Highly Efficient and Ultra‐small Volume Separation by Pressure‐Driven Liquid Chromatography in Extended Nanochannels

Abstract

The rapidly developing interest in nanofluidic analysis, which is used to examine liquids ranging in amounts from the attoliter to the femtoliter scale, correlates with the recent interest in decreased sample amounts, such as in the field of single-cell analysis. For general nanofluidic analysis, the fact that a pressure-driven flow does not limit the choice of solvents (aqueous or organic) is important. This study shows the first pressure-driven liquid chromatography technique that enables separation of atto- to femtoliter sample volumes, with a high separation efficiency within a few seconds. The apparent diffusion coefficient measurement of the unretentive sample suggests that there is no increase in the viscosity of toluene in the extended nanospace, unlike in aqueous solvents. Evaluation of the normal phase separation, therefore, should involve only the examination of the effect of the small size of the extended nanospace. Compared to a conventionally packed high-performance liquid chromatography column, the separation here results in a faster separation (4 s) by 2 orders of magnitude, a smaller injection volume (10(0) fL) by 9 orders, and a higher separation efficiency (440,000 plates/m) by 1 order. Moreover, the separation behavior agrees with the theory showing that this high efficiency was due to the small and controlled size of the separation channel, where the diffusion through the channel depth direction is fast enough to be neglected. Our chip-based platform should allow direct and real-time analysis or screening of ultralow volume of sample.

Investigation of Unique Protonic and Hydrodynamic Behavior of Aqueous Solutions Confined in Extended Nanospaces

“…By utilizing the ultra-small and enclosed spaces and large surface-to-volume ratio, various kinds of applications, such as chemical and biochemical devices, including DNA [43][44][45][46] and ion separations, [47][48][49][50][51] ion condensation, [52][53][54] and ion rectification [55][56][57] and energy devices, [58,59] have been already developed. Their unique performance has been explained in terms of size-confinement effects on liquid properties, such as viscosity, dielectric constant, pH, and ion mobility and transportation, etc.…”

Reversed-phase Chromatography in an Extended Nanospace: Separating Amino Acids in Short and Long Nanochannels

“…1). 5 The four ends of the nanochannels were linked to microchannels. Each end of the microchannels was connected to a custom-made pressure-driven system.…”

“…RP-HPLC realized on a microscale allows one to study metabolism in an individual cell. 1 Chromatographic separations in extended-nano space (10 1 -10 2 nm scale), developed by our group, [2][3][4][5] can deal with sample volumes that range from attoliters to femtoliters with a high separation efficiency and a separation time of several seconds, and can be applied to single-cell analysis. Liquid chromatography in extended-nano space is based on "open tube" nanofluidics, wherein separation takes place in a single open tubular wallcoated nanochannel.…”

“…Our group has already realized chromatographic separations on microchips with extended-nano channel lengths of 1 mm 3,5 and 5 mm 4 for the normal-phase mode and 2 mm for the reversephase mode. 8 In the present work, we studied the influence of a lengthened and extended-nano channel on the efficiency of the reversed-phase separation of labeled amino acids.…”

Living Single Cell Analysis Platform Utilizing Microchannel, Single Cell Chamber, and Extended-nano Channel

“…The extended-nano chromatography device allowed highly efficient separation for aL volume samples. [14][15][16] In addition, fL immunochemical reaction space was constructed by patterning antibodies in the extended-nano channel to realize almost 100% capture of target protein molecules. 17 The volume of the extended-nano channel (aL-fL) is much smaller than the single cell volume (pL), and the extended-nano space will be promising for living single cell analysis by analyzing small volume of sample from the single cell in the extended-nano space.…”