PRODUCTS OF ACTIVATED LYMPHOCYTES
PRODUCTS OF ACTIVATED LYMPHOCYTES
复制标题
活化淋巴细胞的产物
DOI：
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发表时间：
1973
影响因子：
15.3
通讯作者：
B. Bloom
中科院分区：
文献类型：
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作者：
C. Sorg;B. Bloom
开发了一般方法，并应用于微量活化淋巴细胞产物的生物合成和纯化。本文研究了纯化的蛋白衍生物（PPD）或伴刀豆球蛋白A（ConA）刺激豚鼠淋巴细胞产生的迁移抑制因子（MIF）的活性，所选的方法得到了有关分子的两个特征性化学参数，即SephadexG-75上的Kd和等电聚焦上的等离子点pI。当上清液在G-75柱上分级分离时，即使在对照上清液中也有几个区域相对于培养基对照产生迁移抑制。然而，在PPD-和Con A-刺激的上清液中，至少有一个峰的MIF活性被发现仅在刺激的培养物中，Kd为0.15。使用双标记技术来表征该峰的蛋白质。对照、未刺激的培养物用[14 C]亮氨酸标记，刺激的培养物用[3 H]亮氨酸标记。在混合上清液和G-75过滤后，主要的“比率标记的”宽峰。当此峰的一个窄的部分（Kd约0.15，包含大部分MIF活性）进行分析等电聚焦时，所有的标记物与净电荷比白蛋白低的蛋白质结合。在PPD和Con A刺激的组分中发现了一个独特的ratiolabeled峰，pI约为1.5。5.3.一个微制备等电聚焦技术的开发，并产生MIF活性在同一地区的主要ratiolabeled峰。将需要进一步的研究，以确定是否ratiolabelled蛋白是MIF。通过以下的Kd，pI，和3 H/14 C标记的比例，至少有14个产品的活化淋巴细胞，从头合成或增加量，可以区分。
General methods were developed and applied to the biosynthesis and purification of products of activated lymphocytes available in minute quantities. The activity studied here was the migration inhibitory factor (MIF) produced by purified protein derivative (PPD)- or concanavalin A (Con A)-stimulated lymphocytes obtained from one guinea pig or less. The methods selected yielded results in terms of two chemical parameters characteristic of the molecules involved, namely Kd on Sephadex G-75 and isoionic point, pI, on isoelectric focusing. When supernatants were fractionated on G-75 columns, there were several areas even in control supernatants which produced migration inhibition relative to medium controls. However, in PPD- and Con A-stimulated supernatants, at least one peak of MIF activity was found solely in the stimulated cultures, with a Kd of 0.15. A double-labeling technique was used to characterize the proteins of this peak. Control, unstimulated cultures were labeled with [14C]leucine and stimulated cultures were labeled with [3H]leucine. After mixing the supernatants and G-75 filtration, a major "ratiolabeled" broad peak. i.e. one with increased 3H/14C ratio, was found. When a narrow portion of this peak about Kd 0.15, containing most of the MIF activity, was subjected to analytical isoelectric focusing, all of the label was associated with proteins of lower net charge than albumin. A unique ratiolabeled peak was found in PPD- and Con A-stimulated fractions with a pI of approx. 5.3. A micropreparative isoelectric focusing technique was developed and yielded MIF activity in the same region as the major ratiolabeled peak. Further study will be required to ascertain whether the ratiolabeled protein is MIF. By following the Kd, pI, and 3H/14C labeling ratio, at least 14 products of activated lymphocytes, synthesized either de novo or in increased amounts, could be distinguished.