Image Analysis Report
CODEX® Processor will generate an image analysis report(s) that provides an overview of each experiment processed.
This feature is available in version 1.7.
Report(s) will be saved to the root output folder. Each region will have a corresponding PowerPoint.
The first three pages will summarize the experiment metadata and processing parameters. Important details such as the number of biomarkers (e.g. 18-plex) and processor version (e.g. 1.7) can be found here.
Next section contains a histogram and sample image for every acquired channel. Each row is a cycle, containing its corresponding channels. Top-left image and histogram is always the first cycle, first channel. Image and histogram to their right is the same cycle, second channel.
The displayed image is a full-resolution cropped image that has the greatest standard deviation among those sampled. Each image may be of different location within the region, depending on the expression pattern. For example, cropped images for DAPI cycles 2 and 3 below differ in their sampling location.
(Magenta) A single cycle. (Cyan) Full-resolution cropped image. (Purple) Histogram.
The histogram displays pixel frequency of signal intensities found within the region image. The horizontal intensity axis covers the 16-bit range (0 ~ 65535), while the vertical frequency axis auto-scales to the number of pixels found. Frequency is in logarithmic scale to better represent lower frequencies that correspond to biomarker signal.
Additionally, all cycles except the first contains two histogram plots. All purple plots are of the first cycle, which can be a reference for comparison, especially if the first cycle contains blank cycles. The legend specifies which biomarker the histogram represents.
For easier navigation between biomarkers, images and histograms here are linked to individual biomarker slides. You can:
- Ctrl-click on the images or histograms or,
- Shift-F5 to enter presentation mode and left-click on the images or histograms.
You can return to the histogram slides by clicking the main region image.
Summary section contains basic statistics of biomarker region images. Again, each slide contains four cycles and their corresponding channels. Any non-biomarker channel (i.e. containing "DAPI," "blank" or "empty") will be grayed out.
There are 18 columns for each channel. All pixel intensities are in 16-bit ranges (0 ~ 65535).
Exposure time in milliseconds
Minimum pixel intensity of the region image
Median pixel intensity
95-percentile pixel intensity
Max pixel intensity
Mean pixel intensity
Binarization threshold found by Otsu's method
Area of the region image above the threshold (i.e. signal area)
Mean pixel intensity of the area above the threshold (i.e. signal area)
Standard deviation of the signal area
Mean pixel intensity of the area below the threshold (i.e. background area)
Standard deviation of the noise area
Mean of the signal divided by standard deviation of the noise
These values can be used as a basic guideline to image quality of each biomarker. As the threshold is not adaptive nor trained, the summary information should be considered within context. Explanation of Otsu's binarization can be found here.
The final section contains a detailed slide for each marker acquired. The left-hand side contains image information identical to those shown in the histogram and summary slides.
The main image in the center is the region image scaled for best fit and viewing on 4K displays (i.e. scaled to 288 ppi). This image is not in its original resolution and should not be used for detailed analysis.
(Magenta) Image information. (Cyan) Region image. (Purple) Cropped images.
On the right, there are four cropped images displayed in the original, full resolution. Within the whole region, 13 different areas are cropped at full resolution and compared for their intensity standard deviation. The four areas with the highest standard deviations, which often correspond to most contrast and information, are displayed in order.
The main image links back to the corresponding histogram slide, which can be control-clicked or left-clicked in presentation mode.