Quantification of western blot using imageJ for beginners | western blot quantification | imagej |

This video lecture describes how to quantify western blot using imagej.
1. How to upload western blot image in imagej
2. How to invert image in imagej
3. How to adjust background/ brightness and contrast of western blot image in imagej
4. How to rotate image in imagej to make the lanes in the straight line
5. How to put rectangles in the image to quantify
6. How to plot lanes in imagej
7. How to make the calculation for western blot after the quantification by calculating ratios

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[howtoneuro]

I wish I had this explanation a long time ago! Thank you for sharing this knowledge. Doing experiments is cool, but analyzing data is always more tricky!

[antarasengupta]

Thank you! It was very clear and systematic-easy to understand and follow.

[SM-xnbv]

Excellent job!!! Very clear and consize! Many thanks!

[kobbyblaqboi]

Final ratio was rather Protein r/ actin r I think. Good video! thank you.

[AnnieNaeem]

Thank you so much for making it easy to understand.

[markrobinson]

thank you for saving my life. awesome.

[dikshyapanthi]

Excellent explanation sir . Thank you very much for your video

[ItsDeeable]

Sorry, just need a clarification - do you divide Actin Ratio by Protein Ratio or vice versa? You wrote down Actin R/Protein R but do otherwise. Thanks!

[Hari]

Excellent explanation sir. This is what I needed

[jagdeepsingh]

Thank you for such a beautifully explained video.

[mrblackmamba]

Life saver. Thank you so much sir. Subscribed.

[livinghope]

Thanx for responding my question!
I subscrbed, liked + shared your video

[job]

Many thanks, once again. Please, how do I obtain the relative intensity (Protein expression) for samples if the control does not have any protein expression?

[aynalsaba]

Thank you so much for sharing your knowledge with us. I really appreciate your work.
Sir, i have one query. You have taken 1 experimental control for explaining the analysis process. Sir, i have done western blot on clinical samples and i have taken 3 control samples and 3 patient samples for my western blot process. So the thing is how will i take the ratio for those 3 controls?

[mvbs]

Thank you for the clear explanation!
I had one question, what if my CTRL value is 0, how do I now calculate the ratio's and perform the normalisation?

[jackbilotto]

Hi thanks for the video. If my control protein and protein of interest are in seperate images will the contrast settings affect the result?

[roxanne]

At 4:22 you said there was no need for you to adjust the lanes. What I get are lanes that "don't touch the zero" so two bands have the same curve. How do I separate them?

[vanesamattera]

Thank you for the video, how do you tag the Y axis in the plot?. Is it ok to label it like: Protein/actin (relative to control)?.

[job]

Many thanks! I observed that some faint bands were entirely eliminated due to the adjustments of brightness/contract; how can you account for the bands. What are they, please?

[raquelhanadulset]

I did a western blot with n=4 for both control and treatment to observe the expression of a protein of interest and then normalise against housekeeping gene .. how do I plot the data in this case? Do I calculate the ratios for each n individually? how do I carry out a paired student t-test with ratios o does it need to be with the means of the area under the curve calculated with image j? I would appreciate your help. thanks a lot!!