TABLE 2.
Renewable microcolumn program for evaluating the tunable surface hypothesis
| Manipulation | Solution | Vol (μl) | Flow rate (μl s−1) |
|---|---|---|---|
| Form column (rod in trap position)a | Top layer | ||
| Packing layer | Phosphate-buffered saline-Tween 20 | 100 | 10 |
| Luminex particles | HS or LS phosphate | 50 | 10 |
| Rinse beads | Phosphate buffers (see Results) | 140 | 10 |
| Perfuse with denatured rRNA target | Room temp or 45°C (see Results) | 50 | 0.5 |
| Wash beads | Phosphate buffers (see Results) | 100 | 1 |
| Release columnb | |||
| Reverse flow | 2× SSC-Tween 20 | 300 | 100 |
| Rotate rod to release | |||
| Flush column | 100 | ||
| Clean flow cell | 2× SSC-0.02% Tween 20 | 450 | 200 |
a
For the bead packing protocol, 5 × 104 19.9-μm beads in a 100-μl volume were injected into the rotating-rod microcolumn, followed by 100 μl of Luminex bead suspension containing 5 × 103 beads of each color.
b
Samples collected from the rotating rod were centrifuged and resuspended in 300 μl of 1× SSC-0.01% Tween 20 for direct analysis on the Luminex flow cytometer.