[diablo]

Hi,

I got a problem to solve (it's calculation): the following text was also in the descriptio but i think only the data (scatchard plot analysis ) is needed:

CR3, a heterodimeric complex of the CD11b (165kDa) and CD18(95kDa) membrane glycoproteins
is epxressed by NK cells. Evaluation of the binding affinity was conducted by Scatachrd plot analysis with concentrations of I-labeled D-glucan phsopate ranging from 5x10^-9   -   1x 10^-7 M and 10^6
cells tested in triplicate.



The following data were obtained (by Scatchard plot analysis)


Bound liigand [nM]:     4    9     11   20   24    33  (all x 10^-3)
Ligand nM:                 5   7,5   15   25   50   100

I guess that Ligand = free Ligand ?
Now i should calculate Kd for this interaction.

How can i calculate Kd ? --> do i have to draw a Scatchard plot ?


Plz hit back if u know something, thx alot

Greets Diablooooooo

[Yggdrasil]

You can estimate the k_d by plotting the data on a Scatchard plot.  I would think that Ligand means total ligand.

[diablo]

Hi and thx for your answer,

Yeah that was also my thought, but i get strange values if i calculate:

Total Ligands...[nM]      5 ----7,5-------15-----25--------50-------100
Bound Ligand..[nM] 0,004-- 0,009-0,011-0,02------0.024----0,033

for scatchardplot i have to calculate free ligands  (= Total Ligands minus Bound Ligands)and
additional the average n and n/freeligands (for the plot)

FreeLigands....[nM] 4,996---7,491---14,989---24,98---49,97--99,967
n(average) ...0,4...0,9...1,1....20...24...33 . (all x 10^-16)
n/freeligands...80...120....73.....80...48....33 (all x 10^-9)

if i plot this ( y-intercept = n, x-intercept = n/freeligands) i dont get a straigt line  and so i can't decide Kd (cause Kd = slope of the line )

Btw. the  n(average) i calculated this way: Boundligands/ cells(i.e.10^6) --> is this correct?
Plz help me, this drives me mad 

thx, Diablooooooooo

[Yggdrasil]

The calculations look alright to me, but yest the data do not give a very good Scatchard plot.  Maybe you'll just have to draw the best line you can and estimate the kd that way.

[diablo]

mkay, thx alot for your help & proofreading