# HEMOCYTOMETER QUESTION - HEMOCYTOMETER (Oct/19/2006 )

Its been a while since i used a hemocytometer - whats the best calculation for determining cell density? thanks

-BioDude1-

QUOTE (BioDude1 @ Oct 19 2006, 11:29 AM)
Its been a while since i used a hemocytometer - whats the best calculation for determining cell density? thanks

Average # of cells X dilution factor X 10,000 = (# cells/mL) / 1000 = # cells/ uL

-saudata-

this is how I count.

count the 16 small squares, there r 8 of these - count them all

average them

multiply x 10,000 = # of cells in a ml.

-scolix-

Method A

Count the number of cells in the 4 outer squares (see the left panel of Figure 2).
The cell concentration is calculated as follows:
Cell concentration per milliliter = Total cell count in 4 squares x 2500 x dilution factor
Example: If one counted 450 cells after diluting an aliquot of the cell suspension 1:10, the original cell concentration = 450 x 2500 x 10 = 11,250,000/ml

Method B

Estimate cell concentration by counting 5 squares in the large middle square
The cell concentration is calculated as follows:
Cell concentration per milliliter = Total cell count in 5 squares x 50,000 x dilution factor
Example: If one counted 45 cells after diluting an aliquot of the cell suspension 1:10, the original cell concentration = 45 x 50,000 x 10 = 22,500,000/ml

-jmreddy-

I came across this question about a hemocytometer when my lab mate was having problems seeding the correct amount of cells on her plates. We both came from different labs so our our method of counting is a little different and we get different # of cells. She got 21.5x10^6 total # of cells and i got 20.9x10^6. so that made me wonder exactly how accurate is the # you get from the hemocytometer and whether the difference in cell # is that big of a deal?

-pumpkin21-

21.5x10^6 is so close to 20.9x10^6
anyway, haemocytometer is just used to estimate the number

-strawberry-

taht's what i was thinking also.. that the 2 # were so close, but she was making a big deal of it so it just made me kinda wonder.. thanks!! =)

-pumpkin21-

Hi , I have a question regarding cell counting and the dye exclusion test :

I have always used the dilution factor when doing the counting , but the lab I am in right now the calculation that they do am not able to follow â€“
I â€˜ve been told to count the number of cells in any of the 2 opposite squares take their average .Take the average cell count and multiply it with 10 ^ 4

i.e. if I get 32 n 33 in 2 squares I divide it by 2 to get = 32.5 * 10^4 cells / ml
Is this correct ?? Am not sure
Also when I mix the cell suspension n tryphan blue â€“ I take 100 micro lt of cell susp + 100 micro lt of dye , mix it well n load 10 micro lt onto the hemocytometer .

If I do this type of procedure what sort of dilution factor I have to take into account if any?? Please let me know what to do or is this correct. !! Thanks

-seashell83-

also, you need to make sure you know the correct way of using a hemacytometer.
there's an undergrad in our lab make a mistake by putting tooo much cell suspension into hemacytometer and thus, his calculation will not represent the correct numbers of cell.

-sanjiun81-

A short guide to use hemocytometer

-tonix37-