The development of an eluted stain bioassay (ESTA) for human growth hormone.
The development of an eluted stain bioassay (ESTA) for human growth hormone.
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开发人类生长激素的洗脱染色生物测定法 (ESTA)。
DOI：
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发表时间：
1995
期刊：
影响因子：
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通讯作者：
N. J. Marshall
中科院分区：
文献类型：
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作者：
P. A. Ealey;M. Yateman;R. Sandhu;M. Dattani;M. Hassan;S. J. Holt;N. J. Marshall
MTT-甲瓒生产由静止的Nb 2细胞和那些激活的胎牛血清或人生长激素（hGH）的基本特征进行了描述。利用这些特征开发用于生长激素等催乳素纯化制剂的MTT-ESTA生物测定法。所得体外生物测定法具有灵敏度和精密度，检测限约为0.05 mU hGH/l（19 ng/l），在存在0.3 mU hGH/l（114 ng/l）的情况下，测定内不精密度为2.5%。当利用静止的Nb 2细胞进行生物测定时，观察到大幅度的响应。反应的主要成分显然来自细胞的代谢活化，而不是增加的细胞增殖。这种反应被抗人生长激素所消除。后者的延迟添加证明，在整个96 h的推荐生物测定孵育期内需要激素的存在，以获得最大响应。在高剂量下，剂量-反应关系达到延长的平台期，其涵盖4个数量级的增量激素浓度。在测试的最高剂量10（6）mU hGH/l（385 mg/l）下观察到反应下降。这种自抑制与最近的报告一致，即由于连续受体二聚化的化学计量阻断而导致反应减少，这对于hGH激活促生长和催乳受体至关重要。
The basic characteristics of MTT-formazan production by both quiescent Nb2 cells and those activated by fetal calf serum or human growth hormone (hGH) are described. These characteristics are exploited for the development of an MTT-ESTA bioassay for purified preparations of lactogens such as growth hormone. The resulting in vitro bioassay is sensitive and precise, with a detection limit of about 0.05 mU hGH/l (19 ng/l) and a within-assay imprecision of 2.5% in the presence of 0.3 mU hGH/l (114 ng/l). When utilizing quiescent Nb2 cells for bioassays, large magnitudes of response are observed. The major component of the response is clearly derived from metabolic activation of the cells, rather than increased cell proliferation. The response was abolished by anti-human growth hormone. Delayed addition of the latter demonstrated that the presence of the hormone is required for the entire 96 h of the recommended bioassay incubation period to obtain the maximum response. At high doses, the dose-response relationship reaches a prolonged plateau which covers 4 orders of magnitude of incremental hormone concentrations. A decline in response is observed at the highest dose tested, 10(6) mU hGH/l (385 mg/l). This auto-inhibition is consistent with recent reports of a reduction in response due to stoichiometric blockade of sequential receptor dimerisation which is crucial for activation of both somatogenic and lactogenic receptors by hGH.