Abstract
Lectin microarray (LMA) is a high-sensitive glycan analysis technology used to obtain global glycomic profiles of both N- and O-glycans attached not only to purified glycoproteins but also to crude glycoprotein samples. Through additional use of laser microdissection (LMD) for tissue collection, we developed an LMA-based glycomic profiling technique for a specific type of cells in a tiny area of formalin-fixed paraffin-embedded (FFPE) tissue sections. This LMD–LMA method makes it possible to obtain reproducible tissue glycomic profiles that can be compared with each other, using a unified protocol for all procedures, including FFPE tissue preparation, tissue staining, protein extraction and labeling, and LMA analysis. Here, we describe the standardized LMD–LMA procedure for a “tissue glycome mapping” approach, which facilitates an in-depth understanding of region- and tissue-specific protein glycosylation. We also describe potential applications of the spatial tissue glycomic profiles, including histochemical analysis for evaluating distribution of lectin ligands and a fluorescence LMD–LMA method for cell type-selective glycomic profiling using a cell type-specific probe, composed of a lectin and an antibody. The protocols presented here will accelerate the effective utilization of FFPE tissue specimens by providing tissue glycome maps for the discovery of the biological roles and disease-related alterations of protein glycosylation.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Moremen KW, Tiemeyer M, Nairn AV (2012) Vertebrate protein glycosylation: diversity, synthesis and function. Nat Rev Mol Cell Biol 13:448–462
Gaunitz S, Nagy G, Pohl NLB et al (2017) Recent advances in the analysis of complex glycoproteins. Anal Chem 89:389–413
Narimatsu H, Kaji H, Vakhrushev SY et al (2018) Current technologies for complex glycoproteomics and their applications to biology/disease-driven glycoproteomics. J Proteome Res 17:4097–4112
Kuno A, Uchiyama N, Koseki-Kuno S et al (2005) Evanescent-field fluorescence-assisted lectin microarray: a new strategy for glycan profiling. Nat Methods 2:851–856
Pilobello KT, Krishnamoorthy L, Slawek D et al (2005) Development of a lectin microarray for the rapid analysis of protein glycopatterns. Chembiochem 6:985–989
Hirabayashi J, Yamada M, Kuno A et al (2013) Lectin microarrays: concept, principle and applications. Chem Soc Rev 42:4443–4458
Matsuda A, Kuno A, Ishida H et al (2008) Development of an all-in-one technology for glycan profiling targeting formalin-embedded tissue sections. Biochem Biophys Res Commun 370:259–263
Matsuda A, Kuno A, Kawamoto T et al (2010) Wisteria floribunda agglutinin-positive mucin 1 is a sensitive biliary marker for human cholangiocarcinoma. Hepatology 52:174–182
Kuno A, Matsuda A, Ikehara Y et al (2010) Chapter 7—differential glycan profiling by lectin microarray targeting tissue specimens. Methods Enzymol 478:165–179
Tan B, Matsuda A, Zhang Y et al (2014) Multilectin-assisted fractionation for improved single-dot tissue glycome profiling in clinical glycoproteomics. Mol BioSyst 10:201–205
Yamashita K, Kuno A, Matsuda A et al (2016) Lectin microarray technology identifies specific lectins related to lymph node metastasis of advanced gastric cancer. Gastric Cancer 19:531–542
Espina V, Wulfkuhle JD, Calvert VS et al (2006) Laser-capture microdissection. Nat Protoc 1:586–603
Zou X, Yoshida M, Nagai-Okatani C et al (2017) A standardized method for lectin microarray-based tissue glycome mapping. Sci Rep 7:43560
Nagai-Okatani C, Nishigori M, Sato T et al (2019) Wisteria floribunda agglutinin staining for the quantitative assessment of cardiac fibrogenic activity in a mouse model of dilated cardiomyopathy. Lab Investig 99:1749–1765
Nagai-Okatani C, Aoki-Kinoshita KF, Kakuda S et al (2019) LM-GlycomeAtlas Ver. 1.0: a novel visualization tool for lectin microarray-based glycomic profiles of mouse tissue sections. Molecules 24:2962
Nagai-Okatani C, Zou X, Fujita N et al (2021) LM-GlycomeAtlas Ver. 2.0: an integrated visualization for lectin microarray-based mouse tissue glycome mapping data with lectin histochemistry. J Proteome Res 20:2069–2075
Yamashita S (2007) Heat-induced antigen retrieval: mechanisms and application to histochemistry. Prog Histochem Cytochem 41:141–200
Nagai-Okatani C, Nagai M, Sato T et al (2019) An improved method for cell type-selective glycomic analysis of tissue sections assisted by fluorescence laser microdissection. Int J Mol Sci 20:700
Kuno A, Kato Y, Matsuda A et al (2009) Focused differential glycan analysis with the platform antibody-assisted lectin profiling for glycan-related biomarker verification. Mol Cell Proteomics 8:99–108
Matsuda A, Higashi M, Nakagawa T et al (2017) Assessment of tumor characteristics based on glycoform analysis of membrane-tethered MUC1. Lab Investig 97:1103–1113
Kuno A, Matsuda A, Unno S et al (2014) Differential glycan analysis of an endogenous glycoprotein: toward clinical implementation—from sample pretreatment to data standardization. Methods Mol Biol 1200:265–285
Farr AG, Anderson SK (1985) Epithelial heterogeneity in the murine thymus: fucose-specific lectins bind medullary epithelial cells. J Immunol 134:2971–2977
Klug DB, Carter C, Crouch E et al (1998) Interdependence of cortical thymic epithelial cell differentiation and T-lineage commitment. Proc Natl Acad Sci U S A 95:11822–11827
Acknowledgments
We thank Dr. Takashi Sato, Dr. Hiroyuki Kaji, Dr. Erika Noro, and Prof. Hisashi Narimatsu of AIST for their helpful suggestions. We also thank Dr. Maki Yoshida, Dr. Jun Iwaki, Mr. Shuichi Kakuda, Ms. Misugi Nagai, Ms. Kozue Hagiwara, and Ms. Katsue Kiyohara of AIST for their technical assistance. This work was supported by a project for utilizing glycans in the development of innovative drug discovery technologies (grant number JP19ae0101021) from the Japan Agency for Medical Research and Development (AMED).
Author information
Authors and Affiliations
Corresponding authors
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2022 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Nagai-Okatani, C. et al. (2022). Tissue Glycome Mapping: Lectin Microarray-Based Differential Glycomic Analysis of Formalin-Fixed Paraffin-Embedded Tissue Sections. In: Kilcoyne, M., Gerlach, J.Q. (eds) Glycan Microarrays. Methods in Molecular Biology, vol 2460. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2148-6_10
Download citation
DOI: https://doi.org/10.1007/978-1-0716-2148-6_10
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-2147-9
Online ISBN: 978-1-0716-2148-6
eBook Packages: Springer Protocols