Bio-Rad Image Lab™ Software - Manual (Page 143)

Normalizing Volume Data
User Guide | 143
 Delete — deletes a band from a lane. Use this feature to remove bands
that are not relevant to your analysis. Click Delete, and then click on the
band you want to remove.
 Adjust — adjusts the height of a band. Click Adjust. Boundary lines appear
on either side of each band. Move the cursor over a boundary line until you
see a double-headed arrow. Move the boundary line up or down. The
center is recalculated and the band appears there.
Note: You can also adjust band boundaries in the Lane Profile view.
Normalizing Volume Data
There are several reasons why you might want to normalize the volume data in
multichannel images:
 The lanes are loaded with the same volume, but you do not know the total
protein in each lane.
 Pipetting errors result in variations in the lane volumes.
 Differences occur in the transfer of protein from a gel to a membrane.
You can correct for these differences by normalizing the volume data. There are two
ways to normalize volume data:
 Total Lane Protein Normalization – one lane in the normalization channel
is used to calculate the normalization factor.
 Housekeeping Protein Bands Normalization – a single band of a
housekeeping protein is used to calculate the normalization factor. In order
to get accurate results, the housekeeping protein must be stable and
impervious to the treatments in your experiment. It must be the same in
pretreatment and post-treatment.
In both cases, the normalization factor is then used to calculate the normalized
volumes in the lanes for all channels.