Finney Joel, Kuraoka Masayuki, Song Shengli, Watanabe Akiko, Liang Xiaoe, Liao Dongmei, Moody M Anthony, Walter Emmanuel B, Harrison Stephen C, Kelsoe Garnett
Laboratory of Molecular Medicine, Children's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Department of Integrative Immunobiology, Duke University, Durham, NC 27710, USA.
Vaccine. 2025 May 22;56:127157. doi: 10.1016/j.vaccine.2025.127157. Epub 2025 Apr 21.
The discovery of broadly protective antibodies to the influenza virus neuraminidase (NA) has raised interest in NA as a vaccine target. However, recombinant, solubilized tetrameric NA ectodomains are often challenging to express and isolate, hindering the study of anti-NA humoral responses. To address this obstacle, we established a panel of 22 non-adherent cell lines stably expressing native, historical N1, N2, N3, N9, and NB NAs anchored on the cell surface. The cell lines are barcoded with fluorescent proteins, enabling high-throughput, 16-plex analyses of antibody binding with commonly available flow cytometers. The cell lines were at least as efficient as a Luminex multiplex binding assay at identifying NA antibodies from a library of unselected clonal IgGs derived from human memory B cells. The cell lines were also useful for measuring the magnitude and breadth of the serum antibody response elicited by experimental infection of rhesus macaques with influenza virus. The membrane-anchored NAs are catalytically active and are compatible with established sialidase activity assays. NA-expressing K530 cell lines therefore represent a useful tool for studying NA immunity and evaluating influenza vaccine efficacy.
对流感病毒神经氨酸酶（NA）具有广泛保护作用的抗体的发现，引发了人们对将NA作为疫苗靶点的兴趣。然而，重组、可溶解的四聚体NA胞外结构域的表达和分离往往具有挑战性，这阻碍了对抗NA体液免疫反应的研究。为了解决这一障碍，我们建立了一组22个非贴壁细胞系，它们能稳定表达锚定在细胞表面的天然、具有历史代表性的N1、N2、N3、N9和NB型NA。这些细胞系用荧光蛋白进行了条形码标记，可使用常用的流式细胞仪对抗体结合进行高通量、16重分析。在从源自人类记忆B细胞的未选择克隆IgG文库中鉴定NA抗体方面，这些细胞系至少与Luminex多重结合测定法一样有效。这些细胞系还可用于测量恒河猴经流感病毒实验感染后引发的血清抗体反应的强度和广度。膜锚定的NA具有催化活性，并且与已建立的唾液酸酶活性测定法兼容。因此，表达NA的K530细胞系是研究NA免疫和评估流感疫苗效力的有用工具。
