tagged: REDFIN Solo, Spot editing, warping

How to move an entire spot border to compensate for warping mistakes

Posted by ludesi in How to's in REDFIN, New features in REDFIN

If you are using 2D gel image analysis software such as REDFIN Solo, SameSpots, or Delta2D, you will be familiar with what we call the 4th generation workflow. In short, it can be summarized as consisting of the following 4 steps:

1. Image warping
2. Creation of a fusion image of all warped images
3. Spot detection in the fusion image
4. Pushing of the spot borders to all individual warped images

In such a workflow, the image warping step is the most crucial step, as this is where you, as the user, can influence the quality of the generated 2D gel image analysis data the most. This is where you correct for distortions in the gel pattern to ensure that when the spot borders are pushed back to the individual warped images, the underlying protein spots are indeed at the correct positions.

But what if they are not?

Anyone who has warped 2D gel images knows that creating perfectly warped images is an art in itself. There is always that particularly difficult area where even the most experienced of us stalls. Should you warp this spot on top of this one, or is it that one, or maybe neither? There often seems to be no right solution.

Lets also be honest and admit that sometimes we simply run out of patience to fine-tune the warping that little bit more…

Even using highly automated image warping tools can be a problem, as there is the danger of mistakes creeping in that we simply are not aware of until we inspect the results.

In short, warping mistakes happen and this is why we need spot editing tools later on in order to correct for them.

An ideal spot editing tool for 4th generation software allows you to move an entire border in order to compensate for insufficient or incorrect warping. It is against the philosophy of such workflows to create an entirely new spot border at the correct position, as then not all spot borders will be the same anymore.

Also, moving an existing spot border to the correct position is simply easier and faster – an advantage that is not to be frowned upon when you are working your way through a long list of spots…

So how do you do it?

Open your image analysis results in REDFIN and switch to the Spot View. Clicking on the Edit Spots button will open the Spot Editing Tools. Choose the Move Border tool.

Move your mouse over the spot border and left-click.

Drag the spot border so that it encompasses the underlying spot as best as possible.

Release the mouse to place the border.

Done!

REDFIN Solo will automatically re-quantify the spot and re-calculate the  statistics on the fly.

With this tool you can rest assured that any warping mistakes that results in mis-placed borders can be easily corrected, without compromising the consistency of the overal spot borders and thus the data in itself.