PMab-235: A monoclonal antibody for immunohistochemical analysis against goat podoplanin

Yoshikazu Furusawa,Shinji Yamada,Takuro Nakamura,Masato Sano,Yusuke Sayama,Shunsuke Itai,Junko Takei,Hiroyuki Harada,Masato Fukui,Mika K Kaneko,Yukinari Kato

doi:10.1016/j.heliyon.2019.e02063

Yoshikazu Furusawa, Shinji Yamada + Show 9 more

Open Access

https://doi.org/10.1016/j.heliyon.2019.e02063

Copy DOI

Journal: Heliyon	Publication Date: Jul 1, 2019
Citations: 32	License type: cc-by

Affiliation: Tohoku University, Tokyo Medical and Dental University

Abstract

Sensitive and specific monoclonal antibodies (mAbs) against not only human but also mouse, rat, rabbit, dog, cat, bovine, pig, and horse podoplanins (PDPNs) have been established in our previous studies. However, anti-goat PDPN (gPDPN) has not been established yet. PDPN has been utilized as a lymphatic endothelial cell marker especially in pathological diagnoses; therefore, mAbs for immunohistochemical analyses using formalin-fixed paraffin-embedded tissues are needed. Although we recently demonstrated that an anti-bovine PDPN mAb, PMab-44 cross-reacted with gPDPN, PMab-44 did not detect lymphatic endothelial cells in immunohistochemistry. In this study, we immunized mice with gPDPN-overexpressing Chinese hamster ovary (CHO)–K1 (CHO/gPDPN) cells, and screened mAbs against gPDPN using flow cytometry. One of the mAbs, PMab-235 (IgG1, kappa), specifically detected CHO/gPDPN cells by flow cytometry. Furthermore, PMab-235 strongly detected lung type I alveolar cells, renal podocytes, and lymphatic endothelial cells of colon by immunohistochemistry. These findings suggest that PMab-235 may be useful as a lymphatic endothelial cell marker for goat tissues.

Highlights

Podoplanin (PDPN)/T1alpha/Aggrus is a type I transmembrane sialoglycoprotein, which is expressed in many cell types, such as pulmonary type I alveolar cells, renal podocytes, mesothelial cells, and epithelial cells or lymphatic endothelial cells of many organs [1]
The immunohistochemical analyses revealed that PMab-235 strongly stained type I alveolar cells of lung (Fig. 4A and B), podocytes of kidney (Fig. 5A and B), and lymphatic endothelial cells of colon (Fig. 6A and B)
Because anti-goat PDPN (gPDPN) monoclonal antibodies (mAbs), which are useful for immunohistochemical analysis to detect lymphatic endothelial cells, have not been reported, specific detection of lymphatic endothelial cells was difficult

Summary

Introduction

Podoplanin (PDPN)/T1alpha/Aggrus is a type I transmembrane sialoglycoprotein, which is expressed in many cell types, such as pulmonary type I alveolar cells, renal podocytes, mesothelial cells, and epithelial cells or lymphatic endothelial cells of many organs [1]. PDPN could induce platelet aggregation by binding to the endogenous receptor of PDPN, C-type lectin-like receptor-2 (CLEC-2) [2]. Sialic acid-deficient PDPN recovered its activity after additional sialylation, indicating that the sialylated core of Thr is critical for PDPN-induced platelet aggregation. Our previous studies demonstrated that PDPN expression in Chinese hamster ovary (CHO)–K1 cells promoted pulmonary metastasis in both an experimental and a spontaneous mouse model [14]. PDPN-expressing cells, which were covered with platelets, were found to be arrested in the lung microvasculature 30 minutes after injection. Lung metastasis resulting from PDPN expression decreased the survival of the mice. Inhibition of platelets with aspirin reduced the formation of PDPN-promoted metastasis, indicating that

Methods

Results

Conclusion