Improved DiOlistic labelling technique for neurons in situ: Detailed visualisation of dendritic spines and concurrent histochemical-detection in fixed tissue

IF 4.2 3区生物学 Q1 BIOCHEMICAL RESEARCH METHODS

Methods Pub Date : 2024-06-23 DOI:10.1016/j.ymeth.2024.06.009

Ryan J. Bevan , Gloria Cimaglia , James E. Morgan , Philip R. Taylor

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引用次数: 0

Abstract

DiOlistic labelling is a robust, unbiased ballistic method that utilises lipophilic dyes to morphologically label neurons. While its efficacy on freshly dissected tissue specimens is well-documented, applying DiOlistic labelling to stored, fixed brain tissue and its use in polychromatic multi-marker studies poses significant technical challenges. Here, we present an improved, step-by-step protocol for DiOlistic labelling of dendrites and dendritic spines in fixed mouse tissue. Our protocol encompasses the five key stages: Tissue Preparation, Dye Bullet Preparation, DiOlistic Labelling, Confocal Imaging, and Image Analysis. This method ensures reliable and consistent labelling of dendritic spines in fixed mouse tissue, combined with increased throughput of samples and multi-parameter staining and visualisation of tissue, thereby offering a valuable approach for neuroscientific research.

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改进的原位神经元 DiOlistic 标记技术：树突棘的详细可视化和固定组织中的同步组织化学检测。

DiOlistic 标记是一种稳健、无偏见的弹道方法，利用亲脂性染料对神经元进行形态标记。虽然这种方法在新鲜解剖组织标本上的效果有目共睹，但将 DiOlistic 标签应用于储存、固定的脑组织，并将其用于多色多标记研究，则面临着巨大的技术挑战。在此，我们提出了一种改进的、分步骤的方案，用于在固定的小鼠组织中对树突和树突棘进行 DiOlistic 标记。我们的方案包括五个关键阶段：组织制备、染料子弹制备、DiOlistic 标记、共聚焦成像和图像分析。这种方法可确保在固定的小鼠组织中对树突棘进行可靠、一致的标记，同时提高样本的处理量，并对组织进行多参数染色和可视化，从而为神经科学研究提供一种有价值的方法。

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来源期刊

Methods 生物-生化研究方法

CiteScore

9.80

自引率

2.10%

发文量

222

审稿时长

11.3 weeks

期刊介绍： Methods focuses on rapidly developing techniques in the experimental biological and medical sciences. Each topical issue, organized by a guest editor who is an expert in the area covered, consists solely of invited quality articles by specialist authors, many of them reviews. Issues are devoted to specific technical approaches with emphasis on clear detailed descriptions of protocols that allow them to be reproduced easily. The background information provided enables researchers to understand the principles underlying the methods; other helpful sections include comparisons of alternative methods giving the advantages and disadvantages of particular methods, guidance on avoiding potential pitfalls, and suggestions for troubleshooting.