Zoom Plate Assay Protocol - Bead-Based Fluorescent Sandwich Assay
- 5 µm diameter silica beads conjugated with capture antibody, 1% solid stored in 1% BSA-PBST
- Antigen standard
- Detection antibody conjugated with Alexa Fluor 647, 100 nM in 1% BSA-PBST
- Blocking buffer: 10% BSA in PBS
- Rinsing buffer: 1% BSA-PBST
Bead mixture incubation
|30min||Each bead mixture include 10 µl beads, 10 µl antigen standard, and 1 µl detection antibody|
Blocking with 50 µl blocking buffer
This step can be done concurrently with Step 1.
Inject at the center of the reaction zone. Pipette tip should be close to the membrane but do not touch the membrane.
Add 20 µl of bead mixture onto the membrane
|0||Inject at the center of the reaction zone. Pipette tip should be close to the membrane but do not touch the membrane.
For higher sensitivity, you can add 20 µl of rinsing buffer onto the membrane before adding the primary antibody to reduce background.
Rinse with 50 µl rinsing buffer for 4 times
When injecting, rest the pipette tip against the wall near the reaction zone. Do not pipette over the reaction zone as this is easy to introduce bubble.
The plate is now ready for fluorescent signal detection.