Hello everyone, I am currently trying to do a cck-8 assay, Please I need an assistance, I am having trouble to get the exact volume of cell suspension to seed. I need 5000 cells and I have checked my cell number, and I have 2.31x10^6 cells /ml, 3.09x10^6 cell/ml, 9.65x10^6 cell/ml and 4.88x10^6 cell/ml respectively?
Hello Innocent Agida
You may please follow the calculations provided below.
You need 5000 cells per well for a 96 well plate.
So, consider 5000 cells for 100 wells (Take extra wells to avoid loss of volume in the final well).
You use the formula:
C1V1=C2V2
Where
C1 = the number of cells that you have (say 2.31 x 10^6 cells /ml)
V1 = volume of original cell suspension to be taken (X ml)
C2 = The total number cells required for 100 wells ( 5000 x 100 = 500 000)
V2 = Total volume for 100 wells (say 100ul per well, therefore 100ul x 100 = 10000ul).
Using the formula
2.31 x 10^6 cell/ml x Xml = 500000 x 10 ml
X = 2.164 ml
You would add 2.16 ml of the original cell suspension (2.31x10^6 cells /ml) to 7.84 ml of culture media to get a total of 500000 cells in 10 ml. Mix well. When you add 100ul of this suspension to each well, you will be adding 5000 cells per well.
Similarly, for 3.09 x 10^6 cell/ml
Use the formula
C1V1=C2V2
C1= the number of cells that you have (3.09 x 10^6 cell/ml)
V1= volume of original cell suspension to be taken (Xml)
C2= The total number cells required for 100 wells ( 5000 x 100 = 500 000)
V2 = Total volume for 100 wells (say 100ul per well, therefore 100 x 100 = 10000ul).
Using the formula
3.09 x 10^6 cell/ml x Xml = 500000 x 10 ml
X = 1.618 ml
You would add 1.61ml of the original cell suspension containing (3.09x10^6 cell/ml) to 8.39ml of culture media to get a total of 500000 cells in 10 ml. Mix well. When you add 100ul of this suspension to each well, you will be adding 5000 cells per well.
Similarly, for 9.65x10^6 cell/ml
Use the formula
C1V1=C2V2
C1= the number of cells that you have (9.65x10^6 cell/ml)
V1= volume of original cell suspension to be taken (Xml)
C2= The total number cells required for 100 wells ( 5000 x 100 = 500 000)
V2 = Total volume for 100 wells (say 100ul per well, therefore 100 x 100 = 10000ul).
Using the formula
9.65x10^6 cell/ml x Xml= 500000 x 10 ml
X = 0.518 ml
You would add 518ul of the original cell suspension containing (9.65x10^6 cell/ml) to 9.482 ml of culture media to get a total of 500000 cells in 10 ml. Mix well. When you add 100ul of this suspension to each well, you will be adding 5000 cells per well.
Finally, for 4.88x10^6 cell/ml
Use the formula
C1V1=C2V2
C1= the number of cells that you have (4.88x10^6 cell/ml)
V1= volume of original cell suspension to be taken (Xml)
C2= The total number cells required for 100 wells ( 5000 x 100 = 500 000)
V2 = Total volume for 100 wells (say 100ul per well, therefore 100 x 100 = 10000ul).
Using the formula
4.88x10^6 cell/ml x Xml = 500000 x 10 ml
X = 1.024 ml
You would add 1.02 ml of the original cell suspension containing (4.88x10^6 cell/ml ) to 8.98 ml of culture media to get a total of 500000 cells in 10 ml. Mix well. When you add 100ul of this suspension to each well, you will be adding 5000 cells per well.
Good Luck!
