*School of Computer and Information, Hefei University of Technology Confocal microscopy with structured illumination and increased depth of focus 12th International Workshop on Research and Education in Mechatronics Thema 05.04.2019 htdghbhdgbdf Dipl.-Ing. (FH) Philipp Heidingsfelder#1, Prof. Dr. Jun Gao*2, Prof. Dr.-Ing. Peter Ott#3 #Institute for Mechatronics, Optical Engineering Lab, Heilbronn University *School of Computer and Information, Hefei University of Technology © HS HN • Autor (2006) • xyz.ppt

Contents Motivation Confocal Microscopy Structured Illumination Thema 05.04.2019 Contents Motivation Confocal Microscopy Structured Illumination Optical System Results Summary and Outlook 05.04.2019 © HS HN • Autor (2006) • xyz.ppt

Structured Illumination Thema 05.04.2019 Confocal Microscopy Structured Illumination Inspection of microcircuits and MEMS Optical sectioning property Reduction in the presence of flared and scattered light in the image  High-end applications in biotechnologie Improved axial and lateral resolution Great complexity Combine confocal microscopy and structured illumination with moderate effort Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019 © HS HN • Autor (2006) • xyz.ppt

Structured Illumination Principle of Confocal Microscopy Beam Splitter F´T Fobj Sample Defocused Plane Sample Detector Exit Pupil z Detector pinhole Focal Plane Tube Lens Illumination Pinhole Objective Illuminating Beam Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination Principle of Structured Illumination Projection of a sinusoidal fringe pattern on the sample  Conrast evaluation for defocussing the sample [C. Kranitzky, C. Richter, C. Faber, M.C. Knauer, G. Häusler; Institute of Optics, Information and Photonics, University of Erlangen-Nuremberg] Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination Signal Evaluation For each z-position 4 shifts of the field stop 4 irradiance values for each pixel E1, E2, E3, E4  Contrast Evaluation: [C. Kranitzky, C. Richter, C. Faber, M.C. Knauer, G. Häusler; Institute of Optics, Information and Photonics, University of Erlangen-Nuremberg] Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination CCD1 1 4 Slit Period Grating Tube Lens Tube Lens Grating Incoherent Illumination Beam Splitter Exit Pupil Objective Piezo Linear Actor Projected Grating in Focal Plane of CCD1 z Sample Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination Thema 05.04.2019 For the simulation the Optical Transfer Function (OTF) of the Optical System is needed Pattern Illumination Entrance Pupil Exit Pupil Detector Plane The Point Spread Function (PSF) is defined as O2 S2 ∆z S2' ∆z' O2' ∆W α α' O1 O1' Beam Splitter S1 S1' Object Plane NA: Numerical Aperture m: Magnitude of the microscope ∆W: Optical Path Differnce (OPD) in the exit pupil circ: Circular-Function F : Fourier Transformation λ: Wavelength Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019 © HS HN • Autor (2006) • xyz.ppt

Structured Illumination F {EP} F {PSF} IF -1{FS·OTF}I Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination CCD1 1 4 Slit Period Grating CCD2 Beam Splitter Tube Lens Grating Tube Lens Incoherent Illumination Beam Splitter Exit Pupil Objective Piezo Linear Actor Projected Grating in Focal Plane of CCD1 z Sample Focal Plane CCD2 Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination Simulated Focal Depth Response for focused (CCD1) and defocused camera (CCD2) Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination CCD_1 1 4 Slit Period Grating CCD_2 Beam Splitter Tube Lens Grating Tube Lens Incoherent Illumination Beam Splitter Exit Pupil Objective Piezo Linear Actor Focal Plane CCD_1 Projected Grating z Sample Focal Plane CCD_2 Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination Light Source CCD Camera Grating Step Motor and Piezo Actor Tube Lens Beam Splitter Mirror Objective Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination Step Height Standard: 14.33 µm Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination Softmark Structured Illumination Mode Confocal Microscopy Mode Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination Summary Interesting possibility to combine structured illumination with confocal microscopy. Possibility to accelerate the measuring time with second camera. Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination Outlook Realization of the 2-camera-system Downsizing the optical system Analyzing the influence of the pattern illumination and compare it with the waveoptical model Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019

Structured Illumination Thema Thank you very much for your attention! 05.04.2019 Motivation Confocal Microscopy Optical System Results Summary Outlook Structured Illumination 05.04.2019 © HS HN • Autor (2006) • xyz.ppt