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. 2019 Sep 16;9:13295. doi: 10.1038/s41598-019-49840-4

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Detection of NE activity in L. pneumophila-infected lung tissue. (A) Lysates prepared from uninfected or L. pneumophila-infected lungs were labeled with 1 µM PK105b and analyzed by in-gel fluorescence (n = 3–5). *Indicates high-molecular weight species of unknown identity. (B) Total NE expression in the samples in (A) was measured by immunoblotting with an NE-specific antibody. (C,D) Densitometry analysis of active NE in (A) and total mature NE in (B), respectively. (E) Ratios of active to total mature NE in the samples. (F) Labeling of NE by PK105b in infected lung tissue was confirmed by immunoprecipitation with an NE-specific antibody. (G) Competetion of PK105b labeling in an infected lung lysate with PK101. (H) PK105b labeling in neutrophils isolated from infected lung tissue, as analyzed by in-gel fluorescence (refer to Fig. S1 for sorting strategy). Error bars are shown as mean ± SEM.