Development of Negative Controls for Fc-C-Type Lectin Receptor Probes.
Development of Negative Controls for Fc-C-Type Lectin Receptor Probes.
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Fc-C 型凝集素受体探针阴性对照的开发。
DOI：
10.1128/spectrum.01135-23
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发表时间：
2023-06-15
影响因子：
3.7
通讯作者：
Brown, Gordon D. D.
中科院分区：
文献类型：
--
作者：
Hatinguais, Remi;Kay, Madalaine;Salazar, Fabian;Conn, Daniel P. P.;Williams, David L. L.;Cook, Peter C. C.;Willment, Janet A. A.;Brown, Gordon D. D.
关键词：
Fc-C 型凝集素受体 (Fc-CTLR) 探针是可溶性嵌合蛋白，由 CTLR 的胞外结构域与人 IgG 恒定部分 (Fc) 融合构成。这些探针是研究 CTLR 与其配体相互作用的有用工具，其应用类似于抗体，通常与广泛使用的针对 Fc 片段的荧光抗体（抗 hFc）结合使用。特别是，Fc-Dectin-1 已被广泛用于研究病原真菌表面 β-葡聚糖的可及性。然而，Fc-CTLR 没有通用的阴性对照，因此很难区分特异性与非特异性结合。我们在此描述了 Fc-CTLR 的 2 个阴性对照：仅由 Fc 部分组成的 Fc 对照，以及预计无法结合 β-葡聚糖的 Fc-Dectin-1 突变体。使用这些新探针，我们发现虽然 Fc-CTLR 实际上没有表现出与白色念珠菌酵母的非特异性结合，但烟曲霉静息孢子以非特异性方式强烈结合 Fc-CTLR。尽管如此，使用我们在此描述的对照，我们能够证明烟曲霉孢子暴露出少量的β-葡聚糖。我们的数据强调了涉及 Fc-CTLR 探针的实验中适当的阴性对照的必要性。重要性虽然 Fc-CTLR 探针是研究 CTLR 与配体相互作用的有用工具，但由于在涉及真菌和潜在其他病原体的检测中缺乏适当的阴性对照，其使用受到限制。我们开发并表征了用于 Fc-CTLR 测定的 2 种阴性对照：Fc 对照和 Fc-Dectin-1 突变体。在这份手稿中，我们描述了这些阴性对照与酵母聚糖（一种含有 β-葡聚糖的颗粒）和 2 种人类致病真菌（白色念珠菌酵母和烟曲霉分生孢子）的使用。我们证明烟曲霉分生孢子非特异性结合 Fc-CTLR 探针，证明在此类测定中需要适当的阴性对照。
Fc-C-type lectin receptor (Fc-CTLRs) probes are soluble chimeric proteins constituted of the extracellular domain of a CTLR fused with the constant fraction (Fc) of the human IgG. These probes are useful tools to study the interaction of CTLRs with their ligands, with applications similar to those of antibodies, often in combination with widely available fluorescent antibodies targeting the Fc fragment (anti-hFc). In particular, Fc-Dectin-1 has been extensively used to study the accessibility of β-glucans at the surface of pathogenic fungi. However, there is no universal negative control for Fc-CTLRs, making the distinction of specific versus nonspecific binding difficult. We describe here 2 negative controls for Fc-CTLRs: a Fc-control constituting of only the Fc portion, and a Fc-Dectin-1 mutant predicted to be unable to bind β-glucans. Using these new probes, we found that while Fc-CTLRs exhibit virtually no nonspecific binding to Candida albicans yeasts, Aspergillus fumigatus resting spores strongly bind Fc-CTLRs in a nonspecific manner. Nevertheless, using the controls we describe here, we were able to demonstrate that A. fumigatus spores expose a low amount of β-glucan. Our data highlight the necessity of appropriate negative controls for experiments involving Fc-CTLRs probes. IMPORTANCE While Fc-CTLRs probes are useful tools to study the interaction of CTLRs with ligands, their use is limited by the lack of appropriate negative controls in assays involving fungi and potentially other pathogens. We have developed and characterized 2 negative controls for Fc-CTLRs assays: Fc-control and a Fc-Dectin-1 mutant. In this manuscript, we characterize the use of these negative controls with zymosan, a β-glucan containing particle, and 2 human pathogenic fungi, Candida albicans yeasts and Aspergillus fumigatus conidia. We show that A. fumigatus conidia nonspecifically bind Fc-CTLRs probes, demonstrating the need for appropriate negative controls in such assays.
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Dectin-1介导β-葡聚糖的生物学作用。
DOI：
10.1084/jem.20021890
发表时间：
2003-05-05
期刊：
The Journal of experimental medicine
影响因子：
--
作者：
Brown GD;Herre J;Williams DL;Willment JA;Marshall AS;Gordon S
通讯作者：
Gordon S
影响因子：
20.3
作者：
Herre, J;Marshall, ASJ;Brown, GD
通讯作者：
Brown, GD
影响因子：
64.8
作者：
Aimanianda, Vishukumar;Bayry, Jagadeesh;Latge, Jean-Paul
通讯作者：
Latge, Jean-Paul
DOI：
10.3390/jof6030151
发表时间：
2020-08-26
期刊：
Journal of fungi (Basel, Switzerland)
影响因子：
--
作者：
Valsecchi I;Stephen-Victor E;Wong SSW;Karnam A;Sunde M;Guijarro JI;Rodríguez de Francisco B;Krüger T;Kniemeyer O;Brown GD;Willment JA;Latgé JP;Brakhage AA;Bayry J;Aimanianda V
通讯作者：
Aimanianda V
影响因子：
3.4
作者：
Luther, Kathrin;Torosantucci, Antonella;Ebel, Frank
通讯作者：
Ebel, Frank