DNA microarray enhancement using a continuously and discontinuously rotating microchamber.
It is demonstrated that the most efficient way to enhance DNA microarray analysis consists of a maximal reduction of the total device volume (to keep the concentration of the available DNA as high as possible), combined with the creation of a strong lateral convective transport of the sample. In the present study, DNA microarray hybridizations are performed in a set of rotating, circular microchambers covering exactly the spotted area of the microarray and with a depth varying between 70 and 1.6 microm. Rotating the microchamber substrate while keeping the microarray stationary, the rotating microchamber bottom wall literally drags the sample past the microarray spots with a velocity which is independent of the fluid layer thickness. Interestingly, it was found that transporting the sample in a discontinuous mode (with stop periods of several minutes) not only yields a more stable and reproducible operation, it also yields significantly larger hybridization intensities (typically a factor of 2-3 larger) than a continuous rotation. This seems to be due to the fact that the velocity field disturbs the binding process at the binding site level. Working under limiting DNA sample mass conditions, the system yielded in a short, 30-min experiment already a 5-fold increase of the hybridization intensity, as compared to a conventional microscope slide/coverslip system operated overnight under diffusion-driven conditions. Compared to a commercial pump-around hybridization system, the gain was even more impressive, precisely due to the fact that the pump-around system requires larger volumes, which with a fixed amount of available genetic material leads to the application of more diluted samples.
Vanderhoeven
Johan
J
Department of Chemical Engineering, Vrije Universiteit Brussel, Brussels, Belgium, MicroArrayFacility Lab, Flemish Institute for Biotechnology (VIB), Leuven, Belgium. Johan.Vanderhoeven@vub.ac.be
Pappaert
Kris
K
Dutta
Binita
B
Van Hummelen
Paul
P
Desmet
Gert
G
D018076:DNA, Complementary / Q000032:analysis; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods*; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation* / Q000379:methods*
D018076:DNA, Complementary