Calibration

Calibration can be done at any time. However, quantitative measures that involve x and y components in their calculation (i.e., flat area and volume) will not be accurate until the series is calibrated. If a series is calibrated after traces have been made, the contours will be resized proportionately with the underlying images.

Instruction	Image(s)
Go to the calibration grid section (typically section 0).
2. Name two palette squares calibrate_x{1} and calibrate_y{1}. Note: Using curly braces in trace palette names allow you to easily change the number associated with each trace by clicking the plus button to the right of the palette. For example, "calibrate_x1" becomes "calibrate_x2" after incrementing with the + button.
3. Zoom in until you see about 12 boxes across your screen. That is how close you should be in order to accurately trace across 10 boxes from corner to corner. Avoid extreme edges and far corners, as this is where most distortion occurs.
4. With the calibrate_x{1) palette selected, use the "open trace" tool, and trace across 10 boxes from corner to corner in the x-direction. (link to open trace section here)
5. Select the calibrate_y{1} palette, and repeat #4 by tracing across 10 boxes in the y-direction.
6. Move to another region on the calibration grid. Use the "+" button to the right of the palette to automatically increase your numbers in curly braces by +1, and lay down your second -x and -y calibration traces by repeating steps 4 and 5 above.
7. Repeat step 6 until you have a total of 5 calibration traces in x and y.
8. When your traces are complete, select all your traces (Ctrl+A).
9. From the series dropdown menu, select "Calibrate pixel size."
10. A dialogue box will appear asking for the length of your calibration trace in microns. If you are in the Harris lab, enter 4.63 and select ok. This dialogue box will pop up for every calibration trace you have selected. Continue to enter 4.63, and select ok. Once you have done this for all of your traces, PyReconstruct will automatically change the series magnification by averaging your calibration traces (a progress bar will appear informing you that this is occurring). *IMPORTANT NOTE* This number will depend on the calibration grid used. For the grids commonly used in the Harris lab, each box is 0.463 microns. Therefore, the length of 10 boxes (your calibration trace) is 4.63 microns.
11. After series calibration all of your existing traces, including your calibration traces, will be scaled proportionately with the image magnification. You can check to see if a series has been calibrated by opening the trace list, and checking the length of your calibration traces. They should be close to 4.63 microns.

Importing / exporting

Traces

When selecting Series > Import > Traces..., you will be directed to the following menu:

Series

This is where you will locate the jser file for the series you would like to import from (i.e. the importing series).

Object Regex Filters

The regex filters for the specific objects you would like to import. Leave this field blank if you would like to import all objects.

Section Range

The sections whose trace data you wish to import.

Duplicate Threshold

The amount of overlap between two traces necessary for the traces to be considered functional duplicates of each other.

When importing traces from another series, traces between the current and importing series are cross-checked so that no duplicate traces are imported. Any two traces with the same name and share a fractional overlap reaching at least the duplicate threshold are considered functional duplicates, and only the trace from the current series is kept.

It is highly suggested to keep this threshold between 0.9 and 1.

Favor series

In the case of two traces of the same name between the series that overlap, but are not duplicates, the trace belonging series marked as favored will be kept, while the other is discarded.

Check Series Histories

If this option is checked, the histories of each series will be reviewed when importing traces. In the case where the traces on one series have been modified and those on other series have not, the traces on the series that has undergone modification will be used by default.

For example: for trace d01 on section 5, if the current series has been modified since the divergence between the two series but the importing series has not, d01 traces on section 5 of the current series will be used by default.

Flag Import Conflicts

If this option is checked, the program will flag any remaining traces that have not been resolved through any of the checks listed above.

KH Lab Wiki

Series Operations