1 To perform a cell count with the hemacytometer you need at

1) To perform a cell count with the hemacytometer, you need at least 20 L of solution, typically 10 L of cell suspension and 10 L of Trypan Blue. This is referred to as a 1:2 dilution. However, when the cell density is too high to count efficiently, the solution can be further diluted with PBS. Adding to the 20 L of cell count solution at your bench, make a 1:3 and 1:10 dilution with PBS.

2) Prepare 25 mL of destain solution: 20% methanol, 10% acetic acid; 70% water.

Solution

(1)You said you want 1:2 dilution of cell suspension with Trypan Blue.

Then take 10 L of cell suspension and add 10 L of Trypan blue to it, to get 1:2 dilution.

10 L of cell suspension: 10 L of Trypan blue (1:2 dilution)

Now you want to dilute your cell suspension with PBS to get 1:3 and 1:10 dilution

For 1:3 dilution

10L of cell suspension+ 20 L of PBS

For 1:10 dilution

10 L of cell suspension+ 90 L of PBS

(2)We need to add 20 ml of methanol, 10ml of acetic acid and 70 ml of water to get 100 ml of destain solution with 20% methanol, 10% acetic acid; 70% water.

But you need only 25 ml of destain solution

Volume of methanol needed for 25 ml of destain solution=25x20/100=5 ml

Volume of acetic acid needed for 25 ml of destain solution=25x10/100=2.5 ml

Volume of water needed for 25 ml of destain solution=25x70/100=17.5 ml

Now 5ml of methanol+ 2.5 ml of acetic acid+ 17.5 ml of acetic acid will give you 25 ml of destain solution with 20% methanol, 10% acetic acid; 70% water