Sampling Dilutions {|class="wikitable" style="text-align: center; color: black;" |Cell Density |Dilution |PBS ( µL) | Cells( µL) |Trypan Blue

| align="center" style="background:#f0f0f0;"|' | align="center" style="background:#f0f0f0;"|Sampling | align="center" style="background:#f0f0f0;"|' | align="center" style="background:#f0f0f0;"|' | align="center" style="background:#f0f0f0;"|' | align="center" style="background:#f0f0f0;"|' |- | Dilutions||Cell Density ( 10^6/ml)||Dilution||PBS (µL)||Cells (µL)||Trypan Blue(µL) |- | ||1-2||4||100||50||50 |- | ||2- 4.5||8||125||25||50 |- | ||4.5 - 7||12||120||15||45 |- | ||>7||16||137.5||12.5||50 |}

1. Take xµL ofPBS in 96 well plate
2. Add required volume of cell culture. Mix once
3. Bring the plate back to microscope, add trypan blue to the PBS + Cell mixture just before counting the sample.

Trypan Blue is toxic to cells. If left too long with trypan blue, even healthy cells will die.

Calculation of LCD :

LCD = Cell Count/ ( 100* 4) * Dilution

Tips :

1. Mix cells before sampling
2. Take cell sample from top of the liquid
3. Mix trepan blue into the PBS + cel solution slowly and well before loading