Bio-Rad Fluorescent Assay Kits - Manual (Page 7)

2. Sambrook, J., Fritsch, E. F., and Maniatis, T., Molecular Cloning: A
Laboratory Manual, Second Edition, Cold Spring Harbor Laboratory Press,
Cold Spring Harbor, New York, 16.66 (1989).
3. Seed, B., and Sheen, J. Y., Gene, 67, 271–277 (1988).
Section 7
Additional Supplies
Catalog
Number
Product Description
VersaFluor Fluorometer
170-2402 VersaFluor Fluorometer 100/120/220 , includes a standard
cuvette holder, 100 standard cuvettes, one excitation filter, and
one emission filter
170-2420 Optical Filter Excitation , EX360/40 (340–380 nm)
170-2421 Optical Filter Emission , EM460/10 (455–465 nm)
VersaFluor Disposable Cuvettes
170-2415 Standard Cuvette , 12.5 x 12.5 mm (OD) 4-sided optically clear
disposable, polycarbonate, 3.5 ml, 100
Fluoromark Microplate Fluorometer
170-6941 Fluoromark Microplate Fluorometer 120V , includes software,
5 excitation filters (355, 390, 485, 544 nm and time-resolved filter),
and 5 emission filters (405, 460, 538, 590 nm, and time-resolved
filter)
Accessories
223-9522 Disposable Transfer Pipet
170-6963 96-Well Fluorescence Microplate , solid black, 25
170-6964 96-Well Fluorescence Microplate , solid black with clear bot-
toms, 25
9
Section 5
Appendix
5.1 Microplate Assay
The 4MU standard curve (Section 4.3) and ß-galactosidase assay
(Section 4.4) may be scaled down 1:10 to be used in a 96-well microplate
platform. The assay is performed directly in microtiter plates.
Fluorescence units are determined in a microplate fluorometer.
1. Add 50 µl assay buffer (1x Reaction Buffer plus 0.6 mM MUG and
12.0 mM ß-mercaptoethanol) to each sample well.
2. Add samples (0.2–0.4 µg of total protein).
3. Incubate the microtiter plate for 15 minutes in a 37 °C incubator.
4. Stop the reactions by adding 150 µl 1x Stop Buffer. Pipet up and
down to mix.
5. Read samples in a microplate fluorometer.
5.2 Microplate Fluorometer Guidelines
Use the following guide for making measurements on the Bio-Rad
Fluoromark™. For other instruments used, refer to the instruction manu-
al regarding specific operation details.
Fluoromark settings:
Excitation filter 355 nm
Emission filter 460 nm
Gain 5
Flash 10
Section 6
References
1. Alam, J. and Cook, J., Analytical Biochemistry, 188, 245–254 (1990).
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