Female C57BL/6 mice were purchased from the Centre for Animal Resources, Singapore and housed in specific pathogen-free BSL2 facilities at the National University of Singapore (NUS). Mice were anesthetized with ketamine ( body weight) and infected with a sub-lethal dose (100 PFU/mouse) of the influenza virus A/Puerto Rico/8/34 (H1N1) by intratracheal instillation. Five mice were sacrificed at each time point: before infection (control) and 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, and 28 days post infection (dpi). Lung tissues were collected. The left lobes were fixed in 10% neutral buffered formalin solution (Sigma-Aldrich, HT501128-4L) overnight and processed with Tissue Processor (Leica, TP1020, Shanghai, China) and embedded in paraffin blocks. For each lobe, 15 transverse sections were cut with microtome (Leica, RM2165, Nussloch, Germany) from the middle part of the lobe and with 50 μm in between sections. Sections were mounted on polylysine-coated slides (Thermal Fisher Scientific, J2800AMNZ). Ten sections were used for H&E and five sections for immunofluorescent staining. The right lobes were used for lavage, measuring virus titer and other analysis. All animal protocols were approved by the Institutional Animal Care and Use Committee at NUS and Massachusetts Institute of Technology.