Luminex xMAP Antibody Coupling Kit - Manual (Page 19)

Page 15 of 20
4. Prepare a solution of phycoerythrin-labeled anti-species IgG detection antibody at 4
μg/mL in PBS-1% BSA. Prepare a 1:2 dilution series of that detection antibody
solution to a concentration of 0.0625 μg/mL as shown in the following table.
NOTE: For optimal results use Costar round-bottom 96-well plates and Luminex
Magnetic Plate Separator with MagPlex microspheres.
5. Aliquot 50 μL of the microsphere solution prepared in Step 3 into two entire columns
of wells of the plate (duplicate sets of 8 wells, 16 wells total).
6. Add 50 μL of PBS-1% BSA, as a blank sample, into the wells in Row A containing
the microsphere solution.
7. Add 50 μL of each of the diluted detection antibody solutions prepared in Step 4 into
the appropriate wells of the plate (as shown in the plate layout below).
1 2 3 4 5 6 7 8 9 10 11 12
A Blank Blank
B 1:64 1:64
C 1:32 1:32
D 1:16 1:16
E 1:8 1:8
F 1:4 1:4
G 1:2 1:2
H 1:1 1:1
Example plate layout using columns 1 & 2
Dilution Tube
Volume of
PBS-1% BSA
Volume of Detection
Antibody
Concentration
1:1 - - 4 μg/mL
1:2 500 μL 500 μL from Tube 1:1 2 μg/mL
1:4 500 μL 500 μL from Tube 1:2 1 μg/mL
1:8 500 μL 500 μL from Tube 1:4 0.5 μg/mL
1:16 500 μL 500 μL from Tube 1:8 0.25 μg/mL
1:32 500 μL 500 μL from Tube 1:16 0.125 μg/mL
1:64 500 μL 500 μL from Tube 1:32 0.0625 μg/mL