Have you found any related forum topics? If so, cross-link them.The first is to treat each row as a horizontal lane and use ImageJs gel analysis function. The quantification will reflect the relative. There are two built in methods for analyzing a dot blot in ImageJ. Use the wand and get a quantification value from the intensity of my band. The western blot image was exported to ImageJ software to measure band density. From the generated plots I mark the bell shaped curve with the line tool. Knockdown was verified with ScanLater western blot analysis. To quantify my western blot bands: I need to place a box around each band. But since it is difficult to place the line on the bell-shaped curve exactly at the same spot it’s hard to tell if it overlapping boxes affect the quantified band intensities. This protocol will allow you to relatively (no absolute values) quantify protein bands from western blot films. Hello, I have a question regarding western blot analysis using imageJ. Images acquired on LI-COR instruments should be analyzed using. I quantified the bands separately and with overlapping boxes. After you have dragged the last rectangle over the last lane, indicate that you are finished by selecting: Analyze > Gels > Plot lanes (or command -3). Use software designed for Western blot analysis that is compatible with your detection system. Keep in mind, though, that the quantitative intensity data captured in your original image HAS changed. I don’t know if the overlapping of boxes of two bands affects the quantified values of each band Would this affect quantification considering that the white space around the two bands is overlapping but the overlapping boxes don’t reach over the actual band? Plots of CHEMI_10242021_154542_fis_br_se_cropped_FIS_marked_bell curves.tif (523.5 KB)Ĭan the boxes from different bands overlap? I need a certain box size to fit the largest band, but this leads to the box around one band overlapping in the white space of the box of the next band (not over the next band, only the white space). *Use the wand and get a quantification value from the intensity of my band.ĬHEMI_10242021_154542_fis_br_se_cropped_FIS_marked.tif (65.6 KB) From the generated plots I mark the bell shaped curve with the line tool levels were quantified using NIH Image J 1.48 software, and the relative quantity of protein with respect to -tubulin or lamin was calculated.I have a question regarding western blot analysis using imageJ.
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