A report on an experiment I did of doing electrophoresis with proteins

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H. Image analysis on the gel
After the gel has been destained the objective is to quantify the mass of the proteins from the instensity of the dye
that is stuck to it. An exact quantification has its own complications and I shall explain how far one can go in this
goal.
Basically the idea is to see how the intensity of the gel varies as a function of the protein mass and this requires a
way of quantifying the intensity. For this the gel is photographed and the sum of the pixel values is taken as a proxy
for the intensity since one knows that bighter points in the picture have higher pixel values. Then all analysis can be
done using this set of numbers as obtained by the following process of “digitizing” the gel.
The exact steps in the process are as follows,
• The last destaining solution is not transferred out of the tray.
• A pair of soft smooth transparent plastic sheets is cut out of the size little bigger than the gel and these are
joint at one edge. Usually this is made by cutting out from the edge of plastic packets.
• Keeping the gel dipped inside the solution a hard plastic is slid underneath it and the gel is lifted out of the
solution on it. This method ensures that no air bubbles are trapped between the gel and the plastic.
• Then gently the gel is slipped on to one of the pair of soft palstic sheets from the hard plastic by sliding it
onto it keeping the leading edge always in touch with a thin film of water. This ensures that no air bubbles get
trapped in this stage.
• The second plastic film is covered over it in a similar way ensuring that no air bubbles are trapped.
FIG. 11: The gel as it looks just after taking out of the destaining process
• This setup of the gel trapped between pairs of soft tranparent plastic sheets is gently put on a scanner and the
image is taken at a resolution of 600dpi.
• From the image the relevant part is cut out (generally a thin rectangular strip corresponding to the dilution
series) and rotated if necessary.
FIG. 12: The strip of the scanned image of the gel along the dilution series