Keywords: Microfluidics, Micro/Nano Fluidic Device, Particle/Cell Sorting, Pinched Flow Fractionation, Hydrodynamic Filtration.
Summary. We proposed novel microfluidic methods for continuous, rapid and precise particle separation, named Pinched Flow Fractionation and Hydrodynamic Filtration. Here, the mechanistic features of those separation schemes and their applications to the continuous cell sorting are presented.
Microfluidic devices have several advantages over conventional fluidic systems due to their small dimensions, stable laminar flow patterns, easiness in duplicating the specific structures to make a large-scale array, and the flexibility in design. In this decade, therefore, a number of microfluidic systems have been developed for various applications such as biopolymer analysis, biological or environmental sensors, cell manipulation systems including analysis, sorting, treatments, and cultivation, and precise synthesis of novel materials. Recently, several hydrodynamic methods have been proposed for continuous particle sorting without employing outer fields such as electrical, magnetic, and gravitational forces. Our group has also proposed a technique for continuous, rapid and precise particle/cell separation as follows.
Pinched Flow Fractionation (PFF)
We have proposed a concept of ‘Pinched Flow Fractionation (PFF) , and achieved the successful separation of various types of sub-micron to micron-size structures including polymer beads, cells, droplets and macromolecules. In this method, particles can be separated according to their diameters simply by introducing liquid flows with and without particles into a microchannel, i.e., no outer field controls are needed. Also, in PFF method, separated particles can be continuously collected using multiple branch channels connected to the pinched segment. If the multiple collection channels are uniformly arranged in the PFF method, all branch channels are not effectively used since the liquid flow in the pinched segment is equally distributed to branch channels. To overcome this disadvantage, “asymmetric pinched flow fractionation (AsPFF)”  was proposed. The microfluidic device for AsPFF has a short drain channel, and a large portion of the liquid flow goes through the drain channel. Liquid flow containing particles is then efficiently distributed to other branch channels. Therefore, almost all branch channels can be effectively used for particle collection. The effluent positions of particles in PFF are determined by the microchannel structure designed beforehand and it is difficult to change the size range of the separated particles after fabrication. Therefore, we proposed “tunable pinched flow fractionation (tunable PFF)” , in which the effluent positions of the particles can be precisely tuned by controlling the flow rates distributed to each outlet using microvalves. And alternative method of precise tuning of PFF liquid flow using electroosmosis made a prompt change of flow distribution in the microchannel . Using PFF microchannel systems, micro-droplets were also separated based on their sizes only by introducing the emulsion from the inlet of the devices .
Hydrodynamic Filtration (HDF)
We have proposed another scheme for continuous separation of particles or soft matters utilizing laminar flow profiles in a microchannel network with multiple branch channels, named Hydrodynamic Filtration (HDF) , in which both concentration and classification of particles can be performed at the same time, by simply introducing a liquid containing a particle mixture. In this method, the hydrodynamics of laminar flow is also utilized in a microchannel having multiple branch points and side channels, not necessitating any outer field controls. In addition, not the cross-sectional size of the microchannel, but the flow profile inside the channel determines the size limit of the concentrated/classified particles. Therefore, a problem of channel clogging, which is similar to the mesh clogging or membrane fouling in the case of existing filtration methods, can be avoided. However, the inevitable contamination of small particles into the concentrated large-particle fraction decreases the separation efficiency. To overcome these disadvantages, we proposed an improved HDF scheme  for particle separation, employing flow splitting and recombining. By employing the modified HDF technique employing flow splitting and re-combining, the separation efficiency was improved. Several applications of these systems have been demonstrated for the separation of various kinds of biological materials including blood cells, hepatocytes, emulsions, bubbles and non-spherical cells/particles, and the cell treatment for a limited span of time [8-10].
 M. Yamada, M. Nakashima, M. Seki, Anal. Chem., 76, 5465 (2004).
 J. Takagi, M. Yamada, M. Yasuda, M. Seki, Lab Chip, 5, 778 (2005).
 M. Yamada, Y. Sai, M. Seki, J. Chromatogr. A, 1127, 214 (2006).
 T. Kawamata, M. Yamada, M. Yasuda, M. Seki, Electrophoresis, 29, 1423 (2008).
 H. Maenaka, M. Yamada, M. Yasuda, M. Seki, Langmuir, 24, 4405 (2008).
 M. Yamada, M. Seki, Lab Chip, 5, 1233 (2005).
 M. Yamada, M. Seki, Anal. Chem., 78, 1357 (2006).
 M. Yamada et al., Biomed. Microdev., 9, 637 (2007).
 R. Aoki, M. Yamada, M. Yasuda, M. Seki, Microfluid. Nanofluid., 6, 571 (2009).
 S. Sugaya, M. Yamada, M. Seki, Biomicrofluid, 5, 024103 (2011).