Use of mTRAQ derivatization reagents on tissues for imaging neurotransmitters by MALDI imaging mass spectrometry: the triple spray method

Abstract

During drug development, matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry is used for visually elucidating the distribution of substances such as biomarkers, candidate compounds, and metabolites in the tissues. However, it is difficult to make relative comparisons between tissue sections and there are still many challenges. Here, we report a new method of “triple spray” for the comparison of analyte distribution in multiple tissue slices. This method targets amino acids and amines, and it incorporates the application of the internal standard in the on-tissue derivatization step. With further development, it has the potential to alleviate problems caused by the matrix effect. Initially, we measured three serial sections of rat brain to verify the efficacy of this method. In the hypothalamus, where gamma-aminobutyric acid (GABA) is known to be present in high concentration, the GABA levels of the three serial section showed little variation (CV = 1.62%). Subsequently, we compared the GABA level in the brain between stroke-prone spontaneous hypertensive rats (SHRSP) and Wistar-Kyoto (WKY) rats with three individuals each. It showed significant differences between these models at the pre-selected region of interest (p < 0.05). Our results show that the triple spray allows for relative comparison among multiple tissue slices with high reproducibility. Graphical abstract

Advances in MALDI Mass Spectrometry Imaging Single Cell and Tissues

“…The on-tissue chemical derivatization (OTCD), which is spray chemical derivatization first followed by matrix application (Harkin et al, 2021), has been used to improve ionization efficiency to effectively detect analytes directly from both fresh frozen tissue and FFPE tissue. Now OTCD has developed rapidly and used to detect many biological molecules, such as N-glycan (Nishikaze et al, 2013;Holst et al, 2016;Zhang et al, 2020;Saito et al, 2021), drugs (Barre et al, 2016, amines (Chacon et al, 2011;Manier et al, 2014), fatty acids (Wu et al, 2016;Wang et al, 2019;Iwama et al, 2021), amino acids (Toue et al, 2014;Esteve et al, 2016;Guo et al, 2020), poisons (Beasley et al, 2016), plant hormones (Enomoto et al, 2018), peptides (Franck et al, 2010), steroids (Guo et al, 2020;Angelini et al, 2021;Song et al, 2021), neurotransmitters (Ito and Hiramoto, 2019;Palanisamy et al, 2020;Shariatgorji et al, 2020), and so on. It is worth noting that the coating method of derivatization solution and matrix is also important because it affects not only the efficiency of OTCD but also the quality of imaging results.…”

On‐tissue chemical derivatization in mass spectrometry imaging

“…Understanding of the distribution of neurotransmitters and their metabolites would divulge insightful information into behavior, sleep, cognitive function, and diseases such as Parkinson's and Alzheimer's. The largest portion of examples mentioned throughout this review has made possible the localization and tracing of previously undetectable neurotransmitters in animal tissue (Davison et al, 2019;Esteve et al, 2016;Ito & Hiramoto, 2019;Manier et al, 2014;Shariatgorji et al, 2014Shariatgorji et al, , 2015Shariatgorji et al, , 2016Shariatgorji et al, , 2019Shariatgorji et al, , 2020Sugiyama et al, 2019). OTCD of aminoneurotransmitters with TAHS and CA reagents to target the amine group have been successful in localizing neurotransmitters in mice brain tissue (Esteve et al, 2016).…”

Chemical derivatization: recent advances in mass spectrometry imaging

“…取代衍生 化反应主要应用于生物小分子的研究分析中, 有利于 增加小分子的质子亲和性和电离效率. 常见的取代反 应 衍 生 化 试 剂 有 1-溴 丁 烷 [129] 、 mTRAQ(Mass differential tags for relative and absolute quantification) [136] 、 二 甲 基 嘧 啶 鸟 氨 酸 (dimethyl pyrimidinyl ornithine, DiPyrO)标记试剂 [134] 、d0-/d3-6-N-甲 基 罗 丹 明 6G-N-羟 基 琥 珀 酰 加 成 甲 酸 酯 (d0/d3-6-N-methylrhodamine 6G-N-hydroxysuccinimidyl formate, d0/d3-MRSF) [70] 、 9-氟 烯 基 甲 氧 基 羰 基 (9-fluorenylmethoxycarbonyl chloride, FMOC-Cl) [86] 、乙 氧 亚 甲 基 丙 二 酸 二 苄 酯 (dibenzyl ethoxymethylene malonate, DBEMM) [130] 、 乙 氧 亚 甲 基 丙 二 酸 苄 乙 酯 (benzyl ethyl ethoxymethylene malonate, EBEMM) [130] 及稳定同位素标记氨氧基串联质量标签(stable isotope labeled aminoxy tandem mass tags, SIL-aminoxyTMTs) [135] 等. 在 MS 分析中, 取代衍生化 试剂主要有三种功能.…”

“…在 MALDI-MSI 分析表征中, 取代衍生化在生物 分子原位定量研究中具有重要作用, 该衍生化技术 常用于对含有氨基、 羟基等的小分子物质的组织原位 分 析 . 如 , Ito 等 人 [136]…”