Flow cytometry Tutorial | Flow Cytometry Data Analysis

Flow cytometry Tutorial | Flow Cytometry Data Analysis | Flow cytometry Gating

Рет қаралды 132,926

Күн бұрын

This video lecture explains
1. Principle of flow cytometry
2. Overview of instrumentation of flow cytometry
3. Hydrodynamic focusing
4. Flow cytometric data analysis
5. Flow cytometry gating
6. Forward scatter vs Side scatter plot
7. Forward scatter height vs forward scatter area plot for doublet exclusion
8. Single parameter histograms to identify cells with a particular marker expression
9. Two parameter density plots for further analysis
10. Sequential gating to identify specific T subsets
11. Backgating
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Пікірлер: 71

@shiah4205 3 ай бұрын

Thank you very much .I’ve been looking for interpretation for about 4 hours and finally found your video.Can’t thank you enough

@BiologyLectures 2 ай бұрын

You are welcome.

@brandontran9211 2 жыл бұрын

Perfect video! Was struggling in undergrad, but now I understand, especially data analysis section, keep it up and thank you :)

@BiologyLectures 2 жыл бұрын

We are glad that this video was helpful to you. Please subscribe our channel and like ans share the video to support us. Thank you

@polartechie 2 жыл бұрын

Omg that FSX vs SSC debris explaination helped a ton too!

@BiologyLectures 2 жыл бұрын

We are very glad that this video was helpful to you:-)

@12823matthewkao Жыл бұрын

Data analysis section straight to the point!

@BiologyLectures Жыл бұрын

Thank you.

@ivyvien3364 2 жыл бұрын

Very clearly explained! Awesome video :D

@BiologyLectures 2 жыл бұрын

Thank you very much. Please consider subscribing us.

@polartechie 2 жыл бұрын

Fascinating, thanks so much!

@BiologyLectures 2 жыл бұрын

You are most welcome. Please consider subscribing us to support.

@hildegunnink2561 2 жыл бұрын

Hi! Great video, really helped a lot, thank you! I just had two questions if you don't mind my asking (or do you prefer through the fb page rather than the youtube comment section): 1. what exactly is an isotope control, that you mentioned at 12:30? 2. I loved your explanation of the plot analysis, but I don't quite understand the lines graphed in figure 3c (minute 18:53), would you mind enlightening me a bit more on how they are drawn and what they mean? Is it a gating mechanism? thank you so much in advance!

@BiologyLectures 2 жыл бұрын

Dear Hilde Gunnink, Thank you very much for your feedback. We would be very happy, if you suubscribe our channel to support us. Answers to your questions: 1. Isotype controls are primary antibodies that lack specificity to the target, but match the class and type of the primary antibody used in the application. Isotype controls are used as negative controls to help differentiate non-specific background signal from specific antibody signal 2. These lines are showing four quadrants to separate the four different subgroup of cell populations. Hope this helped.

@xozislAoshsnala 2 ай бұрын

Feeling smarter now. Thank you!!

@BiologyLectures 2 ай бұрын

You are most welcome.

@dikshyapanthi7681 3 жыл бұрын

Excellent explanation 😊❤️

@BiologyLectures 3 жыл бұрын

Thank you.

@emmanuelluwaya7974 Жыл бұрын

Excellent explanation. I like it a lot

@BiologyLectures Жыл бұрын

Glad you liked it!

@lee-annspencer1385 3 жыл бұрын

Thank you very much 🙏🏾

@BiologyLectures 3 жыл бұрын

You are most welcome. Please consider subscribing us.

@dakshinapanthi722 3 жыл бұрын

Excellent explanation 😊

@BiologyLectures 3 жыл бұрын

Thank you.

@Hari788 3 жыл бұрын

Excellent explanation sir.

@BiologyLectures 3 жыл бұрын

Thank you.

@prmei8011 25 күн бұрын

great video! Could you explain more about granularity and complexity?

@naranarubi 2 жыл бұрын

Thank you, helped a lot

@BiologyLectures 2 жыл бұрын

You are most welcome:-)

@mEi592 2 жыл бұрын

How would you know if the qc data passed or failed?

@royzhang5192 2 жыл бұрын

it's useful, thanks so much

@BiologyLectures 2 жыл бұрын

You're welcome!

@Theolove111 2 ай бұрын

Thank you so much!!

@BiologyLectures 2 ай бұрын

You are welcome.

@kurapikach4624 2 жыл бұрын

Found this video to be very helpful but I was wondering what cells are represented by the Q4 dots in the far right graph at 15:27

@BiologyLectures 2 жыл бұрын

These are double negative cells i.e. CD3 and CD19 double negative cells.

@swarnadipghosh1735 5 ай бұрын

Very informative video, thanks!

@BiologyLectures 5 ай бұрын

Thanks a lot.

@ISTARI22 Жыл бұрын

Thank you for the video but all these gating process are they hypothesis or its for sure that those cell types are as you mentioned Monocytogenes, granulocytes etc. How do I double confirm these cell population subset?

@BiologyLectures 11 ай бұрын

These are the cells as mentioned in the video. However, if you want to double confirm you can perform staining for the specific marker of these cells and check under microscope 🔬

@simplynano18 Жыл бұрын

Thank you!!

@BiologyLectures Жыл бұрын

You are most welcome

@beyzaozdemir394 Жыл бұрын

Thank you bro!

@BiologyLectures Жыл бұрын

You are most welcome.

@motlalepulemosala7334 9 ай бұрын

Good morning. I'm busy with my research project on flow cytometry. I'd like to know what the cut-off ranges are or what percentage range is bright and dim. I need to do a statistical analysis, but I need a cut-off range for the bright and dim results. Kind regards

@BiologyLectures 9 ай бұрын

Good morning! In flow cytometry, determining bright and dim populations often depends on the specific markers or fluorochromes used in your experiment, as well as the instrument's sensitivity and settings. There isn't a universal percentage range that defines bright and dim cells because it can vary based on several factors. However, I can provide you with some general guidance on how to set your cutoff ranges: Use Positive and Negative Controls: Utilize positive and negative controls in your experiment. Positive controls should be strongly expressing the marker of interest, while negative controls should be cells that do not express the marker. These controls can help you establish appropriate gates for bright and dim populations. Consider Histograms: Analyze histograms of your samples. Histograms can give you an idea about the distribution of fluorescence intensity. You can set gates based on these histograms to separate bright and dim populations. Use Fluorescence Minus One (FMO) Controls: FMO controls are helpful for setting gates in multicolor flow cytometry experiments. FMOs contain all fluorochromes except the one you're interested in. By using FMO controls, you can establish gates that differentiate specific fluorescence from background or nonspecific staining.

@motlalepulemosala7334 9 ай бұрын

My research is about the CD123 in leukaemia. I did literature review on the flow cytometry part of it and I'm not getting any solution. The statistician asked me to give him the percentage range for bright and dim results, so that he can do the data analysis for me. I didn't run the flow cytometry, I only obtained the patient results from the system. Based on your response, I'm guessing I'll have to go to the lab and find someone who runs them to check all that you've suggested? My exams are starting in November, and I need to complete my research write-up before then. 👌🌸

@quangvinhnguyen114 Жыл бұрын

Many thanks for great explanation but Can I have a question? What does the colors of the dots indicate (as in 19:00). Is it the density? Like the number of cells in red area will higher than yellow one, and yellow > green > blue right? sorry for my poor english

@BiologyLectures 11 ай бұрын

Yes, that is correct

@quangvinhnguyen114 11 ай бұрын

@@BiologyLectures thank you!

@BiologyLectures 11 ай бұрын

@@quangvinhnguyen114 You are most welcome

@markwang9259 3 жыл бұрын

Sorry, could you explain what is doublets? Thanks

@BiologyLectures 3 жыл бұрын

Occasionally we would see two cells stuck together referred to as as a "doublet."

@AmruMagdy Жыл бұрын

thank you :

@BiologyLectures Жыл бұрын

You are most welcome

@khalilelaj4840 2 жыл бұрын

Superrrrrrr

@BiologyLectures 2 жыл бұрын

Thank you so much.

@Verstehdieaktie Жыл бұрын

Thanks

@BiologyLectures Жыл бұрын

You are welcome

@sarahfaress5957 2 жыл бұрын

In a density plot, what the do the red, green, blue color signify?

@BiologyLectures 2 жыл бұрын

Thank you for raising this question. In density plot, the red color represents the highest density of events within the cell population. With decreasing density, the color transitions from yellow over green to blue

@sarahfaress5957 2 жыл бұрын

@@BiologyLectures Thanks for the reply. So for example at 8:55, in the lymphocyte population, the red would be the highest density of a certain pop of lymphocytes (say B cells), then yellow would be the area with the second highest density of cell, say T cells? I'm just trying to understand how to read it.