Section 6- MicroArray
6. MicroArray
The capability to pre-select viable fluorescent-tagged hybridization probes for gene expression in micro arrays can
eliminate potentially flawed samples and improve research effectiveness. The NanoDrop
®
ND-1000 Spectrophotometer
measures the absorbance of the fluorescent dye, allowing detection at dye concentrations as low as 0.2 picomole per
microliter.
Fluorescent Dye Selection
There are currently nine fluorescent dyes that are hard-coded for use with the MicroArray module (see table below). Users
can also enter & save fluorescent dyes not coded within the ND-1000 software using the ‘Dye/Chromophore Editor’ button
found in the main menu. Dyes can be selected using the scroll arrows or by highlighting the Dye 1 or Dye 2 box. The
respective absorbance wavelength, extinction coefficient, and 260nm and 280nm % corrections will be automatically
utilized for measurement and concentration calculation. The default settings from NanoDrop remain Dye 1 set to Cy3 and
Dye 2 set to Cy5. In addition to the fluorescent dyes available from the drop-down menu, an option entitled ‘None’ is also
available. Selecting ‘None’ disables the respective calculations & numeric displays corresponding to that dye.
Note: Please refer to the dye manufacturer for the appropriate correction factors for user entered dyes.
Sample Volume Requirements
Field experience has indicated that 1 ul samples are sufficient to ensure accurate and reproducible results when
measuring aqueous nucleic acid samples containing incorporated fluorescent dyes. However, if you are unsure about the
surface tension properties of your sample or your pipettor accuracy, a 1.5-2 ul sample is recommended to ensure that the
liquid sample column is formed and the light path is completely covered by sample.
Measurement Concentration Range
The NanoDrop
®
ND-1000 Spectrophotometer will accurately measure fluorescent-dye and nucleic acid concentrations up
to 100 pmols/ul (Cy3) and 750 ng/ul (DNA) respectively without dilution. A table of sample concentration ranges is listed
below.
Sample
Type
Detection Limit
(pmol/ul)
Approx. Upper
Limit (pmol/ul)
Typical Reproducibility
(minimum 5 replicates)
(SD= pmol/ul; CV= %)
Cy3, Cy3.5, Alexa Fluor 555
and Alexa Fluor 660
0.20 100
sample range 0.20-4.0 pmol/ul:
± 0.20 pmol/ul
sample range >4.0 pmol/ul: ± 2%
Cy5, Cy5.5 and Alexa Fluor
647
0.12 60
sample range 0.12-2.4 pmol/ul:
± 0.12 pmol/ul
sample range >2.4 pmol/ul: ± 2%
Alexa Fluor 488 and Alexa
Fluor 594
0.40 215
sample range 0.40-8.0 pmol/ul:
± 0.40 pmol/ul
sample range >8.0 pmol/ul: ± 2%
Alexa Fluor 546 0.30 145
sample range 0.30-6.0 pmol/ul:
± 0.30 pmol/ul
sample range >6.0 pmol/ul: ± 2%
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