You did an experiment that required to have a large number 5

You did an experiment that required to have a large number, 5,animals /group. At the day of the assay you pooled the spleens of each group. You made single cell suspension, washed the cells and reconstituted the spleens in 30 mls of media. Subsequently you diluted your stock 5 fold and took a small sample from this tube and you diluted it further 1:2 in Trypan Blue You counted viable white cells present in the spleen. You counted 105 cells in 15 squares of the 25 squares of the L5 region of the Hemocytometer. Use the stock cells of the above experiment and do the followings:

- Make serial two-fold dilutions. Calculate the concentration of cells/1 ml in the 6th dilution tube Indicate with volumes of your choice of how you can do 2 fold dilutions

- Calculate the Total number of cells present in 7th dilution tube (remember the volumes used for these dilutions are of your choice)

Solution

The number of viable cells = dilution factor* No. of cells* 10^4

= 5*105*10^4

= 525 * 10^4 cells in stock / ml

for 30 ml the cells are 1575 *10^5

dilutions

a) The concentration of cells/ 1ml in a 6th dilution tube is 0.72 * 10^4 cells/ml = 7200 cells/ml

Initial volumer = 1 ml

serial two fold dilution - 1/3 dilution

1ml has 525 * 10^4 --------1st tube175*10^4 --------58 * 10^4 ------19.5 * 10^4 ------6.5* 10^4 ------2.1* 10^4------6th tube   0.72 * 10^4

7th dilution tube will have 2400 cells or 0.24 * 10^4 cells