Advanced Image Processing using Imaris (25. 11. 2008)

Urs Ziegler ziegler@zmb.uzh.ch

Zentrum für Mikroskopie und Bildanalyse

Imaris – integrated modules

• Open and Save data

• Contrast and BrightnessDisplay Adjustments

• Navigation through dataSliceSectionEasy 3D

• Snapshots

• Surpass

• Isosurface

• Spot detection

• Filament tracing

• Contour

• Statistics and Measurements

• Animations

Todays seminar

Surface rendering: Isosurface

Identification of voxelsbelonging to an object. The criteria for the identification is the grey value of the voxel: threshold value.

grey level < threshold value = background

grey level > threshold value = object

surface voxel = voxel belonging to an object while some of its neighbors do not.

Isosurface

Data displayed as a volume (MIP or Blend) with intensities adjusted that only structures of interest are visible

→ Threshold in isosurface rendering

Isosurface

Select a new isosurface

Select channel

Select if smoothing and / or background subtraction should be performed

Isosurface

Select threshold

(optional: split touching objects)

Spot detection

Data displayed as a volume (MIP or Blend) with intensities adjusted that only structures of interest are visible

→ Measure diameter of structures in slice mode

Spot detection

Filter detected spots according to selected criteria (Quality ~intensity at center)

Tracking spots

1. Select Track Spots over time

2. Select which type of motion model should be applied

Spot trackingMotion models

Brownian Motion

• This algorithm models the motion of each Spot as a Brownian motion. This model is appropriate, if your Spots actually perform a Brownian motion.

Autoregressive Motion

• This algorithm models the motion of each Spot as an autoregressive process. This model is appropriate if your Spots perform any kind of continuous motion.

Filter detected tracks according to selected criteria

Filtering, statistics and counting

Example: Chlamydia infected cells

• Counting cells based on DNA stained nuclei

• Evaluation of infection based on the size of inclusion (fluorescently labeled Chlamydia)

• Attention: DNA found in nuclei and Chlamydia

• 1. Detect nuclei via spot function

Spots are only detected in the nuclei because of filtering for low intensity in channel 2 (green labeled Chlamydia)

Filtering, statistics and counting

Example: Chlamydia infected cells

• Counting cells based on DNA stained nuclei

• Evaluation of infection based on the size of inclusion (fluorescently labeled Chlamydia)

• Attention: DNA found in nuclei and Chlamydia

• 2. Detect inclusions via isosurface

Isosurfaces are filtered for size (>100um, green labeled).

Filtering, statistics and counting

Example: Chlamydia infected cells

• Counting cells based on DNA stained nuclei

• Evaluation of infection based on the size of inclusion (fluorescently labeled Chlamydia)

• Attention: DNA found in nuclei and Chlamydia

• View or export values / statistic

Individual objects can be selected

Animation

Add a keyframe Number of frames

Animation can be designed using keyframes which store scenes shown in the display. Frames between keyframes are interpolated.

Keyframes (blue, white if selected)

Save animation (format: uncompressed avi for ease of use)

Questions and Outlook

Next lunch seminar: Tuesday, 20th of January 2009

Advanced Image Processing: Spot Detection

Animation Generation

If time: starting Deconvolution

Questions and input: www.zmb.uzh.ch or ziegler@zmb.uzh.ch

Imaris

• 7 floating licenses bought by several research groups (2005)

• License server maintained by the computing center:2701@idlicense1.uzh.ch

• Maintenance of Imaris paid by the Center for Microsocpy and Image Analysis

• Download of Imaris and coordination for acquisiton of new modulesand licenses at the University via the Center for Microsocpy and Image Analysis: :