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I have taken microscope pictures for proteins in HeLa cells, in order to quantify the intensity (immunofluorescence). The proteins are both at the nuclear envelope, and in order to quantify the intensity I draw a line that goes across the whole cell,through the nucleus, and measure the intensity on that line (ImageJ). Which value should I use as background?

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  • $\begingroup$ The area outside of the cell typically. Adjust contrast until the background black $\endgroup$ – rhill45 Feb 10 '15 at 13:57
  • $\begingroup$ and what about the cytoplasmic zone? (both proteins are nuclear) also, should I take an average of certain number of values? $\endgroup$ – Katz Feb 10 '15 at 15:28
  • $\begingroup$ It sounds to be in your best interest to set your analysis boundary around the cell but can you post a sample image so we can see your contrast issues $\endgroup$ – rhill45 Feb 10 '15 at 18:20
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    $\begingroup$ sorry but cannot upload any images. I consulted several people and I have been told to use outside cell as background as well. Thanks! $\endgroup$ – Katz Feb 15 '15 at 14:42

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