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A capillary isoelectric focusing-whole column imaging detection (CIEF-WCID) method was used to determine the isoelectric point (pI) of norovirus virus-like particles (VLPs). The VLPs were produced from noroviruses that represented the two genogroups, genogroup I (Funabashi, Seto, and Norwalk) and genogroup II (Hawaii, Kashiwa, and Narita). Using the imaged CIEF-WCID detection technique, separation was accomplished using a short (4-5 cm) internally coated capillary (100-/[micro]m diameter) and a whole-column optical absorption imaging detector operated at 280 nm. CIEF-WCID experiments showed the similarity of the pI values of VLPs from genogroups I and II, with pI values of 5.9, 5.9, 6.0, and 6.0 for Funabashi, Norwalk, Seto, and Hawaii. The two other VLPs displayed pI values of 5.5 (Kashiwa) and 6.9 (Narita). The VLP peaks were shown to be reproducibility resolved. CIEF-WCID shows great promise for norovirus detection in public health, clinical, and food safety applications, as CIEF-WCID overcomes several limitations of the currently used genetic and immunological methods.

The capillary isoelectric focusing (CIEF) cartridge uses a 4 cm long, 75 pm i.d., 150 [[micro]meter] o.d. capillary. The two ends of the capillary are connected to two pieces of 3 mm long, 150 [[micro]meter] i.d., 170 [[micro]meter] o.d. dialysis hollow fibers. These pieces are inserted into electrolyte reservoirs attached to the two ends of the capillary. The fibers isolate the sample solution inside the capillary from the electrolytes in the reservoirs but allow small ions to pass freely, so that the isoelectric focusing process can take place normally inside the capillary. With this cartridge, samples can be injected conveniently and quickly from one end of the capillary. After the sample injection, the separation voltage is turned on and the focusing process begins. When the cartridge is combined with a real-time, on-line absorption imaging detector, the analysis time of the instrument for a sample is only 4 min, including sample injection. The analysis time can be further reduced to about 30 s by using a 1 cm long capillary as the separation column. To preserve the fast speed of this CIEF instrument, a fast, on-line desalting and carrier ampholyte mixing device using a dialysis hollow fiber was also designed. Protein samples 1-10 [[micro]liter] in volume can be desalted and mixed with carrier ampholytes and injected into the separation column in an on-line fashion in 2 min. The new CIEF-desalting device was also developed with multiple capillaries, which increases the sample throughput of the CIEF method. A three capillary cartridge was constructed and tested. Total analysis time for three samples in the three capillaries is only 5 min.

A practical, inexpensive method of analyzing proteins combines Schlieren optics with high-performance capillary isoelectric focusing (IEF). The high- concentration gradient created in a 12 cm by 20 micron (i.d.) capillary allows sensitive detection by a He-Ne laser/light beam position sensor. The new IEF/Schlieren detector tandem matches the sensitivity of a UV absorbance detector even as it permits a smaller detection volume. Using the tandem, femtomole-level proteins are separated and detected directly without prior derivatization.

A practical, inexpensive method of analyzing proteins combines Schlieren optics with high-performance capillary isoelectric focusing (IEF). The high- concentration gradient created in a 12 cm by 20 micron (i.d.) capillary allows sensitive detection by a He-Ne laser/light beam position sensor. The new IEF/Schlieren detector tandem matches the sensitivity of a UV absorbance detector even as it permits a smaller detection volume. Using the tandem, femtomole-level proteins are separated and detected directly without prior derivatization.