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# Counting beads binding to RBCs

### #1 biology_06er

biology_06er

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Posted 24 February 2016 - 03:13 PM

Hi guys,,

Cheers,

biology_06er

### #2 biology_06er

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Posted 24 February 2016 - 04:50 PM

Another question, to check if my maths is correct!! The calculation people use for absolute count is: (#cells counted in population targeted/beads counted)*bead concentration

I have been using the calculation originally used by my supervisor. Example: I make up a master mix of buffer/beads and resuspend my sample in 500uL of this buffer. The final concentration of the beads in this master mix is now 10.21beads/uL (stock vial is 10.21/uL and I take 160uL of this, add to 16mL buffer). I have collected 5uL of blood and into 500uL buffer so a 100x dilution.

Then I collect say 1million events in targeted population and my beads in my gated sample are 500. So the volume of beads I have sucked up is 500/10.21=48.97uL. Therefore, 1mill events/48.98*100 DF* 1000 = cells per mL

Hmm, so I think I might have understood my first question...if my bead concentration from the vial is 1000beads/uL. And in my population I am saying I am collecting 500beads in my gate but my total bead number is 1000 (including the ones bound to RBCs) and the number of RBCs I have collected is 1,000,000. Therefore the volume of beads I am collecting is 500/1000=