Biochemical and immunological characterization of a novel monoclonal antibody against mouse leukotriene B4 receptor 1

Fumiyuki Sasaki, Tomoaki Koga, Kazuko Saeki, Toshiaki Okuno, Saiko Kazuno, Tsutomu Fujimura, Yasuyuki Ohkawa, Takehiko Yokomizo

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Leukotriene B4 (LTB4) receptor 1 (BLT1) is a G protein-coupled receptor expressed in various leukocyte subsets; however, the precise expression of mouse BLT1 (mBLT1) has not been reported because a mBLT1 monoclonal antibody (mAb) has not been available. In this study, we present the successful establishment of a hybridoma cell line (clone 7A8) that produces a high-affinity mAb for mBLT1 by direct immunization of BLT1-deficient mice with mBLT1-overexpressing cells. The specificity of clone 7A8 was confirmed using mBLT1-overexpressing cells and mouse peripheral blood leukocytes that endogenously express BLT1. Clone 7A8 did not cross-react with human BLT1 or other G protein-coupled receptors, including human chemokine (C-X-C motif) receptor 4. The 7A8 mAb binds to the second extracellular loop of mBLT1 and did not affect LTB4 binding or intracellular calcium mobilization by LTB4. The 7A8 mAb positively stained Gr-1-positive granulocytes, CD11b-positive granulocytes/monocytes, F4/80-positive monocytes, CCR2-high and CCR2-low monocyte subsets in the peripheral blood and a CD4-positive T cell subset, Th1 cells differentiated in vitro from naïve CD4-positive T cells. This mAb was able to detect Gr-1-positive granulocytes and monocytes in the spleens of naïve mice by immunohistochemistry. Finally, intraperitoneal administration of 7A8 mAb depleted granulocytes and monocytes in the peripheral blood. We have therefore succeeded in generating a high-affinity anti-mBLT1 mAb that is useful for analyzing mBLT1 expression in vitro and in vivo.

Original languageEnglish
Article numbere0185133
JournalPloS one
Volume12
Issue number9
DOIs
Publication statusPublished - Sep 2017

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leukotrienes
monoclonal antibodies
Monoclonal Antibodies
receptors
mice
monocytes
Monocytes
granulocytes
Granulocytes
Blood
Leukotriene B4
T-cells
G-Protein-Coupled Receptors
Leukotriene B4 Receptors
Clone Cells
Immunization
clones
leukocytes
blood
Leukocytes

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • General

Cite this

Sasaki, F., Koga, T., Saeki, K., Okuno, T., Kazuno, S., Fujimura, T., ... Yokomizo, T. (2017). Biochemical and immunological characterization of a novel monoclonal antibody against mouse leukotriene B4 receptor 1. PloS one, 12(9), [e0185133]. https://doi.org/10.1371/journal.pone.0185133

Biochemical and immunological characterization of a novel monoclonal antibody against mouse leukotriene B4 receptor 1. / Sasaki, Fumiyuki; Koga, Tomoaki; Saeki, Kazuko; Okuno, Toshiaki; Kazuno, Saiko; Fujimura, Tsutomu; Ohkawa, Yasuyuki; Yokomizo, Takehiko.

In: PloS one, Vol. 12, No. 9, e0185133, 09.2017.

Research output: Contribution to journalArticle

Sasaki, Fumiyuki ; Koga, Tomoaki ; Saeki, Kazuko ; Okuno, Toshiaki ; Kazuno, Saiko ; Fujimura, Tsutomu ; Ohkawa, Yasuyuki ; Yokomizo, Takehiko. / Biochemical and immunological characterization of a novel monoclonal antibody against mouse leukotriene B4 receptor 1. In: PloS one. 2017 ; Vol. 12, No. 9.
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