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. 2016 Apr 4;11(4):e0152904. doi: 10.1371/journal.pone.0152904

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© 2016 Lai et al

This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

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Fig 1 — (A) Defatted human milk was loaded onto a CM-Sepharose column and the unbound fraction (UB) was collected. The milk (lanes 1) and the unbound fraction (lanes 2) were analyzed by immunoblot for HAI-1, HAI-2, and matriptase (MTP), as indicated. (B) Flow-chart summarizing the purification scheme for the HAI-1 and HAI-2 complexes in human milk. (C) Fractions 12 through 34 eluted from the CM-Sepharose chromatography were analyzed by Western blot for HAI-2. (D) The pooled HAI-2 containing fractions eluted from the CM-Sepharose were subjected to immunoaffinity chromatography on a HAI-1 mAb M19-Sepharose column followed by a HAI-2 mAb DC16-Sepharos column. Fractions 1 through 4 eluted from the mAb DC16-Sepharose were analyzed by Western blot for HAI-2 species (left panel). The pooled HAI-2 containing fractions were analyzed by immunoblot for matriptase species under non-boiled (lane 1) and boiled (lane 2) conditions (right panel).