Appeal No. 2005-1344 Page 15
Application No. 08/468,610
the opposite polarity from the net electrostatic charge on the target protein or
peptide at the pH of the desorption.
In the context of haemoglobins the opposite is true. Boardman refers to
the illustration in Figure 1a and states (page 210, column 1, first full paragraph),
as is show in Fig. 1a, at pH 5 the carboxylic groups of the resin are
almost wholly undissociated, and adsorption of the haemoglobins is
complete and irreversible. The elution volume – pH data of the
haemoglobins may thus be represented by a family of steep curves
falling rapidly from very high values to zero in the narrow range pH
5-6, the curves for the more acidic proteins being displaced to the
left of those for proteins with higher isoelectric points.”
To clarify, according to Boardman (page 210, column 1, first full paragraph), in
the citrate buffer used for the chromatographic experiments, the isoelectric point
of bovine and sheep foetal carboxyhaemoglobin lies between 4 and 5. Since a
protein will carry a net negative charge at a pH above its pI, at a pH of 6, which is
above the pI for bovine and sheep foetal carboxyhaemoglobin, each of the
carboxyhaemoglobin proteins will have a net negative charge. See, Boardman,
page 210, column 1, first full paragraph.
We encourage appellants to clarify this issue.
II. Becker Declaration:
We recognize appellants’ arguments relating to the Becker declaration
and the “Rohm and Haas product literature” discussed therein. Brief, pages 7-8.
However, upon review of the Becker declaration, we note that declarant states
that the Amberlite IRC 50® resin used by Broadman “becomes fully protonated
(neutralized)[, completely uncharged,] only at a pH of between 2.5 and 4.0,
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