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首页> 外文会议>Conference on Optical Diagnostics of Living Cells Ⅴ, Jan 23-25, 2002, San Jose, USA >Multicolor immunophenotyping of tissue sections by Laser Scanning Cytometry (LSC)
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Multicolor immunophenotyping of tissue sections by Laser Scanning Cytometry (LSC)

机译：通过激光扫描细胞仪（LSC）对组织切片进行多色免疫分型

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摘要

In lymphatic organs the quantitative analysis of the spatial distribution of leukocytes would give relevant information about alterations during diseases (leukemia, HIV, AIDS) and their therapeutic regimen. Analysis of them in solid tissues is difficult to perform but would yield important data in a variety of clinical and experimental settings. We have developed an automated analysis method for LSC suitable for archived or fresh biopsy material of human lymph nodes and tonsils. Sections are stained with PI for DNA and up to three antigens using direct or indirect immunofluo-rescence staining. Measurement is triggered on DNA-fluorescence (Argon Laser). Due to the heterogeneity in cell density measurements are repeatedly performed at different threshold levels (low threshold: regions of low cellular density, germinal centers; high threshold: dense regions, mantle zone). Data are acquired by single- (Ar) or dual-laser excitation (Ar-HeNe) in order to determine data from single- (FITC), up to triple-staining (FITC/PE-Cy5/APC). Percentage and cellular density of cell-subsets is quantified in different structural regions of the specimen. Comparison with manual analysis of identical specimens showed very good correlation. With LSC a semi-automated operator-independent and immunophenotyping of lymphatic tissues with simultaneously up to four antibodies is possible. This technique should yield new insight into processes during diseases and should help to quantify the success of therapeutic interventions.
机译：在淋巴器官中，对白细胞空间分布的定量分析将提供有关疾病（白血病，HIV，AIDS）及其治疗方案变化的相关信息。在实体组织中对它们进行分析很困难，但会在各种临床和实验环境中产生重要数据。我们为LSC开发了一种自动分析方法，适用于人类淋巴结和扁桃体的存档或新鲜活检材料。使用直接或间接免疫荧光染色法对切片进行DNA和最多三种抗原的PI染色。 DNA荧光（Argon激光）触发测量。由于细胞密度的异质性，在不同的阈值水平（低阈值：细胞密度低的区域，生发中心；高阈值：致密区域，地幔区域）重复进行测量。通过单次（Ar）或双激光激发（Ar-HeNe）采集数据，以便确定从单次（FITC）到三重染色（FITC / PE-Cy5 / APC）的数据。在样品的不同结构区域中定量细胞亚群的百分比和细胞密度。与相同样本的人工分析比较显示出很好的相关性。使用LSC，淋巴组织的半自动独立于操作者的免疫分型和同时最多四种抗体的免疫表型成为可能。这种技术应该对疾病过程产生新的见解，并应有助于量化治疗干预的成功。

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