ImageJ 101 For Every PhD with Image Data!
ฝัง
- เผยแพร่เมื่อ 27 พ.ค. 2024
- Things about ImageJ that I wish someone told me about (Macro and Batch Processing Features) #imagej (long title)
A picture is worth a thousand words, and when you acquire an image for your research, be sure to consider every pixel as a potential source of data!
My favorite software is certainly ImageJ because it is free, open-source, and has many powerful applications that convert scientific images to publishable data and beautiful figures.
To celebrate my long friendship with ImageJ, the best buddy during Ph.D. and postdoc studies, I made this video and hope this will pay it forward to the next generation of scientists.
In this video, let's take look at the common applications of ImageJ:
🧫Counting Cell
📐Length and area measurement
🌡Gel Band intensity
🔬Fluorescence quantification
🗿3D volume calculation
🦠Cell migration speed
Recently, I have also discovered the tool of automating Image Analysis and really wished I had the knowledge sooner!
Timestamps:
Start 0:00
History of Image J and common application 1:34
Basics of Length and Area analysis 4:10
Basics of Intensity Measurement 7:45
3D volume calculation 9:00
ImageJ FIJI macro scripts and Batch processing script 10:21
ImageJ Download:
imagej.net/Fiji/Downloads
Tutorials and examples:
imagej.nih.gov/ij/docs/exampl...
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About me:
I am a biologist finishing the 6th-year postdoc. I grew up in Hong Kong, have worked in the US and now living in France, now searching for my next role in private sectors of science.
Recruiters, please contact me for working opportunities: / verabschan
Before I conclude my time as a postdoc, I really hope to make TH-cam videos that are tailored for the younger science community. These are basically what I would share if you find me as a postdoc in your lab. I hope my personal tips will provide some positivity, support, and encouragement to those who are on the journey of academic research.
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Vlog videos:
How I arrived in France + Long-Stay Visa tips + Euraxess
• Long-Stay Visa Tips Wh...
How I handle the unpredictable future (and work productively from home)
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Hello, glad i will be defending my PhD thesis tomorrow and just stumbled on this app and video. Its very educating. Thank you
Great introduction to ImageJ! I'm currently using it for my PhD, and your video has been incredibly helpful in simplifying some of the things I need to figure out. Thank you so much
Fantastic intro to ImageJ; many many thanks! This is super helpful! :)
Hi Cesar, I wishes someone would have taught me as I started using ImageJ! Please share with anyone who might need this!
I love your little story about measuring a friend with the software :')
haha, hope you don't think I am creepy!
@@PhDCoffeeTime No, I could do the same !
Thank you for a great intro!
I guess I have found my new fav channel! Tysm!!!
Thank you so much. This comment has made my day!
Excellent video! Thank you so much!
Glad it is help to you. Please share it with anyone who needs this ImageJ advice!
Hi! I use imageJ to mesure leaf. I really liked your video, you made the use of ImageJ look very easy. Thanks for shearing!
What a coincidence that I was using leave as example :)
I hope the Macro script saves you time. If you have any questions about certain imageJ features, please feel free to ask here :)
@@PhDCoffeeTime Thanks a lot, I will do if I need it! :-)
Great introduction, thank you.
Glad you enjoyed it!
Awesoome!Thank you! You inspired me to, DEFINETLY!
It's a pleasure!
Thanks! I learned a lot.
Hello Dr Chan, Thank you so much for this video.
I have some Images I need to quantify the protein expression but I have issues as I am seeing online that using the intensities is not a good way to do the quantification. Please, can you kindly give me some guidelines?
Hi Dr Chan, thanks for great video! I really interested in that pixel to calculate the volume, is it possible to use imagej to calculate the height of the crop ? How could I valid that results ? Thanks for the help!
Hi Dr. Chan! PhD Coffee Time is great, and I love the work you're doing. I was wondering if you had any suggestions for using the BoneJ plugin to quantify osteons and osteon shape variation in cortical bone. Thanks for any input you can provide!
Hi Jessica, I have not used BoneJ myself, but it looks like this is a relevant paper for you: www.ncbi.nlm.nih.gov/pmc/articles/PMC3193171/
@@PhDCoffeeTime Thanks for your response! I hope you are well. :)
Hi Dr. Chan. I love the simplicity of the video. I am learning about western blot image analysis using ImageJ and this video has helped me understand the basics of image analysis. Would you recommend any textbook or course for beginners that I can take online to advance my knowledge in image analysis? Thank you.
Hi David, for western blot you may find this tutorial on dot-blot relevant: imagej.nih.gov/ij/docs/examples/dot-blot/index.html
When I learned macro languge on ImageJ batch processing, I have really enjoyed reading this book by Kota Miura: biii.eu/kota-miura-ed-bioimage-data-analysis-textbook-wiley
Thank you so much. Can I
use skeletonize 2D/3D for CAM assay to measure vascular density?
Thank you, Dr. Chan. I would like request to give me guideline how to calculate the percent (%) of shadow of a photo.
Excellent explanation... thank you
This would be helpful! Thanks!
Hi Bosong, thank you! Please help to share the video links to anyone who might find this helpful in their research. Also, feel free to leave me any question about ImageJ, you may be featured in the next video :)
Hi Dr Chan, Thanks for great video. I need to caculate area of eyes skin after levato resection, and I've been looking for video teaching that stuff but I cant find anything. Would you mind showing me the way to do that or sending me any video clip or documents so I can learn along that? Thanks for your helps!
Hi guys, please feel free to come back to this video as you need the technical demos.
Timestamps:
History of Image J and common application @1:34
Basics of Length and Area analysis @4:10
Basics of Intensity Measurement @7:45
3D volume calculation @9:00
ImageJ FIJI macro scripts and Batch processing script @10:21
Hi Madam
Please I would like to know how to do automation in height measurements in a Biphasic chemical reactor
Crucial video for many applications! Used to use it for western blot band intensity...Next image analysis I did was in my biotech job, using Incucyte, but that has it's own algorythm and software. I'm thinking to use it for histology, but is it appropriate for H&E stains? For IHC with fluorescence I can imagine it's super! (Like the nucleus counts).
By the way: hahahahaha on the tshirt :D
Yes, you could use RGB separation for the image and threshold for the nuclei (rich in BLUE colour).
Then you threshold the nuclei from the Blue channel which has mostly nuclei channel.
Partical Analysis will be the next step and you will have automated nuclei count from the Blue image.
Feel free to send me an image and I could take look at it!
It was actually a nice button up shirt, so it was quite a mission! I used the sleeve length of a T-shirt that we both own as a "ruler".
It was priceless moment when I showed him all the photos I kept for the ImageJ project, I said "I have measured you on ImageJ from the sample number of X when you were wearing this same shirt from different angle, and I decided to order this size." And it fits him perfectly...
@@PhDCoffeeTime You're such a good friend! :D
Awsome tips keep up
Thanks! I love imageJ :)
Hi. Thanks for your video. I need support regarding reconstruction of 3D image from micro CT. Do you have?
Nice one
Please consider sharing with any students who might need this video :)
Thank you for this, this was so helpful! I am an undergraduate working on research where I need to measure leaves, and this is exactly what I was looking for :-). I was wondering, do you know how to add interactivity in the macro script? I have many leaves I need to measure, but the scale to measure is different for each one, such that I need to remeasure the line each time. Do you know how to edit the macro script so it will stop and ask me to make the line, and then it continues with the rest of steps? Thank you!
Hi Vera S. thank you for the question. Yes, you can add the pixel interactively to the macro. Let me begin by saying that the process is still mostly manual as the scale bars are not the same. However, the macro script can help you save a few clicks.
You will still need to close and open each picture manually and use a "line" to measure the scale bar ("m" on keyboard) in pixel.
Then you could enter (or copy/paste) the length of the scale bar in pixel and run the new macro script. That will save you time from going to Analyze/Set scale every time!
Here is an example in case I wasn't clear, if the first image (Image A) has a 10mm scale bar and pixel length of 903.6824, the macro could look like this:
run("Set Scale...", "distance=903.6924 known=10 unit=mm");
Now when you open the second image (Image B) the scale could be at a different magnification, so I would do the following to minimize manual clicking:
(1) close Image A (so that it clears the scale set for the image
(2) open Image B (have everything in the same folder, so just change the name on the macro script to open, that will save you a few clicks)
(3) run a line along the new scale
(4) hit M to measure, copy the length in pixel, e.g. 800.2341
(5) in the macro, change the "distance = 903.6824" into "distance = 800.2341" a new pixel value.
(6) draw a line to verify if you have the right scale bar by using "line" and "measure".
(7) After calibration, the length of the scale bar should be "10 mm"
Let me know if this is helpful for your situation.
Now only I started using ImageJ. Can you tell me about the segmentation and reconstruction of the image One more thing I want to ask is if imagej has any function to auto-connect the vessels like the auto-fill function?
I came to this video just after measuring my gf in a picture with imageJ to buy her a shirt of the right length hahaha, I'm glad to know that I'm not the only one.
I liked the program so now I want to learn a bit more about it :)
Oh wow, I am pleased to know I am not the only one!
Yes imageJ is really practical and I hope this short video is a good summary about its main features. Feel free to suggest any video request here :)
Hi, I have a question and hope to get my answer from you.
Actually I have some pictures (with green florescent dots as cells) randomly taken from my sample. I counted the cells/picture by ImageJ and I was wondering how to extrapolate these certain number of cells counted as it's very hard to take lots of pictures from my samples.
Hi Mansoureh, There is a plugin called "Particle Analysis"
imagej.net/Particle_Analysis
You could check by optimizing the image threshold to represent accurate count of cells in each picture (in 1-2 image), and compare the first dataset with manual counting.
@@PhDCoffeeTime thanks for your quick response. Actually, I know how to count cells with the "analyze particle" plugin. My main issue is how to extrapolate the data produced from each sample. Imagine that I have 5 images taken from different random spots of a sample. I want to know the number of cells supposed to be counted for about 30 images without taking them by a microscope. The only way is using Excel to extrapolate data but my data has diversity so cannot be fitted a line to extrapolate it. I was wondering if there is a specific option in ImageJ? thanks for your support.
Thank you for sharing about your research, it looks like you have a data visualization problem, remember imageJ is mostly for image analysis but there are options to export data for further data indexing and visualization with python.
Here are more details about imageJ data exporting.
imagej.net/Saving_and_Exporting
However, programming will take time too, the balance would be to estimate how long it would take to use the manual method, and perhaps the few hours of programming is worthwhile if you intend to use this approach repetitively for a large project.
If you know someone with a programming background, try taking your exported CSV files and consult a computer scientist, this could be a simple data analysis problem that could be figured out quickly.
Let me know how it goes!
Hi, I have a query how to calculate probably paramter for irregularities (PPi)
I have no idea, do you have examples?
Vera, do you use activity diagrams in your work?
Hi Saadat, I mostly use Gantt chart because it shows time component of a project. Do you use activity diagram in your work?
@@PhDCoffeeTime Yeah. I use it for the conceptualisation of my Master Thesis. I used WhiteStar UML, but the output is so ugly, so I had to redo it in a graphic editor (thanks for the Inkscape suggestion) to have a nice figure.
By the way, which Gantt chart do you use? :)
Hello! Could anyone tell me why my Image J Folder is missing the ij.jar component in the folder? Since it is missing I can't run the program. I had to download the application for one of my under-graduate classes but I'm really unfamiliar with the program so any information would be appreciated!
Hi Pamela, it's not straight forward to resolve that without seeing your computer. I would try to uninstall from Program completely then reinstall ImageJ (I receommend ImageJ Fiji btw)
For more specific troubleshooting, I hope this page will have answers to your problem!
imagej.net/Frequently_Asked_Questions
Please let me know if any of these work!
This video is Good epitizer for imagej
I think its time to explore imagej in details and we looking forward to hearing from you soon
Any video suggestions on ImageJ? What are you analyzing these days? I can see how I can create a mini-tutorial on a broadly interesting topic.
@@PhDCoffeeTime pictures editing, particles size measurements .
It will be glade if you make a mini tutorial on plant cell measurements using imageJ
@@Ibrahim920 If you have plant cell images (first make sure your supervisor approve!) that I can use for the mini tutorial, please send to phdcoffeetime@gmail.com!
I will create that video with the parameters you have suggested and acknowledge your contribution.
@@PhDCoffeeTime ok let me call for approval to supervisor
@@Ibrahim920 Thanks for considering this idea!
Hallo, my name is Syefira and i'm a collage student. Here, I would like to ask about imageJ/fiji plugin BoneJ due to the final project i'm working on. Can imageJ/fiji plugin boneJ be used to find radiopharmaceutical uptake levels in bone? And using BoneJ to find the count value of the ROI technique from bone scan?
Thank you very much, I hope i coud get a better information from you.
Have a good day
Hi, Syefira. Thanks for the question. Could you specify what is the image (2D/3D) data you have, for example, are they black and white images with white areas representing radioactive signals (drug uptake?). I would be happy to take a look, you could send me an email via phdcoffeetime@gamil.com
Thank you for this great video. Would you please guide me on how I can plot the histogram for particles volume? I could not find anything by search!
Can you please tell
How to annotate non uniform particles. It will be more than a help
Hi Mohit, I am not sure I follow your question! If you have a more complex analysis to explain with text, don't hesitate to email me (phdcoffeetime@gmail.com). I am happy to take a look at the image.
Hi I want to use image J software to measure infarct size of brain tissue. researchers use scanners to take images, can i use camera or mobile phone to take images. pls suggest
Are these tissues stained? or have a different shade of grey? You can measure the area by thresholding (select the relevant RGB layer if stained in color). Let me know if this helps!
Hi, I am in trouble with the measurement of my plant that is in pots next to each other. My aim is to the measurement of the growth of plants one by one. I tried to find you on LinkedIn but can't open your profile and send you pictures to show clearly what I mean :( I would really appreciate it if you can help me somehow?
Hi Asl V, you can send me an email (phdcoffeetime@gmail.com), I will look into it when I have free time. Hope I have a good answer for you!
Does anyone know how to export photo that are annotated on imagej ? I’m trying to circle label mitochondria. But when I export them the circles are not there.
Hi Kelssey, I personally find the annotation is easier to be done on Inkscape, another free software (but completely worth the download!)
I keep ImageJ for adding scale bars, and find it much more efficient labeling and annotating on Inkscape environment:
Hope this video helps you: th-cam.com/video/9MX2lap_ot0/w-d-xo.html
Hi, I'm working on a project with image j and would like to ask u of how to proceed further, it would be great if u help me a bit with it.
Thanks🙂
Hi Omkar, I can look into the problems, could you send me your questions through Instagram/ Facebook/ Twitter? If you want to stay private, you could use the direct message function
@@PhDCoffeeTime 😃, Thankyou very much for the reply it means a lot. Yeah, what's ur instagram profile name?
@@omkarpawar7958 It is "phdcoffeetime" look forward to hearing from you.
@@PhDCoffeeTime Hey, i have msged u on instagram 🙂👍.
I use imageJ to measure sizes of microplastics and boy is it a pain. Thanks for sharing some tips.
Hey I did the same with oyster larvae, how many images you have? Do you use "Particle Analysis Tool"? Or just measured by hand?
Hello Vera!
Hi mam I'm an postgraduate agricultural student in statistics from India in currently going to do an image analysis using maize(cob) image to predict yield actually I referred an article released by CIMMYT they have used this software to analyse bt they had cob analyser option in plugins folder bt while I open it is not there can u plz help me it'll be very helpful for my thesis plzzz
Hi, I am not sure I understand your question. Could you share the link to the article? And clarify what you are looking for? I will look into that
How can point counting on rock thin section can be done by image J.
Hi Nishu, it depends on your image
(1) If the threshold is obvious, you could use "Particle Analysis"
imagej.nih.gov/ij/docs/menus/analyze.html
(2) In my case, it was a very manual function, I used "multipoint" and tag each barnacle by hitting "t" on the keyboard.
Hope this helps!
@@PhDCoffeeTime Thank you, i will try and let you know if any doubt
Hi Madam
Please I would like to know how to do automation in height measurements in a Biphasic chemical reactor
Hi thanks for the question. I would need more information before I can help. Could you send in a more detailed email to phdcoffeetime@gmail.com?
e.g. What is the type of image data you have? Are there publication related to your question that I can take a look at.
Hello Madam
Please I have pictures of the aggregates coated with the bitumen and I want to know the percentage of the bitumen,I found methods based on the substraction of the background and the color component and by the passage of the calculation of surfaces
can you help me and thanks PhD.
What is the type of the image? If it is coloured, extract the closest RGB channel to bitumen (if it's blue, use the Blue layer), conver the layer to 8-bit (Image > Type > 8-bit) for thresholding (Image > Adjust > Threshold....), set scale then press M to measure (Analyze > Measure)
Thanks for answering me.Please, can you send me your email address, I will explain it to you what I want exactly.
@@benamladjmohamed4397 sure, you can send the question to phdcoffeetime@gmail.com
@@PhDCoffeeTime thanks, i sent you my question to your email address.
Well.. It was a kind of introduction video about Image J but it does not look like an informative or educational video. It is more like an advertisement-type video. Hope you can make some educational and detailed videos about this software which would be helpful. Thanks.
Thanks for your suggestions, well noted for future videos. Are you searching for a specific ImageJ operations that you cannot find a good tutorial about? Please fee free to share more with me.
May I know how yo calculate the diameter of a circle in mm?
Hi Ali,
Thanks for the question. You can do the following after calibrating (set scale with a know scale bar, see the video how to convert pixel to mm)
(1) draw a line across the longest end of the object, hit M to measure diameter in length
or
(2) If you have many images, you may want to use the Analyze>Analyze Particles funtion
Set Measurements (Analyze>Analyze Particles), you can choose "Feret Diameter" (defined as the longest distance between any two points along the selection boundary), so all the diameters of the detected objects will be measured.
Read more about Analyze Particle here: imagej.nih.gov/ij/docs/guide/146-30.html
Let me know if this works well for you!
@@PhDCoffeeTime Many thanks for your valuable feedback, sure I'll follow accordingly 👍👍
@@aliasghar6288 Let me know if it works well, if you have more complex issues, feel free to send me an email via phdcoffeetime@gmail.com (I try to check every weekend in my free time)
why do only 3 colors (RGB) are considered in imagej?
Great question! RGB are the three additive primary colors, red, green, and blue, and they can combine to create any color!
In fact, the RGB values for black and white are rgb(0, 0, 0) and rgb(255, 255, 255) , respectively.
Great summary on Wiki here, hope this helps you understand the concept of RGB.
en.wikipedia.org/wiki/RGB_color_model
:D
it will be really really helpful if u can help me i'll forever be grateful i'm on verge to end my thesis please help mam kindly look into that mam.
Hi Sandhyaprakash, I don't understand where you have sent your questions. Could you write to me via email? It's phdcoffeetime@gmail.com
I will do my best to help.
hi mam i'm trying sending u replies
Hi Sandhyaprakash, I don't understand where you have sent your questions. Could you write to me via email? It's phdcoffeetime@gmail.com
I will do my best to help.