Establishment of a Radioimmunoassay of Somatostatin in Human Plasma and in Rat Hypothalamus

人體血漿及大白鼠下視丘體抑素放射免疫分析法之建立

何橈通(L. T. Ho) ； 林興中(H. C. Lam) ； 王振濤(J. T. Wang) ； 鄧錦泉(K. T. Tang) ； 彭敬成(J. C. Perng) ； 劉岳芬(Y. F. Liu) ； 陳如菱(R. L. Chen)

《The Chinese Journal of Physiology》 29卷2期 (1986/12) Pp. 91-103

摘要

利用Carbodiimide，將環狀Somatostatin-14(SRIF-14)結合在血藍質(Hemocyanin)上去疫種大白兎，可獲得高靈敏度，高特異性，而又力價高(1:5,000-100,000)的抗體血清。而製成的125^I-Tyr-SRIF經過CM-52陽離子交換柱管的純化，其免疫特異活性(SP.AC.)約在1013 mCi/mg左右，比起市售的商品或其他國外報告的追蹤劑還要高。試驗的靈敏度在16 pg/ml左右，能測範圍在20至2000 Pg/ml之間。人類血漿標本的準備必需在含有Aprotinin的試管內，立卽離心取得。老鼠的腦下垂體經過磨勻後，以鹽酸萃取，並加溫至95℃，然後離心，取上清液測定。所得到的醋酸緩衝液（pH5.2）、血漿劑量、血漿回收劑量、腦下垂體萃取劑量及腦下垂體回收劑量等標準曲線群，均有良好的平行性。SRIF在人血漿和老鼠腦下垂體的回收率分別是98.8±6.3%和86.9±6.8％。抗體血清的特異性很高，只與H2-SRIF和Tyr1-SRIF產生交互反應，而分別是環狀SRIF-14免疫反應活性的15％和170％。結合數據的斯克伽投圖成一直線，Kd是3.52×10^(-12)M和結合容量是4.06×l0^(-10)M 。試驗中和試驗間之變異係數分別是4.5％和12％。將合成的環狀SRIF(Stilamin)經由靜脈注射到正常自願者身上，可得到重現性甚高的血漿SRIF-LI時間－劑量曲線，而獲知其在血中的半衰期約為1.5分鐘，代謝廓清率約為50ml/kg/min。

並列摘要

Ho, L. T., H. C. Lam, J. T. Wang, K. T. Tang, J. C. Perng, Y. F. Liu and R. L. Chen: Establishment of a radioimmunoassay of somatostatin in human plasma and in rat hypothalamus. Chinese J. Physiol, 29(2): 91-103, 1986: Synthetic cyclic somatostatin-14 (SRIF-14) was conjugated with carbodiimide onto hemocyanin to immunize rabbits, and anti-sera with high titers (1:5,000-100,000) and good sensitivity and specificity wereobtained. 125^I-Tyr1-SRIF was made and purified on CM-52 cation exchange column. The immunoreactive specific activity (SP.AC.) of the tracer was around 101 3mCi/mg, whichwas much higher than the commercial products and other reports. The sensitivity of the assay was around 16 pg/ml, and the detectable range 20-2000 pg/ml. Human plasma was freshly prepared in the presence of aprotinin. Rat hypothalamus was homogenized and extracted with HC1 followed by heating at 95℃ then centrifuged, and the supernatant assayed. The standard curves of the acetate buffer (ph 5.2), plasma doses, plasma recovery doses, hypothalamic extract doses, and hypothalamic extract recovery doses were all in good parallelism with each other. The recovery of SRIF from human plasma and from rat hypothalamus were 98.8±6.3% and 86.9±6.8%, respectively. The specificity of the antisera were very high, and they only cross-react with H2-SRIF and Tyr1-SRIF at 15% and 170%, respectively, of the immunoreactivity to cyclic SRIF-14. The Scatch-ard plot of the binding data showed a straight line with a Kd of 3.52×10^(-12)M and binding capacity of 4.06×10^(-10)M. The intra- and inter-assay coefficient of variation were 4.5% and 12%, respectively. When the synthetic cyclic SRIF (Stilamin) was infused intravenously into normal volunteers, there were reproducible plasma time-dose curves of SRIF-LI (SRIF Like Immunoreactivity), which revealed its plasma half life around1.5 minutes, and metabolic clearance rate about 50ml/kg/min.

並列關鍵字

Human plasma ； Pharmacokinetics ； Radioimmunoassay ； Rat Hypothalamus ； Scatchard plot ； Somatostatin