Preparing 48-Well Plate Feeders







In preparing 48-well plates, a dilution must be made from the regular

density of 3.5 x 105  cells/ml.   The required density for 48-well 

plates is:  8.2 x 104  cells/ml  (650 ml/well  or  31.2 mls/plate).



Example:



For a total of 5 x 48-well plates, the calculation is:



5 plates x 31.2 mls  =  156 mls  +  10 mls Extra  =  166 mls  (Total)



166 mls x 8.2 x 104 cells/ml  =  1.36 x 107 cells  (Total)



1.36 x 107 cells  :  3.5 x 105 cells/ml  =   38.86 mls



Dilution:  38.9 mls (of cell suspension  @ 3.5 x 105 cells/ml) + 127.1  mls Media.



Take a count at this point, to verify the cell density of 8.2 x 104 cells/ml.   

Plate-out this feeder media (dilution) into the 48-well plates.    

REMEMBER TO USE PLATES THAT HAVE BEEN GELLED, 2 HOURS PRIOR THIS.    

Dispense 650 ml of feeder media/well (31.2 mls/plate).   

Mix plates to distribute cells uniformly and incubate.





No. of Plates     Total # of Cells     Dilution (mls)     Total Vol. (mls)

 5 plates         1.36 x 107 cells     38.9  +  127.1      166  mls

10 plates         2.64 x 10 7 cells    75.0  +  247.0      322

15 plates         3.92 x 10 7 cells   112.0  +  366.0      478

20 plates         5.20 x 10 7 cells   148.5  +  485.5      634





Feeders/STO  Media:  DMEM,  7% FCS,  1 X GPS.