Laboratory of Endocrinology and Animal Reproduction, Faculty of Veterinary Medicine, University of Liege, Belgium.
Acta Vet Scand. 2010 Feb 3;52(1):9. doi: 10.1186/1751-0147-52-9.
The involvement of placental lactogen (PL) in the regulation of foetal growth has been investigated in different species by in vivo immunomodulation techniques. However, when circulating antibodies are present together with the hormone, the procedure for hormonal measurement becomes considerably complex. The aim of this study was the immunoneutralization of bovine placental lactogen (bPL) concentrations in bovine foetal circulation by direct infusion of rabbit anti-bPL purified immunoglobulins (IgG) via a foetal catheter (in vivo study). The ability of a RIA based on guinea pig anti-bPL antiserum, for the measurement of bPL concentrations in samples containing exogenous rabbit anti-bPL immunoglobulins, was also analyzed in in vitro and in vivo conditions.
Six bovine foetuses were chronic cannulated on the aorta via the medial tarsal artery. Infusion of rabbit anti-bPL IgG was performed during late gestation. Pooled rabbit anti-bPL antisera had a maximal neutralization capacity of 25 microg bPL/mL of immunoglobulin. Interference of rabbit anti-bPL immunoglobulin with radioimmunoassay measurement using guinea pig anti-bPL as primary antibody was first evaluated in vitro. Polyclonal anti-bPL antibodies raised in rabbit were added in foetal sera to produce 100 samples with known antibodies titers (dilutions ranging from 1:2,500 till 1:1,280,000).
RESULT(S): Assessment of the interference of rabbit anti-bPL antibody showed that bPL concentrations were significantly lower (P < 0.05) in samples added with dilutions of rabbit antiserum lower than 1:80,000 (one foetus) or 1:10,000 (four foetuses). It was also shown that the recovery of added bPL (12 ng/mL) was markedly reduced in those samples in which exogenous rabbit anti-bPL were added at dilutions lower than 1:20,000. Concentrations of foetal bPL were determined in samples from cannulated foetuses. In foetuses 1 and 6, bPL concentrations remained almost unchanged (<5 ng/mL) during the whole experimental period. In Foetus 3, bPL concentrations decreased immediately after IgG infusion and thereafter, they increased until parturition.
CONCLUSION(S): The use of a bPL RIA using a guinea pig anti-bPL as primary antiserum allowed for the measurement of bPL concentrations in foetal plasma in presence of rabbit anti-bPL IgG into the foetal circulation. Long-term foetal catheterization allowed for the study of the influence of direct infusion of anti-bPL IgG on peripheral bPL concentrations in bovine foetuses.
通过体内免疫调节技术，已经在不同物种中研究了胎盘催乳素（PL）在调节胎儿生长中的作用。然而，当循环抗体与激素同时存在时，激素测量的程序会变得相当复杂。本研究的目的是通过胎儿导管直接输注兔抗牛胎盘催乳素（bPL）纯化免疫球蛋白（IgG），对牛胎儿循环中的牛胎盘催乳素（bPL）浓度进行免疫中和（体内研究）。还分析了基于豚鼠抗 bPL 抗血清的 RIA 在体内和体外条件下测量含有外源性兔抗 bPL 免疫球蛋白的样品中 bPL 浓度的能力。
通过内侧跗骨动脉对 6 只牛胎儿进行慢性主动脉内插管。在妊娠晚期进行兔抗 bPL IgG 输注。兔抗 bPL 抗血清的最大中和能力为 25μg bPL/mL 免疫球蛋白。首先在体外评估兔抗 bPL 免疫球蛋白对使用豚鼠抗 bPL 作为初级抗体的放射免疫测定测量的干扰。在兔中产生的多克隆抗 bPL 抗体被添加到胎儿血清中，以产生 100 个具有已知抗体滴度的样品（稀释度范围从 1:2500 到 1:1280000）。
评估兔抗 bPL 抗体的干扰表明，当添加的兔抗血清稀释度低于 1:80000（一只胎儿）或 1:10000（四只胎儿）时，样品中的 bPL 浓度显着降低（P<0.05）。还表明，在添加的外源性兔抗 bPL 稀释度低于 1:20000 的情况下，添加的 bPL（12ng/mL）的回收率明显降低。在插管胎儿的样品中测定了胎儿 bPL 的浓度。在胎儿 1 和 6 中，整个实验期间 bPL 浓度几乎保持不变（<5ng/mL）。在胎儿 3 中，bPL 浓度在 IgG 输注后立即下降，此后直至分娩时增加。
使用豚鼠抗 bPL 作为初级抗血清的 bPL RIA 允许在牛胎儿循环中存在兔抗 bPL IgG 的情况下测量胎儿血浆中的 bPL 浓度。长期胎儿导管化允许研究直接输注抗 bPL IgG 对牛胎儿外周 bPL 浓度的影响。
