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OS-SIM - Optical Sectioning Structured Illumination Microscopy

Learn how to build a low-cost, DMD-based Structured Illumination Microscope (SIM) for optical sectioning. This system removes out-of-focus blur and dramatically improves image contrast in fluorescence microscopy.

What is Optical Sectioning SIM?​

Optical Sectioning SIM (OS-SIM) illuminates the sample with structured patterns (stripes) instead of uniform light. By acquiring multiple images with different pattern phases and combining them mathematically, you can extract in-focus information while suppressing out-of-focus background.

The RMS Reconstruction Algorithm​

Three images are acquired with phase shifts (0°, 120°, 240°), then combined:

ISIM(x)=(I1−I2)2+(I2−I3)2+(I1−I3)2I_{\text{SIM}}(x) = \sqrt{(I_1 - I_2)^2 + (I_2 - I_3)^2 + (I_1 - I_3)^2}

This computation removes out-of-focus blur, effectively creating optical sections without physically scanning.

Two Configurations​

Direct Projection Setup​

The DMD pattern is directly imaged onto the sample. Simple and easy to align, but includes all diffraction orders.

4f Interference Setup​

Uses a Fourier filter to remove higher diffraction orders, creating cleaner interference patterns for improved optical sectioning.

What You'll Learn​

  • Principles of structured illumination microscopy
  • DMD (Digital Micromirror Device) control
  • Optical sectioning without mechanical scanning
  • Pattern projection and phase shifting
  • Image reconstruction algorithms
  • 4f optical relay systems
  • Fourier filtering techniques

Tutorials in this Section​

  • OS-SIM Tutorial - Complete guide to building a DMD-based SIM system

Key Components​

Optical Components​

  • 488nm laser (fiber-coupled)
  • 10× Plan Objective (NA = 0.25)
  • Tube lens (f = 100mm)
  • Projection lens (f = 200mm for direct setup)
  • Relay lenses (2× f = 50mm for 4f setup)
  • Dichroic mirror and emission filters
  • Spatial filter (for 4f configuration)

Electronics​

  • DMD: DLP300S chip + DLPC1438 controller (from Anycubic Photon Ultra)
  • Controller: Raspberry Pi Zero 2
  • Interface: Custom bridge board (based on OpenMLA)
  • Power: 12V power supply

Mechanical​

  • OpenUC2 cubes and mounting plates
  • Custom DMD mounting parts

How It Works​

  1. Pattern Generation: DMD creates structured patterns (stripes)
  2. Projection: Pattern is imaged onto the sample
  3. Phase Shifting: Pattern shifts in 3 steps (0°, 120°, 240°)
  4. Image Acquisition: Camera captures one image per phase
  5. Reconstruction: Mathematical combination removes out-of-focus light

Safety Warning ⚠️​

Laser Safety: This system uses a 488nm laser. Always wear appropriate safety goggles.

DMD Diffraction: The DMD acts as a grating, creating multiple diffraction orders. Ensure all stray beams are properly blocked to prevent eye exposure.

Key Advantages​

  • No Mechanical Scanning: Purely electronic pattern switching
  • Low Cost: Uses harvested DMD from 3D printers
  • UC2 Integration: Works with existing UC2 fluorescence setups
  • True Optical Sectioning: Removes out-of-focus blur computationally
  • Fast Acquisition: Electronic pattern switching is rapid

Applications​

  • Fluorescence microscopy with improved contrast
  • Thick sample imaging without confocal scanning
  • Live cell imaging with optical sectioning
  • Educational demonstrations of computational microscopy
  • Research into structured illumination techniques

Technical Capabilities​

  • Optical sectioning without Z-scanning
  • Pattern frequency adjustable via DMD programming
  • Compatible with various fluorophores (488nm excitation)
  • Real-time or post-processing reconstruction
  • Integration with existing wide-field systems

Perfect for researchers and educators interested in advanced fluorescence microscopy techniques without the cost of commercial systems!