Market size for gene chips, lab on chips, sensors, flow controllers, nozzles, valves, pressure measurement, actuators, relays, sata storage, strain sensors $4-8Bln in 2003 USD.
(from DARPA estimates, so how good can those be!!?)
Look at Burns, Johnson, et. al. Science 282, 484 (1998) for (first?) bio lab on a chip paper.
Issues: data acquisition from chip to computer. diagnoses times/latencies
Review Capillary electro-chromatography
Overall architectural considerations:
- kinematic properties, transport properties, thermodynamic properties, surface tension, vapor pressure, surface accomodation
- Fluidic interconnects - standards? universal interconnects?
- pumps and valves - fabrication, integration, elastomers
- fluid injection = volume vs. sensitivity (if you only have one cell worth of protein, and add fluid you need sensitive sensors
- Confining molecules for detection
- kinetics of chemical reactions
- molecule activitiy
- intensifying and analyzing role of surfaces in catalytic reactions
- applying magnetic fields to conotrol reactions
- labelling molecules
- biomolecules with NEMS
- amplification of other than DNA biological structures
- non-newtonian fluids
Viable marketable serviecs with demand/practical application today
- sequenced low-nanogram scale bacterial and mammalian DNA
- support of more efficient PCR prep, see example overview of the issues here:
"Accurate quantification of the sequencing library is essential to achieve high yield and high quality sequencing. Inaccuracy in quantification is addressed by the manufacturers through ‘titration’ runs of the sequencer, which are used to empirically divine the concentration of productive DNA fragments in the sequencing library."
Overview of modern sequencing:
In vitro clonal amplification
As molecular detection methods are often not sensitive enough for single molecule sequencing, most approaches use
an in vitro cloning step to generate many copies of each individual molecule. Emulsion PCR is one method, isolating
individual DNA molecules along with primer-coated beads in aqueous bubbles within an oil phase. A polymerase chain
reaction (PCR) then coats each bead with clonal copies of the isolated library molecule and these beads are
subsequently immobilized for later sequencing. Emulsion PCR is used in the methods published by Marguilis et al.
(commercialized by 454 Life Sciences, acquired by Roche), Shendure and Porreca et al. (also known as "polony
sequencing") and SOLiD sequencing, (developed by Agencourt and acquired by Applied Biosystems).
Another method for in vitro clonal amplification is "bridge PCR", where fragments are amplified upon primers
attached to a solid surface, developed and used by Solexa (now owned by Illumina). These methods both produce
many physically isolated locations which each contain many copies of a single fragment. The single-molecule method
developed by Stephen Quake's laboratory (later commercialized by Helicos) skips this amplification step, directly
fixing DNA molecules to a surface.
Once clonal DNA sequences are physically localized to separate positions on a surface, various sequencing approaches
may be used to determine the DNA sequences of all locations, in parallel. "Sequencing by synthesis", like the popular
dye-termination electrophoretic sequencing, uses the process of DNA synthesis by DNA polymerase to identify the
bases present in the complementary DNA molecule. Reversible terminator methods (used by Illumina and Helicos) use
reversible versions of dye-terminators, adding one nucleotide at a time, detecting fluorescence corresponding to that
position, then removing the blocking group to allow the polymerization of another nucleotide. Pyrosequencing (used
by 454) also uses DNA polymerization to add nucleotides, adding one type of nucleotide at a time, then detecting and
quantifying the number of nucleotides added to a given location through the light emitted by the release of attached
"Sequencing by ligation" is another enzymatic method of sequencing, using a DNA ligase enzyme rather than
polymerase to identify the target sequence. Used in the polony method and in the SOLiD technology offered
by Applied Biosystems, this method uses a pool of all possible oligonucleotides of a fixed length, labeled according to
the sequenced position. Oligonucleotides are annealed and ligated; the preferential ligation by DNA ligase for
matching sequences results in a signal corresponding to the complementary sequence at that position.
- hardware and software for controlling, modeling, engineering flows in the chip, for example: