蔡司 激光共聚焦显微镜

The second exhibition of Carl Zeiss confocal laserscanning microscopeConfocal MicroscopyDr. Matthias Vaupel,Carl Zeiss MicroImaging GmbH,Göttingen, GermanyLSM 700Confocal Principledetectorpinholelenssample beamsplitterlaser mirrorsxyzfocus Main differences in comparison toclassical light & video microscopy:1) Illumination source:Laser instead of lamp2) Image acquisition:Sequential scanning of the samplewith a laser beam 3) 3D Microscopy:Height information by opticalslicingLSM 7003D Imaging: SolarSolar panel laser scribeThe confocal microscopes LSM 700 and CSM 700 –What are they good for?3D microstructure analysis, e. g.z-height propfilexyz-distances and angles intopographyroughnessthin transparent film thicknessvolume parametersbearing area curve, amplitudedensity functionImage processing and analysisReportingMaterials:Soft matter, i.e. polymersMetalPaperGlassSpecifications CSM 700Height measurement:repeatability(1σ) 0.020 μm16-bit resolution in heightHeight measurement15 mm maximum rangeMaximum sample height63 mmLateral measurement:resolution0.160 μmImage pixel resolution1280 x 1024 pixelsIllumination Xenon-Lamp(wavelength range: 400 to 700 nm)Acquisition speed7.5 fps(High-speed) ...100 fps(Wire-frame mode) Color depth3 x 8 bit(RGB)USPs and Top Features of CSM 700USPs:Confocal images in true color-> topo& colored texture (all in focus image) in just 1 step! High-speed-mode 7.5 fps (< 30 sec) at 1280 x 1024 pixelsZeiss objectives give highest Zeiss optical performanceTops:Complete Software packet, incl. Image Processing, Macro Programming, Image Stitching, ReportingFilm thickness measurementAvailable with motorized stage for scanning and stitching of large imagesSpecifications: LSM 700 on AxioScope orAxioImagerHeight measurement:repeatability0.020 μm with closed-loop-detector(Heidenhain-detector) and Axio Imager, 0.080 µm on Axio Scope mot16-bit resolution in heightHeight measurement appr. 15 mm maximum rangeMaximum sample height63 mm with Axio Imager2Lateral measurement:resolution0.120 μmImage pixel resolution4 x 1 ...2048 x 2048 pixelsrepeatability0.010 μmzoom 1x (40x)Acquisition speed5 fps(at 512 x 512 pixels, 154 fps at 512 x 16 pixels)Illumination Laser, 405 nm typical for material applicationsUSPs and Top Features of LSM 700USPs:Additional contrasting methodes: Darkfield, Pol., C-DIC, TICFluorescence: e.g. for porosity measurement and for profilometry with unlimited surface slope due to auto-fluorescence detectionZeiss objectives give highest optical performancespectral imaging of light emitted from the sampleTops:Zoom, high pixel number, and high lateral resolutionFilm thickness measurementAvailable with motorized stage for scanning and stitching of large imagesComplete Software packet, incl. Image Processing, Macro Programming, Image Stitching Additional application tools available via AxioVision: Automatic Analysis of Grains, and of nonmetallic inclusions in steelDefine the no. of slices or slice distance or set according toZXtilted surface in original z-image plane surface after automatic z-levellingstep-height of small structures (e.g. stripes) is conserved with z-levellingArtefact-free z-stack record in a field of view with strong contrastProblem:Sample surface areas with very lowand high reflectivity in the same field of viewSolutions:Change NA via objectiveApply F/Z Noise Cut or “Fill Holes”filter and interpolate incorrect pixels Record several z-stacks with different exposure time/gain factor/lightintensity Possible reasons:scattered light due to roughnesslateral inhomogeneous material distribution surface slope angle is larger than half of the acceptance angle of the objectiveOvercome the theoretical diffraction limitation of surface tilt angle by fluorescenceFluorescence ModePolymer Foil in Orthogonal Projections (Epiplan-Neofluar 50x/0.8)x Reflection Modeyz zλ= 543 nmProfilometry with unlimited surface slope due to auto-fluorescence detectiondisplays removedfrom the height map to profiles and 3D visualisationHeight mapProfile3D ViewRoughness and Waviness –a practical examplewaviness of the road:average height differencealong the road, typicallymeasured on the km-distance scaleroughness of the road:average grain size of theasphalt, typically measuredon the mm-scale©Vaupel©Vaupeluninspiring andambitious formulasahead()∫=l sk dx x z l R 031()∫=l q dx x z l R 021()∫=lku dx x z l R 041()∫=l a dx x z l R 01asphalt, typically measuredRoughness and waviness –Fourier FilteringPrimary profile:P(Section Curve)Roughness:RWaviness:WMeasurement items for roughness curves –Examplelubricant reservoirshigh friction and wear quicklyMeasurement items for roughness curves –BAC: Bearing Area Curve (Load Curve)Curve of the load length ratio of the roughness curve expressed as a function of cutting level cused for QC of steel productionOnly available in ZEN and CSM-Software !Roughness in ZEN 2009Roughness –StandardsThe definitions of roughness parameters are from DIN 4287Profiles2D standards exist only for tactile instruments.In those standards are described:- 1. Type of sensor (tip diameter, material)- 2. Type of acquisition (scan length)- 3. Type of processing (filter, cut-off wavelength)-Standards don‘t exist for optical instruments.Topographies3D standards neither exist for tactile nor for optical methods.Tactile and optical confocal scanning –in Comparison。