Measure rH2AX intensity

Image Analysis
#1

Hi,
I used “speckle counting” pipeline to count rH2X foci. Now I need to measure the intensity of rH2AX staining in the whole nucleus (not only the intensity of rH2AX foci). However, I am confused by the output names in the Excel file. There are 15 outputs which seem related to rH2AX intensity.
Mean_H2AX_Intensity_IntegratedIntensityEdge_rH2AX
Mean_H2AX_Intensity_IntegratedIntensity_rH2AX
Mean_H2AX_Intensity_LowerQuartileIntensity_rH2AX
Mean_H2AX_Intensity_MassDisplacement_rH2AX
Mean_H2AX_Intensity_MaxIntensityEdge_rH2AX
Mean_H2AX_Intensity_MaxIntensity_rH2AX
Mean_H2AX_Intensity_MeanIntensityEdge_rH2AX
Mean_H2AX_Intensity_MeanIntensity_rH2AX
Mean_H2AX_Intensity_MedianIntensity_rH2AX
Mean_H2AX_Intensity_MedianIntensity_rH2AX
Mean_H2AX_Intensity_MinIntensityEdge_rH2AX
Mean_H2AX_Intensity_MinIntensity_rH2AX
Mean_H2AX_Intensity_StdIntensityEdge_rH2AX
Mean_H2AX_Intensity_StdIntensity_rH2AX
Mean_H2AX_Intensity_UpperQuartileIntensity_rH2AX

Could you help me figure out which one I should use?

Thanks a lot,
Rong

#2

Hi Rong,

These mean measurements are generated by the RelateObjects module which performs a per-nucleus mean calculation for all the child objects (i.e, rH2X). So ‘Mean_H2AX_Intensity_IntegratedIntensity_rH2AX’ means 'the mean per-nucleus integrated intensity for all H2AX objects measured from the rH2AX image." Since MeasureObjectIntensity makes 15 measurements (see the help for the module), the RelateObjects module creates means from those 15 features.

Whichever measurement you choose depends on what phenotype you’re interested in. If you want the total per-nucleus rH2AX intensity, then choose IntegratedIntensity; if you want the mean per-nucleus intensity, then choose MeanIntesnity, so on.

Regards,
-Mark

#3

So to ensure I understand it correctly:
Mean_H2AX_Intensity_IntegratedIntensity_rH2AX’ means in this case mean total H2AX intensity in all nuclei
and
Mean_H2AX_Intensity_MeanIntensity_rH2AX’ means mean intensity of one H2AX foci in all nuclei
Am I correct?

Thank you

#4

To clarify:

For each nucleus, the mean of the total (i.e., integrated) rH2AX intensity of all H2AX foci objects contained in that nucleus.

For each nucleus, the mean of the mean rH2AX intensity of all H2AX foci objects contained in that nucleus.
-Mark