Development of an enzyme-linked immunosorbent assay for detection of chicken osteocalcin and its use in evaluation of perch effects on bone remodeling in caged White Leghorns
Development of an enzyme-linked immunosorbent assay for detection of chicken osteocalcin and its use in evaluation of perch effects on bone remodeling in caged White Leghorns
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开发用于检测鸡骨钙素的酶联免疫吸附测定法及其在评估鲈鱼对笼养白来航鸡骨重塑的影响中的应用
DOI：
10.3382/ps.2012-02909
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发表时间：
2013-08-01
期刊：
影响因子：
4.4
通讯作者：
Hou, J. -F.
中科院分区：
文献类型：
--
作者：
Jiang, S.;Cheng, H. W.;Hou, J. -F.
骨钙素（OC）是评估哺乳动物骨转换的敏感生化标志物。由于需要对鸡进行试验，禽类OC的作用尚不清楚。我们的目的是建立一种间接竞争性ELISA检测鸡血清OC的方法，并利用该方法研究鲈鱼对笼养母鸡骨重塑的影响。用重组大肠杆菌OC免疫家兔，制备了抗鸡OC多克隆抗体。以骨提取的鸡OC作为包被蛋白，纯化的鸡OC用于标定。建立的OC酶联免疫吸附试验的检出限为0.13 ng/mL。试验内CV < 7，试验间CV < 12%。将所建立的OC酶联免疫吸附试验与市售的大鼠-中型OC酶联免疫吸附试验对饲养在有或无栖木的常规笼中的蛋鸡进行敏感性比较。选取71周龄来港白母鸡进行4种处理，采集血清样本。处理1是在其生命周期中从未接触过栖木的对照鸡。处理2鸡在雏鸡期（0 ~ 16.9周龄）有栖木，而处理3鸡只在生命周期的产蛋期（17 ~ 71周龄）有栖木。处理4的鸡（0 ~ 71周龄）均可获得栖木。两项检测的相关性为0.62 （P < 0.0001）。鸡用ELISA法检测血清OC水平高于大鼠- mid ELISA法（P < 0.0001）。鸡的酶联免疫吸附试验结果显示，有栖木的母鸡产蛋期间血清OC浓度高于没有栖木的母鸡（P = 0.04）。雏鸡接近栖木对71周龄母鸡血清OC水平无影响（P = 0.15）。总之，鸡骨重塑酶联免疫吸附试验具有灵敏、准确和足够的鉴别能力，可用于测定鸡骨重塑。
Osteocalcin (OC) is a sensitive biochemical marker for evaluating bone turnover in mammals. The role of avian OC is less clear because of the need for a chicken assay. Our objectives were to develop an assay using indirect competitive ELISA for detecting chicken serum OC and use the assay to examine the effects of perches on bone remodeling in caged hens. Anti-chicken OC polyclonal antibody was produced by immunization of rabbits with a recombinant OC from Escherichia coli. Chicken OC extracted from bone was used as a coated protein, and purified chicken OC was used for calibration. The limit of detection of the developed OC ELISA was 0.13 ng/mL. The intra-and interassay CV were < 7 and < 12%, respectively. The sensitivity of the developed OC ELISA was compared with a commercial Rat-Mid OC ELISA in laying hens housed in conventional cages with or without perches. Serum samples were collected from 71-wk-old White Leghorn hens subjected to 4 treatments. Treatment 1 was control chickens that never had access to perches during their life cycle. Treatment 2 chickens had perches during the pullet phase (0 to 16.9 wk of age), whereas treatment 3 chickens had perches only during the egg-laying phase of the life cycle (17 to 71 wk of age). Treatment 4 chickens always had access to perches (0 to 71 wk of age). Correlation between the 2 assays was 0.62 (P < 0.0001). Levels of serum OC using the developed chicken ELISA were higher than that detected using the Rat-Mid ELISA (P < 0.0001). Results from the chicken ELISA assay showed that hens with perch access had higher concentrations of serum OC than hens without perches during egg laying (P = 0.04). Pullet access to perches did not affect serum OC levels in 71-wk-old hens (P = 0.15). In conclusion, a chicken OC ELISA has been validated that is sensitive and accurate with adequate discriminatory power for measuring bone remodeling in chickens.