Gels Mimicking Antibodies in Their Selective Recognition of Proteins and Its Potential Use for Protein Crystallization - Molecules: Nucleation, Aggregation and Crystallization - page 21

was 1.2 mg/mL). The column was emptied and the gel particles were
pressed three times through a 100-mesh net and mixed with a monomer
solution of the composition T20, C3 in the volume ratio 1:2. The same
catalyst was used as in the above experiment with the “ribonuclease col-
umn” although the volumes of the ammonium per-sulfate and TEMED
solutions were increased 2.5-fold. About 40 minutes following poly-
merization the gel was disintegrated by forcing it once through a 100-
mesh net and then packed into a Pasteur pipette. The column was washed
with 6% SDS in 10% acetic acid and equilibrated with 20 mM sodium
phosphate, pH 7.0. The sample consisted of 20 L of a solution of hemo-

globin (10 mg/mL), cytochrome c (2 mg/mL) and lysozyme (2 mg/mL) in
1.0 mM sodium phosphate, pH 7.0. The flow resistance of this bed was
considerably lower than that obtained with the original T6 C5 gel. The
selectivity test (Fig. 6), performed as described for the experiment pre-
sented in Fig. 4, demonstrates that it is possible to combine high speci-
ficity with high flow rate on beds with small gel particles which have the
advantage of permitting shorter run times and less zone broadening.

µ

Results and Discussion

The selectivity of the artificial gel antibodies

In an article published more than a decade ago, we described a method
for the preparation of gels with the property to permit selective recog-
nition of cytochrome c (Liao et al ., 1996). From the chromatograms in
Figs. 1, 2 and 4, one can conclude that the same method can be
employed to adsorb specifically ribonuclease from horse, human
growth hormone and myoglobin from horse. From the many experi-
ments we have done since then, we can conclude that, with great prob-
ability, similar beds can be designed for all proteins. It is interesting to note
that one can synthesize a gel which can recognize several proteins (Fig. 3).
Such a gel can be of interest for rapid “negative purification” of a certain
protein by suspending the gel particles (selective for the non-desirable pro-
teins) in the protein sample and decanting; see section on “The design of
a chromatographic bed for removal of impurities”, (page 19).

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