鲫鱼斯坦尼小体降血钙因子的初步鉴定

采用向春季鲫鱼体腔注射Cacl2，导致身钙升高的方法，使其含有降血钙因子——斯氏降钙素的斯坦尼小体Ⅰ型细胞的分泌颗粒应答性地释放殆尽。然后对这种不含斯氏降钙素的斯坦尼小体组织匀浆和含有斯氏降钙素的斯坦尼小体组织匀浆的电泳带进行比较、分析，从而鉴定出鲫鱼坦尼小体中的降血钙因子是一种表观分子量为48kD的糖蛋白，其解聚物的分子量为24kD。

Perliminary Identification of Stanniocalcin of the Stannius Corpuscles of Carassius auratus

After Carassius auratus was fed in fresh water in lab for 24 hours,3 groups were performed: group 1 received intraperitoneal injection of 0.68?mol/L?CaCl 2 solution (0.1?mL/100?g.b.w/24 h); group 2 were injected with 6% NaCl solution as control;group 3 were untreated as normal control. After the first injection, blood was collected from the fish's blood vessels of the caudal to determine the serum ionic calcium concentration by using calcium electrode and ion analyser at different times. Four hours after the second injection, the CS were removed from the rest of the fish and prepared for electron microscopy or SDS PAGE. The ultrathin sections of CS for electron microscopy were prepared in routine procedure (Double fixed in glutaraldehyde and Osmium tetroxide, embedded in Epon812, double stained with uranyl acetate and plumbum citrate), and CS tissue homogenates were prepared by homogenizing 30?mg of fresh CS tissue in 1?mL of 0.062?5mol/L Tris HCl buffer (pH?6.8). The supernatant for SDS PAGE was obtained after centrifugation (1?500?r/min, 3?min). Discontinuity SDS PAGE was performed with 5% of gels while concentrating and 15% of gels while separating. In order to investigate the effects of reducing reagents on CS, SDS PAGE was carried out under reducing conditions (with mercaptoethanol) or nonreducing conditions (without mercaptoethanol). After electrophorsis, slab gels were either fixed, stained and photographed or prepared for surveying glycoprotein. The result of calcium determined indicated that the serum Ca 2+ increased significantly in CaCl 2 injected group, but not in NaCl injected group. Electron micrographs showed that the CS of the NaCl injected or untreated Carassius auratus contained an abundance of large secretory granules, which are typical for the type I cells; the CaCl 2 injected fish were almost devoid of these secretory granules. Electrophorsis results indicated: A glycoprotein with an apparent molecular weight of 48?kD was observed under nonreducing condition, but it disappeared after CaCl 2 injection. The glycoprotein had an apparent molecular weight of 24?kD only under reducing condition. The glycoprotein was regarded as the stanniocalcin of Carassiue auratus.