measuring depths, i.e. at the maximum of SHG as well as at depths of 60 and 150 μm.
  
  
  RESULTS
  The evaluation did not reveal any significant differences in the SAAID behavior between the Z-stacks of each test area scanned at different acquisition times. However, the acquisition time of 1 s/frame increases the measurement stability without influencing the SAAID behavior. The resolution of subcellular structures decreases significantly at scan times ≤3 s, whereas the acquisition time from 7 to 13 s warrants a high image quality.
  
  
  CONCLUSIONS
  The study has shown that there are no significant differences between the scan speeds per scan in a Z-stack and the resulting SAAID. Acquisition times of 7 s are suitable for the morphological evaluation whereas a further extension to 13 s does not result in any benefits. A scan time per image of 1 s is sufficient for the quantitative evaluation of SAAID thus substantially reducing the possible influence of movement artifacts.
authors.affiliation: Department of Dermatology and Allergology, University Hospital, Jena, Germany.
authors.forename: S
authors.initials: S
authors.lastname: Springer
authors.affiliation: Department of Dermatology and Allergology, University Hospital, Jena, Germany.
authors.forename: M
authors.initials: M
authors.lastname: Zieger
authors.affiliation: Faculty of Physics and Mechatronics, Saarland University, Saarbrücken, Germany.|JenLab GmbH, Jena, Germany.
authors.forename: K
authors.initials: K
authors.lastname: Koenig
authors.affiliation: Department of Dermatology and Allergology, University Hospital, Jena, Germany.|Department of Dermatology and Allergology, SRH Waldklinikum Gera gGmbH, Gera, Germany.
authors.forename: M
authors.initials: M
authors.lastname: Kaatz
authors.affiliation: Center of Experimental and Cutaneous Physiology (CCP), Department of Dermatology, Venereology and Allergology, Charité-Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: J
authors.initials: J
authors.lastname: Lademann
authors.affiliation: Center of Experimental and Cutaneous Physiology (CCP), Department of Dermatology, Venereology and Allergology, Charité-Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: M E
authors.initials: ME
authors.lastname: Darvin
mesh_terms: D000328:Adult; D000465:Algorithms; D005260:Female; D006801:Humans; D007089:Image Enhancement / Q000379:methods*; D000069416:Intravital Microscopy / Q000379:methods*; D008297:Male; D036641:Microscopy, Fluorescence, Multiphoton / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012867:Skin / Q000166:cytology*; D041622:Tomography, Optical / Q000379:methods*
keywords: AF; SAAID; SHG; SHG-to-AF aging index of dermis; autofluorescence; multiphoton tomography; second harmonic generation