Figure 6
Figure 6. Effect of APCE removal on conversion of Met-α2-AP to Asn-α2-AP with time. Plasma was drawn from a person of RR Met-α2AP genotype and divided into 3 aliquots. One aliquot was mixed with an APCE F19 mAb (F19) bound to chromatography beads and incubated at 4°C to remove APCE. The second aliquot was incubated at 4°C with a nonspecific rabbit α-goat Ab (RAG) bound to beads. The third aliquot (RR) received no treatment. After removal of beads, each sample was incubated at 29°C, and Asn-α2AP/Met-α2AP ratios determined at selected times. In addition, F19-bound beads and RAG-bound beads were boiled with SDS to remove antibody-bound protein. Samples were electrophoresed on 10% Bis-Tris SDS-PAGE gels and blotted to nitrocellulose. APCE (97 kDa) was identified by Western blotting using a goat Ab to its amino-terminal region and visualized with a chemiluminescent substrate.

Effect of APCE removal on conversion of Met-α2-AP to Asn-α2-AP with time. Plasma was drawn from a person of RR Met-α2AP genotype and divided into 3 aliquots. One aliquot was mixed with an APCE F19 mAb (F19) bound to chromatography beads and incubated at 4°C to remove APCE. The second aliquot was incubated at 4°C with a nonspecific rabbit α-goat Ab (RAG) bound to beads. The third aliquot (RR) received no treatment. After removal of beads, each sample was incubated at 29°C, and Asn-α2AP/Met-α2AP ratios determined at selected times. In addition, F19-bound beads and RAG-bound beads were boiled with SDS to remove antibody-bound protein. Samples were electrophoresed on 10% Bis-Tris SDS-PAGE gels and blotted to nitrocellulose. APCE (97 kDa) was identified by Western blotting using a goat Ab to its amino-terminal region and visualized with a chemiluminescent substrate.

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