The quantitation of human growth hormone by a radioreceptor assay using an established human cell line

Thore Nederman , Lars Sjödin
{"title":"The quantitation of human growth hormone by a radioreceptor assay using an established human cell line","authors":"Thore Nederman ,&nbsp;Lars Sjödin","doi":"10.1016/0092-1157(87)90023-0","DOIUrl":null,"url":null,"abstract":"<div><p>Membrane receptors on cultured human lymphocytes (IM-9) have been shown to bind human growth hormone (hGH) in a specific manner. The aim of the present study was to develop an <em>in vitro</em> assay of hGH based on this binding. The assay should fulfil established pharmacopoeial requirements for quantitation of hormones. The binding of [<sup>125</sup>I]hGH was studied as a function of time, temperature, cell density, tracer concentration and the concentration of unlabelled hGH and other related hormones. Also, the dissociation of bound hGH and the chemical stability of hGH in the incubation medium were studied. From these studies, the conditions for an appropriate radioreceptor assay were determined. Briefly, 1·5–3·0 × 10<sup>7</sup> cells ml<sup>−1</sup> were incubated with 5–20 × 10<sup>−12</sup> <span>m</span> [<sup>125</sup>I]hGH and three different concentrations of unlabelled hGH chosen from the linear part of the [<sup>125</sup>I]hGH displacement curve. The results were analyzed according to general pharmacopoeial principles. The mean values for growth hormone activity tested by radioreceptor assay were within the fiducial limits (<em>P</em> = 0·05) of the corresponding activity determined by the hypophysectomized rat body-weight gain assay. The <em>in vitro</em> assay was found to be more precise and less resource demanding than the <em>in vivo</em> bioassay of hGH. It is concluded that the <em>in vitro</em> bioassay described here is well suited as a screening method for potency determination of hGH preparations.</p></div>","PeriodicalId":75991,"journal":{"name":"Journal of biological standardization","volume":"15 3","pages":"Pages 199-211"},"PeriodicalIF":0.0000,"publicationDate":"1987-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/0092-1157(87)90023-0","citationCount":"10","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of biological standardization","FirstCategoryId":"1085","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/0092115787900230","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 10

Abstract

Membrane receptors on cultured human lymphocytes (IM-9) have been shown to bind human growth hormone (hGH) in a specific manner. The aim of the present study was to develop an in vitro assay of hGH based on this binding. The assay should fulfil established pharmacopoeial requirements for quantitation of hormones. The binding of [125I]hGH was studied as a function of time, temperature, cell density, tracer concentration and the concentration of unlabelled hGH and other related hormones. Also, the dissociation of bound hGH and the chemical stability of hGH in the incubation medium were studied. From these studies, the conditions for an appropriate radioreceptor assay were determined. Briefly, 1·5–3·0 × 107 cells ml−1 were incubated with 5–20 × 10−12 m [125I]hGH and three different concentrations of unlabelled hGH chosen from the linear part of the [125I]hGH displacement curve. The results were analyzed according to general pharmacopoeial principles. The mean values for growth hormone activity tested by radioreceptor assay were within the fiducial limits (P = 0·05) of the corresponding activity determined by the hypophysectomized rat body-weight gain assay. The in vitro assay was found to be more precise and less resource demanding than the in vivo bioassay of hGH. It is concluded that the in vitro bioassay described here is well suited as a screening method for potency determination of hGH preparations.

利用已建立的人类细胞系,用放射受体测定法测定人类生长激素
培养的人淋巴细胞上的膜受体(IM-9)已被证明以特定的方式结合人生长激素(hGH)。本研究的目的是建立一种基于这种结合的生长激素体外检测方法。该方法应满足既定药典中激素定量的要求。研究了[125I]hGH的结合与时间、温度、细胞密度、示踪剂浓度、未标记hGH及其他相关激素浓度的关系。此外,还研究了结合生长激素的解离和生长激素在培养介质中的化学稳定性。从这些研究中,确定了适当的放射受体测定的条件。简单地说,1·5-3·0 × 107细胞ml - 1与5-20 × 10 - 12 m的[125I]hGH和从[125I]hGH位移曲线的线性部分选择的三种不同浓度的未标记的hGH孵育。根据一般药典原理对结果进行分析。放射受体法测定的生长激素活性平均值在垂体去骨大鼠增重法测定的相应活性的基准范围内(P = 0.05)。发现体外测定比体内测定hGH更精确,所需资源更少。结论是,体外生物测定法很适合作为hGH制剂效力测定的筛选方法。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 求助全文
来源期刊
自引率
0.00%
发文量
0
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:604180095
Book学术官方微信