Bio-Plex Pro Human Immunotherapy Panel Quick Guide
Premixed Panels
Number of Wells
10x Detection
Antibodies, µl
Detection Antibody
Diluent HB, µl
Total Volume, µl
96
300
2,700
3,000
Singleplex Assays
Number of Wells
Singleplex #1
20x Detection
Antibodies, µl
Singleplex #2
20x Detection
Antibodies, µl
Detection
Antibody
Diluent HB, µl
Total
Volume, µl
96
150
150
2,700
3,000
Note:
20x singleplex beads allow multiplexing up to 20 analytes.
6.
Wash the plate three times
with
100 µl
wash buffer.
7.
Vortex
the diluted (1x) detection antibodies.
Add 25 µl
to each well.
8.
Cover the plate with sealing tape and incubate at
850 ± 50 rpm
for
30 min
at RT. Meanwhile, prepare the Bio-Plex Manager Software
protocol; enter standard S1 values and units provided in the assay kit.
9.
With 10 min left in the incubation,
vortex
100x streptavidin-phycoerythrin
(SA-PE) for
5 sec
and quick-spin to collect liquid.
Dilute to 1x
as shown
and protect from light.
Number of Wells
100x SA-PE, µl
Assay Buffer, µl
Total Volume, µl
96
60
5,940
6,000
10. Wash the plate three times
with
100 µl
wash buffer.
11. Vortex
the diluted (1x) SA-PE.
Add 50 µl
to each well.
12.
Cover the plate with sealing tape and incubate at
850 ± 50 rpm
for
10 min
at RT.
13. Wash the plate three times
with
100 µl
wash buffer.
14.
Resuspend the beads in
125 µl
assay buffer. Cover and shake at
850 ± 50 rpm
for
30 sec
.
15.
Remove the sealing tape and
read plate
using the following settings:
Instrument
RP1 (PMT)
DD Gates
Bead Events
Bio-Plex 3D*
Standard
Select MagPlex Beads
50
Bio-Plex 100, 200*
Low
5,000 (low); 25,000 (high)
50
Luminex MAGPIX
N/A, use default
instrument settings
* Or similar Luminex System.
