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### #1 bio08

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Posted 13 April 2010 - 01:33 AM

Hello,
If you want to make a 6 ml cell suspension with 1 mil cells/ml.

You counted the cells in your 4 ml cell suspension and it was 1000 000 cells/ml
So in 4 ml cell suspension you have: 4 ml* 1 000 000 cells/ml= 4 000 000 cells in total

4 mil cells/1 mil  = 1 ml cells, So from the 4 ml cell suspension you take 1 ml and since you need 6 ml for the experiment so you add 5 ml of media and in this way you have 6 ml for the experiment.

Is the calculation right?

Thanks!

Edited by bio08, 13 April 2010 - 01:33 AM.

### #2 Doki

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Posted 13 April 2010 - 01:53 AM

not true.

cells U have :

Quote

1 million/ml cell suspension. Total volume : 4 ml

so total number of cells U have = 4 million.

Cells U need for experiment :

Quote

cell dilution : 1 million/ml
total volume : 6ml
so total number of cells needed = 6 million.

U are short of 2 million cells if I understood it right.

Edited by Nabi, 13 April 2010 - 01:54 AM.

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### #3 bio08

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Posted 13 April 2010 - 02:26 AM

Thanks for the answer!
But if I have 8 mil cells in the 4 ml cell suspension and I need 6 ml with the cell concentration 1 mil/ml

So in 6ml I will have 6 mil cells.

8 mil/ 6 mil= 1,3 ml, So does it mean I will take 1,3 ml of the 4 ml cell suspension and since I need 6 ml in total volume for the experiment so I add 4,7 ml media. In this way I have 6 ml for the experiment.

Is it Right?

Thanks!

### #4 Doki

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Posted 13 April 2010 - 08:32 AM

bio08.

make the calculations simple.

In 4 ml suspension, there are 8 million cells.
so, if U take 1 ml of the suspension, there will be 2 million cells ( 8 million divided by 4 ml).

U need 6 million cells.

if there are 2 million cells in 1 ml of the suspension,
to get 6 million cells, U will need 3 ml of the suspension  ( 1 ml + 1 ml + 1 ml = 2 million cells + 2 million cells + 2 million cells)

so, if U take 3 ml of the original cell suspension U had, U will have 6 million cells.

next

Now, u have 6 million cells in 3 ml; but U need to have 6 million cells in 6 ml. So, to get that, add 3ml of media.

Then, U will get 6 million cells in 6 ml.

Do u have any confusion. Please clear your confusion before you start any further experiments. We can help you here or better ask some seniors to show U how these calculations are done.

Good luck.
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### #5 bio08

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Posted 13 April 2010 - 12:27 PM

Thank you very much Nabi!
Could you please see if my calculations below is right:
1.
I centifuge my cells and dissolve the pellet in 5 ml media.

In 5ml cell suspension I have 1,77 mil cells/ml
In this 5 ml cell suspension I counted the cells and I have 1,8 mil cells/ml

I want 0,5 mil/ml in 17,7 ml cell suspension for an experiment

So 0,5 mil/ml * 17,7 ml= 8,85 mil cells
So in 1 ml I have 1,8 mil cells and I need 8,85 mil cells.
8,85 mil cells/1,8 mil cells=  4,9 ml but I need 17,7 ml so to the 4,9 ml cell suspension I add   12,8 ml media.
Is this calculation right?

My second question is:
2.

I centifuge my cells and dissolve the pellet in 5 ml media.

In 5ml cell suspension I have 250 mil cells/ml
In this 5 ml cell suspension I counted the cells and I have 1,8 mil cells/ml

I want 1 mil/ml and want to make 10 ml cell suspension

So I have 250 mil cells  in 1ml
10 ml *10^6  cells= 10 mil cells  ( in 10 ml I have 10 mil cells)

10 mil cells/ 250 mil cells = 0,04 ml= 40 ul. So I take 40 ul from my 5 ml cell suspesnion, but I need 10 ml in total volume so to the 40 ul cells ( I took from my 5ml cell suspension) I will add 9,96 ml media.

Is this calcultation right or this one below:

In 5ml cell suspension I have: 5ml *250 mil cells/ml= 1250 mil cells

1250 mil/10 mil = 8 ul I will take from the 5 ml cell suspension and I will add around 9,92 ml media

3. Could you also please see if this calculation is right:

I centifuge my cells and dissolve the pellet in 5 ml media.
In this 5 ml cell suspension I counted the cells and I have 0,95 mil cells/ml

I need 0,5 mil cells/ml in total volume 1,5 ml
So 1,5 ml * 0,5 cells/ml=0,75 mil cells/ml

0,75 mil / 0,95 mil = 790 ul I will take from my 5 ml cell suspension and add 710 ul media to get 1,5 ml total volume

Is this calculation right?

Thank you very much again!

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Posted 13 April 2010 - 01:07 PM

so many words about such simple thing

### #7 bio08

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Posted 15 April 2010 - 01:50 PM

I am a beginner so I am sorry for asking but Nabi encouraged me.
I still hope someone can help me

### #8 Doki

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Posted 15 April 2010 - 04:10 PM

vladooo, on Apr 14 2010, 06:07 AM, said:

so many words about such simple thing
vladoo, yeah, U r right. It is a simple problem but too many words were used. Can U please simplify so that bio08 can get the basics right. That would be a big help here.

Quote

I am a beginner so I am sorry for asking but Nabi encouraged me.
I still hope someone can help me

bio8, U seem to be weak in calculations and might need more help. Don't you have any seniors who could help you show how these calculations are done? Like valdoo said, these are some of the simplest calculations U will be doing; so for others U will be needing better understanding.
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### #9 Doki

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Posted 15 April 2010 - 04:24 PM

bio08, on Apr 14 2010, 05:27 AM, said:

1.
I centifuge my cells and dissolve the pellet in 5 ml media.

In 5ml cell suspension I have 1,77 mil cells/ml
In this 5 ml cell suspension I counted the cells and I have 1,8 mil cells/ml

I want 0,5 mil/ml in 17,7 ml cell suspension for an experiment

So 0,5 mil/ml * 17,7 ml= 8,85 mil cells
So in 1 ml I have 1,8 mil cells and I need 8,85 mil cells.
8,85 mil cells/1,8 mil cells=  4,9 ml but I need 17,7 ml so to the 4,9 ml cell suspension I add 12,8 ml media.
Is this calculation right?

You either have 1.77 million cells/ml or 1.8 million cells/ml in the suspension.

If U had 1.8 million cells/ml then your calculations are right.

U started with 5 ml of 1.8 million cells/ml which has 9 million cells in total (1.8 X 5). U needed to make 17.7 ml of 0.5 million/ml cell suspension which will require  8.85 million cells (17.7 X 0.5).  8.85 million cells are in 4.92ml of the cell suspension U had [ (8.85/9) X 5 ].

So, U took 4.92ml from the cell suspension U had (which will have 8.85 million cells) and to make 0.5million/ml cell suspension, U needed to add 12.78ml of medium (17.7-4.92)
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### #10 Doki

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Posted 15 April 2010 - 04:26 PM

bio08, on Apr 14 2010, 05:27 AM, said:

2.
I centifuge my cells and dissolve the pellet in 5 ml media.

In 5ml cell suspension I have 250 mil cells/ml
In this 5 ml cell suspension I counted the cells and I have 1,8 mil cells/ml

I want 1 mil/ml and want to make 10 ml cell suspension

So I have 250 mil cells  in 1ml
10 ml *10^6  cells= 10 mil cells  ( in 10 ml I have 10 mil cells)

10 mil cells/ 250 mil cells = 0,04 ml= 40 ul. So I take 40 ul from my 5 ml cell suspesnion, but I need 10 ml in total volume so to the 40 ul cells ( I took from my 5ml cell suspension) I will add 9,96 ml media.

Is this calcultation right or this one below:

In 5ml cell suspension I have: 5ml *250 mil cells/ml= 1250 mil cells

1250 mil/10 mil = 8 ul I will take from the 5 ml cell suspension and I will add around 9,92 ml media

If U got 2 answers for same mathematical calculations then one or both might be wrong. Can U do it like U did No. 1 and see which one is right?
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### #11 bob1

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Posted 15 April 2010 - 04:30 PM

The easiest thing to do is to go back to the equations -

For dilutions use Vi*Ci = Vf*Cf where V=volume, C=concentration, i= initial and f=final.

from your post above you have a cell suspension at 1.77*106 cells/ml (Ci)

and you want 0.5*106 cells/ml (Cf) in a volume of 17.7 ml (Vf)

```so Vi * 1.77*10^6 cells/ml = 17.7 ml * 0.5*10^6 cells/ml

Vi * 1.77*10^6 cells/ml = 8.85*10^6 cells

Vi = 8.85*10^6 cells
-----------------
1.77*10^6 cells/ml

Vi= 5 ml```

So you take 5 ml of your cell suspension and add it to 12.7 ml (17.7ml - 5ml = 12.7ml) of medium.

If you always go back and state the variables you have and what you are looking for and the equation you are going to use to solve the problem, you shouldn't have difficulties! Also remember to keep your units the same always (i.e. if you are using ml in an equation keep using ml even one of the variables is in litres)

If you just want to know how many cells you have use V=N/C where N= number.  Same equation used for making chemical solutions in the lab!

Edited by bob1, 15 April 2010 - 04:34 PM.

### #12 Doki

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Posted 15 April 2010 - 04:36 PM

bio08, on Apr 14 2010, 05:27 AM, said:

3. Could you also please see if this calculation is right:

I centifuge my cells and dissolve the pellet in 5 ml media.
In this 5 ml cell suspension I counted the cells and I have 0,95 mil cells/ml

I need 0,5 mil cells/ml in total volume 1,5 ml
So 1,5 ml * 0,5 cells/ml=0,75 mil cells/ml

0,75 mil / 0,95 mil = 790 ul I will take from my 5 ml cell suspension and add 710 ul media to get 1,5 ml total volume

Is this calculation right?

Thank you very much again!

Yes, U r right.

U started with 5 ml of 0.95 million cells/ml which has 4.75 million cells in total (0.95 X 5). U needed to make 1.5 ml of 0.5 million/ml cell suspension which will require 0.75 million cells (1.5 X 0.5). 0.75 million cells are in 0.789 ml of the cell suspension U had [ (0.75/4.75) X 5 ].

So, U took 0.789 ml from the cell suspension U had (which will have 0.75 million cells) and to make 0.5million/ml cell suspension, U needed to add 0.711 ml of medium (1.5-0.789)
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