Specific in situ immuno-imaging of pulmonary-resident memory lymphocytes in human lungs.
Specific in situ immuno-imaging of pulmonary-resident memory lymphocytes in human lungs.
复制标题
DOI：
10.3389/fimmu.2023.1100161
复制
发表时间：
2023
影响因子：
7.3
通讯作者：
Pavot, Vincent
中科院分区：
文献类型：
--
作者：
Humphries, Duncan C.;O'Connor, Richard A.;Stewart, Hazel L.;Quinn, Tom M.;Gaughan, Erin E.;Mills, Beth;Williams, Gareth O. S.;Stone, James M.;Finlayson, Keith;Chabaud-Riou, Martine;Boudet, Florence;Dhaliwal, Kevin;Pavot, Vincent
关键词：
肺驻留记忆 T 细胞 (TRM) 和 B 细胞 (BRM) 协调保护性免疫，以抵抗呼吸道病原体的再感染。开发这些人群的原位检测方法将有利于研究和临床环境。为了满足这一需求，我们开发了一种新型原位免疫标记方法，与临床就绪的基于光纤的光学内窥镜检查 (OEM) 相结合，以检测进行离体肺通气 (EVLV) 的人肺中淋巴细胞组织原位驻留的典型标记。最初，使用 CD69 和 CD103/CD20 荧光抗体对来自人肺消化物的细胞（使用流式细胞术确认含有 TRM/BRM 群体）进行染色，并使用 KronoScan 进行体外成像，证明其检测抗体标记细胞的能力。接下来，我们将这些预先标记的细胞注入接受 EVLV 的人肺中，并确认它们仍然可以使用荧光强度和针对背景肺结构的寿命成像来可视化。最后，我们将荧光 CD69 和 CD103/CD20 抗体直接滴注到肺部，并能够在直接向肺泡内输送微剂量荧光标记抗体的几秒钟内进行原位标记后检测 TRM/BRM。 原位、免洗、肺泡内 OEM 成像免疫标记是一种新颖的方法，有可能扩大 EVLV 和临床前模型的实验效用。
Pulmonary-resident memory T cells (TRM) and B cells (BRM) orchestrate protective immunity to reinfection with respiratory pathogens. Developing methods for the in situ detection of these populations would benefit both research and clinical settings. To address this need, we developed a novel in situ immunolabelling approach combined with clinic-ready fibre-based optical endomicroscopy (OEM) to detect canonical markers of lymphocyte tissue residency in situ in human lungs undergoing ex vivo lung ventilation (EVLV). Initially, cells from human lung digests (confirmed to contain TRM/BRM populations using flow cytometry) were stained with CD69 and CD103/CD20 fluorescent antibodies and imaged in vitro using KronoScan, demonstrating it’s ability to detect antibody labelled cells. We next instilled these pre-labelled cells into human lungs undergoing EVLV and confirmed they could still be visualised using both fluorescence intensity and lifetime imaging against background lung architecture. Finally, we instilled fluorescent CD69 and CD103/CD20 antibodies directly into the lung and were able to detect TRM/BRM following in situ labelling within seconds of direct intra-alveolar delivery of microdoses of fluorescently labelled antibodies. In situ, no wash, immunolabelling with intra-alveolar OEM imaging is a novel methodology with the potential to expand the experimental utility of EVLV and pre-clinical models.
登录
查看更多内容
影响因子：
8
作者：
McMaster SR;Wein AN;Dunbar PR;Hayward SL;Cartwright EK;Denning TL;Kohlmeier JE
通讯作者：
Kohlmeier JE
影响因子：
7.3
作者：
Humphries DC;O'Connor RA;Larocque D;Chabaud-Riou M;Dhaliwal K;Pavot V
通讯作者：
Pavot V
影响因子：
10
作者：
Dorward DA;Lucas CD;Doherty MK;Chapman GB;Scholefield EJ;Conway Morris A;Felton JM;Kipari T;Humphries DC;Robb CT;Simpson AJ;Whitfield PD;Haslett C;Dhaliwal K;Rossi AG
通讯作者：
Rossi AG
影响因子：
15.9
作者：
Barker, Kimberly A.;Etesami, Neelou S.;Mizgerd, Joseph P.
通讯作者：
Mizgerd, Joseph P.
DOI：
10.1007/s00259-020-05021-4
发表时间：
2020-09-11
影响因子：
9.1
作者：
Mills, Bethany;Megia-Fernandez, Alicia;Stone, James M.
通讯作者：
Stone, James M.