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. 2019 Nov 20;10:2621. doi: 10.3389/fimmu.2019.02621

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Copyright © 2019 Pizzagalli, Latino, Pulfer, Palomino-Segura, Virgilio, Farsakoglu, Krause and Gonzalez.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Neutrophils exhibit different actions in the early response to vaccination. (A) Graphic representation (up) and 2P-IVM micrographs showing different neutrophil actions in response to influenza vaccination: flowing “F” neutrophils moving inside blood vessels, with long and straight tracks following the vessel structure. Arrested neutrophils “A” exhibit confined tracks and short displacement. Patrolling neutrophils “P” have long, non-straight tracks associated with extensive tissue monitoring. Directed neutrophils “D” exhibit straight tracks with lower migration speed compared to flowing neutrophils. Swarming neutrophils (S) form cell aggregates with high density and large volume. (B) Gating strategies used to define and identify the actions according to cell speed, displacement, straightness, and arrest coefficient on track fragments of fixed length (500 s). (C) Classification of track fragments at different time intervals (arrested: black, patrolling: red, directed: green, flowing: blue, unspecified: gray). (D) Time-course plot showing the percentage of cells performing each of the actions over time, according to our gating strategy.