PMID- 32973205 OWN - NLM STAT- MEDLINE DCOM- 20210528 LR - 20210924 IS - 2055-5008 (Electronic) IS - 2055-5008 (Linking) VI - 6 IP - 1 DP - 2020 Sep 24 TI - Clinically adaptable polymer enables simultaneous spatial analysis of colonic tissues and biofilms. PG - 33 LID - 10.1038/s41522-020-00143-x [doi] LID - 33 AB - Microbial influences on host cells depend upon the identities of the microbes, their spatial localization, and the responses they invoke on specific host cell populations. Multimodal analyses of both microbes and host cells in a spatially resolved fashion would enable studies into these complex interactions in native tissue environments, potentially in clinical specimens. While techniques to preserve each of the microbial and host cell compartments have been used to examine tissues and microbes separately, we endeavored to develop approaches to simultaneously analyze both compartments. Herein, we established an original method for mucus preservation using Poloxamer 407 (also known as Pluronic F-127), a thermoreversible polymer with mucus-adhesive characteristics. We demonstrate that this approach can preserve spatially-defined compartments of the mucus bi-layer in the colon and the bacterial communities within, compared with their marked absence when tissues were processed with traditional formalin-fixed paraffin-embedded (FFPE) pipelines. Additionally, antigens for antibody staining of host cells were preserved and signal intensity for 16S rRNA fluorescence in situ hybridization (FISH) was enhanced in poloxamer-fixed samples. This in turn enabled us to integrate multimodal analysis using a modified multiplex immunofluorescence (MxIF) protocol. Importantly, we have formulated Poloxamer 407 to polymerize and cross-link at room temperature for use in clinical workflows. These results suggest that the fixative formulation of Poloxamer 407 can be integrated into biospecimen collection pipelines for simultaneous analysis of microbes and host cells. FAU - Macedonia, Mary C AU - Macedonia MC AD - Division of Gastroenterology, Hepatology and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA. AD - Epithelial Biology Center, Vanderbilt University Medical Center, Nashville, TN, USA. FAU - Drewes, Julia L AU - Drewes JL AUID- ORCID: 0000-0002-5060-0624 AD - Division of Infectious Diseases, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA. FAU - Markham, Nicholas O AU - Markham NO AUID- ORCID: 0000-0002-1348-1267 AD - Division of Gastroenterology, Hepatology and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA. AD - Epithelial Biology Center, Vanderbilt University Medical Center, Nashville, TN, USA. FAU - Simmons, Alan J AU - Simmons AJ AD - Epithelial Biology Center, Vanderbilt University Medical Center, Nashville, TN, USA. AD - Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN, USA. FAU - Roland, Joseph T AU - Roland JT AD - Epithelial Biology Center, Vanderbilt University Medical Center, Nashville, TN, USA. AD - Department of Surgery, Vanderbilt University Medical Center, Nashville, TN, USA. FAU - Vega, Paige N AU - Vega PN AUID- ORCID: 0000-0003-2799-7587 AD - Epithelial Biology Center, Vanderbilt University Medical Center, Nashville, TN, USA. AD - Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN, USA. FAU - Scurrah, Cherie' R AU - Scurrah CR AD - Epithelial Biology Center, Vanderbilt University Medical Center, Nashville, TN, USA. AD - Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN, USA. FAU - Coffey, Robert J AU - Coffey RJ AD - Division of Gastroenterology, Hepatology and Nutrition, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA. AD - Epithelial Biology Center, Vanderbilt University Medical Center, Nashville, TN, USA. AD - Vanderbilt Ingram Cancer Center, Nashville, TN, USA. FAU - Shrubsole, Martha J AU - Shrubsole MJ AUID- ORCID: 0000-0002-5591-7575 AD - Vanderbilt Ingram Cancer Center, Nashville, TN, USA. AD - Division of Epidemiology, Vanderbilt Epidemiology Center, Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, USA. FAU - Sears, Cynthia L AU - Sears CL AUID- ORCID: 0000-0003-4059-1661 AD - Division of Infectious Diseases, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA. FAU - Lau, Ken S AU - Lau KS AUID- ORCID: 0000-0001-8438-0319 AD - Epithelial Biology Center, Vanderbilt University Medical Center, Nashville, TN, USA. ken.s.lau@vanderbilt.edu. AD - Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN, USA. ken.s.lau@vanderbilt.edu. AD - Vanderbilt Ingram Cancer Center, Nashville, TN, USA. ken.s.lau@vanderbilt.edu. LA - eng GR - R35 CA197570/CA/NCI NIH HHS/United States GR - P30 DK058404/DK/NIDDK NIH HHS/United States GR - UM1 CA183727/CA/NCI NIH HHS/United States GR - K99 CA230192/CA/NCI NIH HHS/United States GR - T32 AI007281/AI/NIAID NIH HHS/United States GR - U2C CA233291/CA/NCI NIH HHS/United States GR - P30 CA068485/CA/NCI NIH HHS/United States GR - P50 CA236733/CA/NCI NIH HHS/United States GR - R01 DK103831/DK/NIDDK NIH HHS/United States GR - T32 DK007673/DK/NIDDK NIH HHS/United States GR - T32 HD007502/HD/NICHD NIH HHS/United States PT - Journal Article PT - Research Support, N.I.H., Extramural DEP - 20200924 PL - United States TA - NPJ Biofilms Microbiomes JT - NPJ biofilms and microbiomes JID - 101666944 RN - 0 (RNA, Ribosomal, 16S) RN - 106392-12-5 (Poloxamer) SB - IM MH - Animals MH - Bacteria/classification/genetics/*isolation & purification MH - Biofilms/classification/*growth & development MH - Colon/*microbiology MH - Fluorescent Antibody Technique MH - Host-Pathogen Interactions MH - Humans MH - In Situ Hybridization, Fluorescence MH - Mice MH - Mucus MH - Paraffin Embedding MH - Poloxamer/*chemistry MH - RNA, Ribosomal, 16S/*genetics MH - Tissue Fixation PMC - PMC7518420 COIS- C.L.S. receives research funding from Bristol Myers Squibb and Janssen and a personal fee from Merck & Co. in 2019 for an advisory role. The authors declare that there are no other competing interests. EDAT- 2020/09/26 06:00 MHDA- 2021/05/29 06:00 PMCR- 2020/09/24 CRDT- 2020/09/25 05:56 PHST- 2020/03/31 00:00 [received] PHST- 2020/08/28 00:00 [accepted] PHST- 2020/09/25 05:56 [entrez] PHST- 2020/09/26 06:00 [pubmed] PHST- 2021/05/29 06:00 [medline] PHST- 2020/09/24 00:00 [pmc-release] AID - 10.1038/s41522-020-00143-x [pii] AID - 143 [pii] AID - 10.1038/s41522-020-00143-x [doi] PST - epublish SO - NPJ Biofilms Microbiomes. 2020 Sep 24;6(1):33. doi: 10.1038/s41522-020-00143-x.