Biomedical Engineering Reference
In-Depth Information
The Quantum Dot Corporation (now part of Invitrogen) has also developed
magnetic microspheres encoded by what appears to be some form of LBL process,
although insuffi cient details have been revealed to be certain of this [44]. The
microspheres have a diameter of 8
m and are encoded with up to four colors of
QDs at 12 different intensity levels; in theory, this could yield up to 455 spectral
codes. The encoding QDs had emission peaks at 525, 545, 565, and 585 nm, while
a fi fth QD conjugated to streptavidin with an emission peak at a longer wavelength
was used as a reporter label. The encoded microspheres were used for the multi-
plexed gene expression profi ling of 100 cRNA sequences and 20 calibrator
sequences. The results clearly demonstrated the advantages of performing multi-
plexed detection with suspension arrays combining QD encoding and magnetic
separation. In comparison with microarrays, the duration of the assay was an order
of magnitude faster, the dynamic range was 2-3 log units broader; the sensitivity
was an order of magnitude better, and reproducibility approached that of an
Affymetrix GeneChip microarray. Moreover, quality control was straightforward
because encoded microspheres can be prepared in gram-sized batches, with each
gram being suffi cient to perform at least 10
9
assays.
μ
2.7.1
Magnetically Encoded Suspension Arrays
The possibility of encoding suspension arrays with a combination of luminescent
and magnetic entities has been described elsewhere [54]. The maximum number
of spectral codes that can be resolved by fl uorescence alone is ultimately limited
by the ability of the detector to distinguish between different colors and intensities.
The number of resolvable codes can be doubled by the simple expedient of incor-
porating additional magnetic encoding elements into some of the fl uorescent
particles. Two suspension arrays with identical spectral codes are prepared, with
the single difference that one of them is based on magnetic particles; the arrays
can therefore be separated by the application of a magnetic fi eld gradient before
reading the spectral code. In a variation on this idea, Kim and Park carried out
dual analyte sandwich immunoassays in which the identity of captured analyte
molecules was indicated by the fl uorescence code of the microsphere, and the
amount of target molecule captured by them was determined from their velocity
in an applied magnetic fi eld [55]. Particles in a suspension array can also be identi-
fi ed by means of magnetic codes alone. The force on a magnetic particle in a
magnetic fi eld depends on its size and composition; thus, particles in a suspension
arrays could in theory be separated and identifi ed on the basis of their magnetic
content. The separation of particles in a magnetic fi eld, which is known as mag-
netophoresis or magnetic spectrometry, has been used to separate populations of
cells labeled with antibodies conjugated to magnetic nanoparticles [56, 57], and
more recently to separate particles on the basis of their size and magnetic moment.
In one of these reports, magnetic particles were injected into a planar fl ow cell
and separated into eight outlet bins by the application of a magnetic fi eld gradient
at right-angles to the direction of fl ow (see Figure 2.11) [58]; similar separations
