The demands of
research technology: Fluorescence, absorbance, and
luminescence assays have long generated demand for a
light standard. Recently, however, new technologies
have heightened that need. The genetic engineering of
fluorescent proteins, for example, has made it necessary
to measure light emissions from single molecules. The
widening adoption of biosensors has required researchers
to discern optical signals from additional new sources
and in real time.
Simultaneously, much research has moved from in vitro
to in vivo assays – to complex, high content screening
in live cells. Since cracking the genome, researchers
have eagerly sought to translate genetic code into protein
expression, and to trace that out along cellular and
physiological pathways. Drug discovery has consequently
moved into proteomics, cellomics, and physiomics, requiring
scientists to build assays around whole cells. This
requires finely controlled, minimal use of light to
guard against phototoxicity. It also requires super-sensitive
detection of the resulting signals.
If scientists wished to study the potential binding
of a receptor and ligand, for example, they might label
the former with one fluorochrome and the latter with
another. Staging this in a live cell, they’d have
to hold excitation to a minimum to avoid phototoxicity.
Their system would need to discern tiny fluorescent
signals, against a large background, in two colors simultaneously,
with high spatial and temporal resolution. To achieve
this, their light generation and detection systems would
need to be extremely well calibrated.
The demands of research collaboration: Adding
to the need for a light standard, virtually all biochemical
research is now done in teams, often spread across multiple
laboratories. For scientists to collaborate, their data
must be comparable and reproducible. That is, if researchers
run the same assay on two microscopes – or two
microplate readers – their results should be the
same. If they run the same assay on a single system
– but at different times – again, results
should be identical.
This is difficult, however, and it’s become increasingly
so in high-content research. To generate reproducible
results in cell-based assays, scientists need consistent
culture, media, coating, and time from plating to challenge.
They must monitor the consistency of the cells under
study, as well as their response to standard compounds.
Many researchers use profiling with TaqMan or microarrays
to check this. Regularly.
Unfortunately, after all this effort and expense, researchers
often lose consistency in their experiments nonetheless,
due to variations in the projection or detection of
light in their systems. |
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