Formalin-fixed paraffin-embedded lung sections were cut at 4 mm, tissue was probed with primary antibodies (listed below) and secondary antibodies with fluorescent conjugates (Invitrogen Alexa Fluor 488, 594, 647), and nuclear staining was performed with 4′,6-diamidino-2-phenylindole (DAPI) (Thermo Fisher, R37606). Immunostaining was performed using the following primary antibodies: mouse anti–Ac-α-Tub (Sigma, T6793), rabbit anti–Ac-α-Tub (Enzo Life Sciences, BML SA4592), rabbit anti-AKR1B10 (Sigma, HPA020280), rabbit anti-CEACAM5 (Abcam, ab131070), chicken anti-KRT5 (BioLegend, 905-901), rat anti-KRT8 (Developmental Studies Hybridoma Bank, University of Iowa; TROMA-I), mouse anti-MUC5AC (Abcam, ab3649), and rabbit anti-MUC5B (Sigma, HPA008246). Imaging of staining panels analyzed in collaboration with investigators at the UMCG (table S2) (e.g., AKR1B10/Ac-α-Tub/KRT8: Fig. 3C; AKR1B10/MUC5AC/KRT8: fig. S13; MUC5B/MUC5AC: Fig. 4B; MUC5B/MUC5AC/KRT5: fig. S14; CEACAM5/KRT8/MUC5AC: fig. S18) was performed using a Carl Zeiss LSM 710 NLO confocal microscope at ×63 objective magnification at the Boston University School of Medicine Multiphoton Microscope Core Facility. Imaging of staining panels analyzed in collaboration with investigators at the UCL (table S3) (e.g., CEACAM5/KRT8/MUC5AC: Fig. 5D; KRT8/MUC5AC/Ac-α-Tub: fig. S15) was performed using a Leica TCS Tandem confocal microscope at ×63 objective magnification.

Note: The content above has been extracted from a research article, so it may not display correctly.



Q&A
Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.



We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.