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Data Acquisition
1. Open the Acquisition template on the BD™ CBA Software.
Note: Following installation on Macintosh computers
using BD CBA Software Version 1.0, the template can
be found in the BD Applications/BD CBA folder/Sample Files/
Mouse Isotyping Files/Instrument Setup folder. For
BD CBA Software Version 1.1 or higher, the template can
be found in the BD Applications/BD CBA folder. It is
labeled “Isotype Kit Acquire Template”. Alternatively, the
Acquisition template may be downloaded via the internet from:
bdbiosciences.com/pharmingen/CBA/downloads.shtml
2. Set acquisition mode and retrieve the optimized instrument settings from
Instrument Setup with the Cytometer Setup Beads on pg. 14.
3. In the Acquisition and Storage window, set the resolution to 1024.
4. Set number of events to be counted at 1800 of R1 gated events.
(This will ensure that the sample file contains approximately
300 events per Capture Bead).
5. Set number of events to be collected to “all events”. Saving all events collected
will ensure that no true bead events are lost due to incorrect gating.
6. In setup mode, run tube no. 1 and using the FSC vs. SSC dot plot, place
the R1 region gate around the singlet bead population (see Figure 3a).
7. Samples are now ready to be acquired.
8. Begin sample acquisition with the flow rate set at HIGH.
Note: Run the negative control tube (0 pg/ml standards) before any
of the recombinant standard tubes. Run the control assay
tubes before any unknown test assay tubes. Run the tubes in the
order listed in Table 2 of BD CBA Mouse Inflammation Assay
Procedure on pg. 12.
To facilitate analysis of data files using the BD CBA Software
and to avoid confusion, add a numeric suffix to each file that
corresponds to the assay tube number (ie, Tube No. 1 containing
0 pg/ml could be saved as KT032598.001). The file name must
be alphanumeric (ie, contain at least one letter).
