Dear Mohammad Asad,

Usually, you can tell your cells are smaller or larger comparing FSC-A vs SSC-A. So, if you results are closer to 0 on each axis, then your cells are smaller. But it's only comparative size. Otherwise, you will need size beads or cells.

I think you are in need to refrence beads,

otherwise you will have rough estimation to size , and it all will depend on which cells you study

The position of your cells on the FSC and SSC plot is completely dependent upon the amplifier and gain settings during that experiment. These settings will directly impact the position and scale of the axis.  You can obtain relative size differences between cells, but even that is limited to larger or smaller because the scale is not fixed and changes with the amplifier and gain settings. FSC-W generally gives a good indication of size, but SSC-A also works.

To obtain any sort of quantitative measurement of size you will need to run reference size beads using the same settings and flow rate, preferably during the same experiment.  One complicating factor is that the beads will be near spherical and your cells may not be.

Dear All!

Thank you very much for your responses. Actually in few papers they were measuring the exact cell size (without using reference beads). Thus, I wanted to know the rationale behind.

Best regards,

Asad

Hi Mohammad! I also agree that you should use size reference beads. Can you please link the papers you are talking about?

PS: does anyone have a good protocol for cell size measurement with beads?

Best regards,

Gabriele