Chapter 5: Sorting 163
Using Auto Drop Delay
1 Set up an experiment for drop delay as described in Setting Up the
Experiment on page 158.
2 Load a tube filled with a suspension of Accudrop beads (approximately 2
drops of beads in 0.5 mL PBS).
3 Adjust the flow rate to achieve these values of events per second:
70 micron = 1,000 to 3,000
85 micron = 800 to 2,000
100 micron = 600 to 1,500
130 micron = 400 to 1,200
Tip If this cannot be achieved using a flow rate setting between 1 and 5,
adjust the bead concentration.
4 Turn on the voltage in the Side Stream window. Click Sort in the Sort
Layout window.
5 Click Cancel at the Confirm dialog.
There is no need to collect the beads. When the drawer is closed, the beads
are sorted to waste.
6 Adjust the micrometer dial (see Figure 1-17 on page 41) to obtain the
brightest bead spot on the center stream.
7 Click the Auto Delay button in the sidestream window.
A dialog similar to Figure 5-5 on page 164 opens. The graph in the left
pane of the dialog represents brightness of the stream (Y axis) versus drop
delay (X axis). Typically the graph should have a flat portion, with several
small peaks in either direction (up or down) or one prominent peak.
NOTE If the original drop delay value was close to the appropriate value
prior to starting auto delay, then you may not get a graph displayed
because few data points were required to determine the optimal drop delay.
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