Skeletal ciliopathies are a group of disorders caused by dysfunction of the cilium, a small signaling organelle present on nearly every vertebrate cell. This group of disorders is marked by genetic and clinical heterogeneity, which complicates accurate diagnosis. In this study, we developed a robust, standardized immunofluorescence approach to accurately diagnose a subset of these disorders. Hereto we determined and compared the cilium phenotype of healthy individuals to patients from three different ciliopathy subgroups, using skin-derived fibroblasts. The cilium phenotype assay consists of three parameters; (1) ciliogenesis, based on the presence or absence of cilium markers, (2) cilium length, measured by the combined signal of an axonemal and a cilium membrane marker, and (3) retrograde intraflagellar transport (IFT), quantified by the area of the ciliary tip. Analysis of the cilium phenotypic data yielded comparable and reproducible results and in addition, displayed identifiable clusters for healthy individuals and two ciliopathy subgroups, i.e. ATD and CED. Our results illustrate that standardized analysis of the cilium phenotype can be used to discriminate between ciliopathy subgroups. Therefore, we believe that standardization of functional assays analyzing cilium phenotypic data can provide additional proof for conclusive diagnosis of ciliopathies, which is essential for routine diagnostic care.

纤毛骨骼病是由纤毛功能障碍引起的一组疾病，纤毛是几乎每个脊椎动物细胞上都存在的小型信号细胞器。这组疾病的特点是遗传和临床异质性，这使准确诊断变得复杂。在这项研究中，我们开发了一种强大的标准化免疫荧光方法来准确诊断这些疾病的一个子集。至此，我们使用皮肤衍生的成纤维细胞确定并比较了健康个体的纤毛表型与来自三个不同纤毛病亚组的患者。纤毛表型测定由三个参数组成；(1) 纤毛发生，基于纤毛标记的存在与否，(2) 纤毛长度，通过轴丝和纤毛膜标记的组合信号测量，以及 (3) 逆行鞭毛内运输 (IFT)，由睫状体尖端的面积来量化。纤毛表型数据的分析产生了可比较和可重复的结果，此外，还显示了健康个体和两个纤毛病亚组（即 ATD 和 CED）的可识别集群。我们的结果表明纤毛表型的标准化分析可用于区分纤毛病亚组。因此，我们认为分析纤毛表型数据的功能测定的标准化可以为纤毛病的结论性诊断提供额外的证据，这对于常规诊断护理至关重要。我们的结果表明纤毛表型的标准化分析可用于区分纤毛病亚组。因此，我们认为分析纤毛表型数据的功能测定的标准化可以为纤毛病的结论性诊断提供额外的证据，这对于常规诊断护理至关重要。我们的结果表明纤毛表型的标准化分析可用于区分纤毛病亚组。因此，我们认为分析纤毛表型数据的功能测定的标准化可以为纤毛病的结论性诊断提供额外的证据，这对于常规诊断护理至关重要。