Molecular Devices FLIPR Tetra, User Manual

Page: 173 / 302

FLIPR
®
Tetra High Throughput Cellular Screening System User Guide
0112-0109 H 173
2. Remove one vial of FLIPR Calcium Assay Reagent
(Component A) and equilibrate to room temperature.
3. Dissolve contents of Component A vial by adding the
appropriate amount of 1X HBSS Buffer plus 20 mM HPES as
outlined in the table below. Mix by vortexing (~1–2 min) until
contents of vial are dissolved. It is important that contents are
completely dissolved to ensure reproducibility between
experiments.
4. Prepare the Loading Buffer by diluting the Component A vial
mixture with an additional volume of 1X HBSS plus 20 mM
HEPES Buffer as outlined in table below. Multiple washes of the
vial are necessary to completely transfer the contents.
CAUTION! The components supplied are sufficient for proper cell
loading. For optimum results it is important NOT to add any additional
reagents or change volumes and concentrations.
Table 8-1 Quantities of 1X HBSS necessary to dissolve Component A
contents.
Plate Format
Explorer Kit
(R8142)
Bulk Kit
(R8141)
Express Kit
(R8143)
96- or 384-well 10 mL 10 mL 20 mL
1536-well 6.5 mL 10 mL 20 mL
Table 8-2 Quantities of 1X HBSS plus 20 mM HEPES necessary to
perform second dilution of Component A.
Plate Format
Explorer Kit
(R8142)
Bulk Kit
(R8141)
Express Kit
(R8143)
96- or 384-well NA 90 mL 480 mL
1536-well NA 55 mL 300 mL
Note: If your cells require probenecid, then a stock solution should
be prepared in 1 M NaOH and added fresh to 1X HBSS buffer plus
20 mM HEPES (adjust pH to 7.4 after addition of probenecid) for
preparation of the Loading Buffer so that the final in-well working
concentration is 2–2.5 mM. Do not store frozen aliquots of Loading
Buffer with probenecid and always prepare fresh probenecid on the
day of the experiment.