Fig.5 . E.coli separation from diluted blood. (A)
Qualitative analysis showing fluorescently labelled (using bacterial
viability kit) E.coli migrating to the inner wall and collected
in O1 while blood cells remain focused at the outer wall and collected
in O2. Scale bar : 100 µm. (B) Quantification of E.coli (1000
CFU/mL) spiked into three diluted blood samples (1:10, 1:5, and 1:2).
Agar plating based quantification show E.coli separation
efficiency between 90% to 82% depending on blood dilution.
For sepsis applications, ideally large volumes (2-7 mL whole blood) will
be necessary for any sample preparation method. Microfluidics, defined
as the science and technology of systems that can handle small volumes
of fluid in channels of ten to a few hundred microns in size, has proven
to be well suited for applications in clinical diagnostics. However,
while impressive progress has been made in high throughput rare
circulating tumor cell isolation, there has been limited success for
bacteria isolation from blood using microfluidics. In general, there is
a trade-of between separation efficiency and throughput for any given
technology. To this end, in comparison to the current state of the art,
our method is well suited for high separation efficiency without
compromising throughput much. In Fig.6, we compare work published using
both active (magnetophoresis 38, acoustophoresis32, 33 and dielectrophoresis 35, 36)
and passive (size-based separation 28, 31, 34, 37) to
our current work for bacteria isolation from blood. While the
experimental conditions (such as dilution ratio and bacteria
concentration) vary between the studies, making the comparison of
performance difficult, Fig 6 shows elegantly shows the compromise
brtween throughput and separation efficiency. While a more fair
comparison would be to use similar experimental conditions, e.i same
bacteria concentration, we found that ost of the work (∼ 70% of the
studies) used high bacteria concentration of ≥ 104CFU/mL28, 31, 32, 34, 35, 36,38 and
most of the work in inertial microfluidics that show good separation
efficiency have lower throughput 28-36 while studies
that highlight higher throughput show lower separation efficiency37, 38. The separation efficiency corresponding to the
three different blood dilutions in the current work is highlighted in
red in Fig.6. Using a single spiral, we demonstrated processing of 1 mL
of blood at extremely high throughput with separation efficiency
>80% for all the blood dilutions tested. To our best
knowledge, this is the highest throughput reported for a single passage
using a single chip at the given high separation efficiency. The
extremely high separation efficiency is attributed to the fact that
pre-positioned blood cells at the outer wall remain fully focused
throughout the length of the spiral channel while bacteria migrate. In
this work, we have focused on separating bacteria from RBCs and
nucleated cells, but given the size of platelets (2 to 4 µm) we
anticipate majority of platelets should also be separated based on the
size difference.