Biochemical and immunological characterization of a novel monoclonal antibody against mouse leukotriene B4 receptor 1.
Biochemical and immunological characterization of a novel monoclonal antibody against mouse leukotriene B4 receptor 1.
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DOI：
10.1371/journal.pone.0185133
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发表时间：
2017
期刊：
影响因子：
3.7
通讯作者：
Yokomizo T
中科院分区：
文献类型：
--
作者：
Sasaki F;Koga T;Saeki K;Okuno T;Kazuno S;Fujimura T;Ohkawa Y;Yokomizo T
白三烯B4（LTB 4）受体1（BLT 1）是一种在各种白细胞亚群中表达的G蛋白偶联受体;然而，由于尚未获得mBLT 1单克隆抗体（mAb），因此尚未报道小鼠BLT 1（mBLT 1）的精确表达。在这项研究中，我们提出了一个杂交瘤细胞系（克隆7A 8），产生一个高亲和力的单克隆抗体mBLT 1的直接免疫的BLT 1缺陷小鼠与mBLT 1过表达细胞的成功建立。使用mBLT 1过表达细胞和内源性表达BLT 1的小鼠外周血白细胞证实了克隆7A 8的特异性。克隆7A 8不与人BLT 1或其他G蛋白偶联受体（包括人趋化因子（C-X-C基序）受体4）交叉反应。7A 8 mAb与mBLT 1的第二个细胞外环结合，不影响LTB 4结合或LTB 4的细胞内钙动员。7A 8 mAb对外周血中的Gr-1阳性粒细胞、CD 11b阳性粒细胞/单核细胞、F4/80阳性单核细胞、CCR 2高和CCR 2低单核细胞亚群以及CD 4阳性T细胞亚群（从初始CD 4阳性T细胞体外分化的Th 1细胞）进行了阳性染色。该mAb能够通过免疫组织化学检测到幼稚小鼠脾脏中的Gr-1阳性粒细胞和单核细胞。最后，腹膜内施用7A 8 mAb耗尽外周血中的粒细胞和单核细胞。因此，我们成功地产生了高亲和力的抗mBLT 1单克隆抗体，这是有用的分析mBLT 1在体外和体内的表达。
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.
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DOI：
10.1084/jem.188.6.1063
发表时间：
1998-09-21
期刊：
The Journal of experimental medicine
影响因子：
--
作者：
Huang WW;Garcia-Zepeda EA;Sauty A;Oettgen HC;Rothenberg ME;Luster AD
通讯作者：
Luster AD
DOI：
10.1084/jem.192.3.433
发表时间：
2000-08-07
期刊：
The Journal of experimental medicine
影响因子：
--
作者：
Haribabu B;Verghese MW;Steeber DA;Sellars DD;Bock CB;Snyderman R
通讯作者：
Snyderman R
影响因子：
7.3
作者：
Galvani S;Sanson M;Blaho VA;Swendeman SL;Obinata H;Conger H;Dahlbäck B;Kono M;Proia RL;Smith JD;Hla T
通讯作者：
Hla T
影响因子：
64.8
作者：
通讯作者：
--
影响因子：
64.8
作者：
AbdAlla, S;Lother, H;Quitterer, U
通讯作者：
Quitterer, U