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. 2022 Apr 20;298(6):101960. doi: 10.1016/j.jbc.2022.101960

Figure 4.

Figure 4

The same glycoform of RPTPζ carries CD33 and Siglec-8 ligands.A, equal aliquots of human cerebral cortex total protein extract from four donors (numbered) were resolved on replicate composite agarose–acrylamide gels and blotted to PVDF. One blot (upper panels) was double-label probed with CD33-Fc (red) and anti-RPTPζ (green) and a replicate blot (lower panels) with Siglec-8-Fc (red) and anti-RPTPζ (green). B, human cerebral cortex extract was size excluded (not shown) and subjected to affinity capture purification as for Figure 3. Equal aliquots of samples from affinity capture were resolved on replicate composite agarose–acrylamide gels, blotted to PVDF, and double-label probed with Siglec-8-Fc (red) and anti-RPTPζ (green) or with CD33-Fc (red) and anti-RPTPζ (green) as indicated. Lanes: (1) precapture; (2) precleared on IgG beads; (3) flow through (unbound) Siglec-8-Fc beads; (4–7) low salt washes; and (8–10) high salt elutions. The double-label gels carried custom molecular weight markers visible in the green images only. IgG, immunoglobulin G; PVDF, polyvinylidene fluoride; RPTPζ, receptor protein tyrosine phosphatase zeta; Siglec, sialic acid–binding immunoglobulin-type lectin.