Separations on Chips

Successful liquid-phase separation on-chip was first carried out in CE, because EOF pumping can be easily achieved in the microscale. For instance, six fluorescein-labeled amino acids are separated by CE on a Pyrex glass chip (10-pm-deep and 30-pm-wide channel) (see Figure 6.5). Separation was achieved in a very short time of about 15 s [324]. Similar CE separation of calcein and fluorescein were also reported [582]. Separation of a binary mixture of rhodamine B and dichlorofluorescein was even achieved in only 0.8 ms using a short separation length of 200 pm [604]. 

In competitive homogeneous immunoassay, separation and quantitation of free and bound labeled antigen (cortisol) were carried out in a fused silica chip. Since the antibody-antigen complex was not detected, an internal standard (fluorescein) was added to aid quantitation. In addition, since most of the total cortisol was bound in the serum, a releasing agent, 8-anilino-l-naphthalenesulfonic acid (ANS), should be added [1006]. In other reports, competitive immunoassay for BSA was demonstrated after performing a CE separation on-chip [105,1005]. 

Transverse IEF was also conducted in a pressure-driven flow for BSA and soybean lectin separation on-chip [1040], Here, Pd electrodes were used (in preference to Au) because of the non-gassing character of Pd. In addition, the protein sample was sandwiched between two buffer streams and was prevented from direct contact with the channel wall (and hence the electrode), a process akin to hydrodynamic focusing [1040],... 

In carrying out CGE separation on chip, why is the channel usually coated (2 marks)... 

Harrison, D.J., Andersson, P.E., Li, P.C.H., Chiem, N., Tang, T., Smith, R., Szarka, R., Tran, T., Integration of biochemical and cellular reactions with separation on-chip. 19th International Symposium on Capillary Chromatography and Electrophoresis, May 18-22, 1997. 

FIGURE 45.11 Three-dimensional multicolor data collected with an AOTF. A mixture of three dyes (FL fluorescein, R6G rhodamine 6G, and ROX carboxy-X-rhodainine) was injected and separated on chip while scanning the AOTF through 19 wavelengths. (Adapted from KarUnsey, J.M. and Landers, J.P., Analytical... 

Another reason for the limited interest in HPLC-like separations on chip appears to be the complexity of plumbing since an external pump typical of standard HPLC instrumentation has to be included. The role of the chip then degrades to a capillary-like column and the advantages attributed to microfluidic devices vanish. In order to avoid attachment of the chip to a bulky mechanical pump, Penrose et al. used centripetal force to propel the mobile phase through a bed of particles packed in a channel. Since no detector was added to the system, the readout was a scan of the bed after visualizing the bands in ultraviolet (UV) light. 

Space for the monolithic column. The wall modification and preparation of the polyfethylhexyl methacrylate-co-ethylene dimethacrylate) monolith was identical with that shown earlier. The ultimate test of the performance of the system included separation of a digest of three proteins— BSA, myoglobin, and cytochrome c. The sequence coverage obtained using the microchip with two monolithic columns run in parallel is shown in Table 47.1 and compared with MALDI-MS analysis data obtained for nonfractionated sample. Clearly, the separation on chip containing monolith affords significantly better results. 

Pellejero I, Garcia S, Urbiztondo MA, Sese J, Pina MP, Santamaria J. silicahte-1 micromembranes for COj/Hj separation on chip Microfabrication process and device performance. Book of Abstracts. 12th International Cortference on Inorganic Membranes, Enschede, the Netherlands, July 9-July 13, 2012. 

Microfabricated instrumentation has become an active area of research over the past 10 years and it promises to play a dominant role in the future of chemical analysis. Although microfabricated devices such as sensors, electrode arrays, and even gas chromatography have been available for some time, the current revolution has been brought about by the implementation of microfluidics and integration of sample preparation and separation on chips. Both CE and LC systems have... 

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