Quantification of intranuclear label – immunocytochemistry

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Quantification of intranuclear label – immunocytochemistry

bcha7650
I have performed immunocytochemistry on an antigen which is both cytoplasmic
and intranuclear and would like to quantify the intensity density of
intranuclear label. I have labelled the nuclei with DAPI.

The workflow I have attempted was to first split the colours, threshold the
DAPI with outlines showing the ROIs then attempt to create a mask. However,
I am not sure how to overlay the mask in the other channel so that I can
measure the intensity density of the red channel using the masks I created
in the blue channel.

Have I got the workflow correct, or is there a better way to do this?

Any help is greatly appreciated

B







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Re: Quantification of intranuclear label – immunocytochemistry

Herbie
Good day,

I'm not a specialist in your field but I think you are not far from a
good solution.

If you have the selections in the blue channel you are to enter all of
them (one after the other) to the ROI-Manager. Please first study the
relevant section "30.14.5 ROI Manager…" of the ImageJ User Guide:
<https://imagej.nih.gov/ij/docs/guide/146-30.html#toc-Subsection-30.14>
(scroll down the page to subsection 30.14.5)

With the selections in the ROI-Manager it is easy to transfer them to
the red channel and do the measurements there. For the latter, "More >>
Multi Measure" may be handy...

Regards

Herbie

::::::::::::::::::::::::::::::::::::::
Am 13.07.20 um 06:24 schrieb bcha7650:

> I have performed immunocytochemistry on an antigen which is both cytoplasmic
> and intranuclear and would like to quantify the intensity density of
> intranuclear label. I have labelled the nuclei with DAPI.
>
> The workflow I have attempted was to first split the colours, threshold the
> DAPI with outlines showing the ROIs then attempt to create a mask. However,
> I am not sure how to overlay the mask in the other channel so that I can
> measure the intensity density of the red channel using the masks I created
> in the blue channel.
>
> Have I got the workflow correct, or is there a better way to do this?
>
> Any help is greatly appreciated
>
> B

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