Guico-Lamm M L, Voss E W, Sherwood O D
Department of Physiology and Biophysics, University of Illinois, Urbana 61801.
Endocrinology. 1988 Nov;123(5):2472-8. doi: 10.1210/endo-123-5-2472.
The physiological role of relaxin during pregnancy and at parturition in the rat is not absolutely established. There are limitations to the experimental approach used in the few studies that examined the influence of relaxin in the pregnant rat. These studies were unphysiological, since they involved administration of porcine relaxin as well as progesterone and estrogen to ovariectomized pregnant rats. A more physiological approach is to use antibodies to neutralize the biological actions of endogenous relaxin in the intact pregnant rat. The purpose of the present study was to produce and characterize monoclonal antibodies suitable for this approach. Six stable and rapidly growing hybridoma clones which produced monoclonal antibodies specific for rat relaxin (MCA-rR) were obtained after the fusion of NSO mouse myeloma cells with lymphocytes from the spleen of a BALB/c mouse immunized with rat relaxin. Five MCA-rR (MCA1-5; all immunoglobulin G1 kappa) inhibited the ability of exogenously administered rat relaxin to increase the interpubic ligament length in estrogen-primed mice. Of the five MCA-rR that neutralized rat relaxin's bioactivity in vivo, MCA1 exhibited the highest relative affinity for rat relaxin. MCA1 was also highly specific for rat relaxin. MCA1 demonstrated no cross-reactivity with rat insulin, rat insulin-like growth factor I and II, or porcine relaxin-proteins that are structurally related to rat relaxin. In view of its high affinity and high specificity for rat relaxin as well as its ability to neutralize rat relaxin's bioactivity in vivo, MCA1 was selected for use in subsequent studies aimed at the neutralization of endogenous relaxin in intact pregnant rats.
松弛素在大鼠孕期及分娩时的生理作用尚未完全明确。在少数研究松弛素对妊娠大鼠影响的实验方法存在局限性。这些研究不符合生理学原理，因为它们涉及给去卵巢的妊娠大鼠注射猪松弛素以及孕酮和雌激素。一种更符合生理学的方法是使用抗体来中和完整妊娠大鼠体内内源性松弛素的生物学作用。本研究的目的是制备并鉴定适用于该方法的单克隆抗体。将NSO小鼠骨髓瘤细胞与用大鼠松弛素免疫的BALB/c小鼠脾脏中的淋巴细胞融合后，获得了六个稳定且生长迅速的杂交瘤克隆，它们产生了针对大鼠松弛素的单克隆抗体（MCA-rR）。五种MCA-rR（MCA1-5；均为免疫球蛋白G1 κ）抑制了外源性给予的大鼠松弛素增加雌激素预处理小鼠耻骨间韧带长度的能力。在体内中和大鼠松弛素生物活性的五种MCA-rR中，MCA1对大鼠松弛素表现出最高的相对亲和力。MCA1对大鼠松弛素也具有高度特异性。MCA1与大鼠胰岛素、大鼠胰岛素样生长因子I和II或猪松弛素（与大鼠松弛素结构相关的蛋白质）均无交叉反应。鉴于其对大鼠松弛素具有高亲和力和高特异性，以及在体内中和大鼠松弛素生物活性的能力，MCA1被选用于后续旨在中和完整妊娠大鼠体内内源性松弛素的研究。
