Titrating for flow cytometry experiments

Рет қаралды 4,652

2 жыл бұрын

Titration of antibodies is one of the most critical aspects of a flow cytometry experiment. In this video, I give a brief overview of the basics of setting up a titration for a flow cytometry experiment.
My concentration will be different from what you do in the lab- so make sure you have enough volume in your first sample in order to remove the necessary volume for the dilution series AND to add the appropriate volume to your samples. For example, if you are doing a 1 in 2 dilution series and adding 100 uL to each sample, you will need to start with a minimum of 200 uL in your first tube!

Пікірлер: 18

@safimoshkani8495 12 күн бұрын

Great instuctor❤❤❤❤❤

@djhell3054 2 жыл бұрын

Thank you so much for posting this! You're the best.

@ajarieger_flow 2 жыл бұрын

Any other requests, let me know! 👍

@berkansavas5870 2 жыл бұрын

It is crazy that today I was trying to set up my intracellular antibody titration for flow cytometry. Can you please make another video to analyze it too? And talk a little about acqusition on cytometer too?

@xuemeiwang1881 Жыл бұрын

fascinating

2 жыл бұрын

Dear Aja! Amazing explanation, quick question: if I'm titrating multiple antibodies at once (but in separate tubes) with different fluorochromes each, would it be necesary to make compensantion controls?

@ajarieger_flow 2 жыл бұрын

Thanks Davis! Because your antibodies are all in separate tubes, compensation controls are not needed (each sample is a single colour is no need to worry about spectral overlap)

@CY2.2.2 8 ай бұрын

On this note however, during the staining process of Blood where you will add a combination of these titrated mABs you will indeed need to account for spectral overlap and thus do a compensation matrixes@@ajarieger_flow

@paporisharma2936 10 ай бұрын

Sweet

@soudas08 3 ай бұрын

How to do a titration for a primary antibody-unconjugated and secondary biotinilyated antibody?

@ajarieger_flow 3 ай бұрын

Thanks for reaching out! This gets more challenging. I have generally first titrated the primary with a constant secondary concentration. I then pick 2-4 primary antibody concentrations that look the best and titrate the secondary antibody with each of those.

@ajarieger_flow 3 ай бұрын

Be aware that non-specific staining will increase at high secondary antibody concentrations, so it’s important to compare the staining of cells with and without primary antibody present (secondary only control) at each concentration

@soudas08 3 ай бұрын

@@ajarieger_flow thank you Dr.Rieger.