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Copy file name to clipboardExpand all lines: docs/03_Investigator/02_FRAME/config_examples/Optical_Alignment_Procedures.md
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## Overview
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The FRAME system requires precise optical alignment to ensure optimal performance. This document provides detailed procedures for aligning the internal optics cube and calibrating the optical path.
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The FRAME system requires precise optical alignment to ensure optimal performance. This document provides detailed procedures for aligning the internal optics module and calibrating the optical path.
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## Internal Optics Cube Preparation
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### Initial Setup Outside the FRAME
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Before installing the internal optics cube in the FRAME, it must be properly aligned using a separate alignment setup.
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Before installing the internal optics module in the FRAME, it must be properly aligned using a separate alignment setup.
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#### Components to Align
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The internal optics cube contains three main optical elements that must be aligned:
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The internal optics module contains three main optical elements that must be aligned:
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1.**Transmitted light module** with camera and folding mirror
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2.**Fluorescence excitation module**
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3.**Objective mounting system**
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#### Alignment Setup
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1.**Create External Test Setup**:
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- Remove the internal optics cube from the FRAME
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- Build a test setup on top of the cube that mimics the FRAME's optical distances
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- Remove the internal optics module from the FRAME
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- Build a test setup on top of the module that mimics the FRAME's optical distances
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- Mount an objective in the test setup to maintain proper optical spacing
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2.**Software Preparation**:
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- Start ImSwitch software to enable camera viewing
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- Ensure proper communication with the cube's electronics
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- Ensure proper communication with the module's electronics
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### Fluorescence Module Alignment
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#### Mechanical Alignment
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1.**Access Adjustment Screws**:
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- Use a 2.5mm hex key to access the three adjustment screws on the back of the cube
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- Use a 2.5mm hex key to access the three adjustment screws on the back of the module
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- These screws control the position and angle of the optical elements
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2.**Laser Centering**:
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- Ensure the laser focus is exactly centered where the objective will be positioned
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- The light beam must exit the internal cube parallel to the optical axis
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- The light beam must exit the internal module parallel to the optical axis
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- This ensures proper Köhler illumination
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3.**Objective Installation**:
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### Objective-to-FRAME Alignment
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After internal cube alignment, align the optical axis of the FRAME with the internal cube.
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After internal module alignment, align the optical axis of the FRAME with the internal module.
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#### Available Adjustments
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Two adjustment mechanisms are available:
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1.**X-axis**: Objective can be moved along the X-axis using the slide mechanism
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2.**Y-axis**: Internal cube can be moved relative to the FRAME using adjustment screws
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2.**Y-axis**: Internal module can be moved relative to the FRAME using adjustment screws
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#### LED Alignment Tool Setup
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- Observe the point light source projected to the back focal plane
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2.**Y-axis Alignment**:
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- Adjust the internal cube position along Y-axis using the adjustment screw
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- Adjust the internal module position along Y-axis using the adjustment screw
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- Move the light point to the center of the back focal plane
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- Use a piece of paper with crosshairs to mark the position
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Once all optical elements are properly aligned:
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1.**Secure Internal Cube**:
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- Tighten the locking knobs to permanently secure the internal cube in the rail
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- Ensure the cube cannot move during operation
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1.**Secure Internal Module**:
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- Tighten the locking knobs to permanently secure the internal module in the rail
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- Ensure the module cannot move during operation
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2.**System Testing**:
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- Perform comprehensive testing of all optical functions
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