Partial purification and characterization of protection-inducing antigens from the muscle larva of Trichinella spiralis by molecular sizing chromatography and preparative flatbed isoelectric focusing.
Partial purification and characterization of protection-inducing antigens from the muscle larva of Trichinella spiralis by molecular sizing chromatography and preparative flatbed isoelectric focusing.
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通过分子大小色谱和制备平板等电聚焦对旋毛虫肌肉幼虫的保护诱导抗原进行部分纯化和表征。
DOI：
10.1111/j.1365-3024.1981.tb00405.x
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发表时间：
1981
影响因子：
2.2
通讯作者：
Despommier,DD
中科院分区：
文献类型：
--
作者：
Despommier,DD
来源于肌肉幼虫的大颗粒部分的可溶部分。用Sephacryl S‐200进行分子分级柱层析。得到了280 nm吸收材料的5个主要峰。免疫电泳分析显示，每个峰都含有抗原，其中大部分抗原出现在峰3、峰4和峰5。初步研究表明，峰4 （mol. wt范围20 000 ~ 10 000）含有保护诱导抗原。峰4的交叉免疫电泳和单维电泳分析显示至少有10种抗原，而分析等电聚焦显示存在pl相差很大的蛋白质，范围在4·0到9·0之间。峰4采用制备平板等电聚焦（PIEF），采用3.5 ~ 9.5梯度和3.5 ~ 5.5梯度对峰4进行分馏。两组的融合火箭免疫电泳（FRIEP）分析表明，有几个抗原与其他抗原分离：一个在pl 4.0，另一个在pl 9.0。其余抗原集中在pl 4·3 ~ 4·9之间。将全峰4、pl 9.0抗原和pl 3.4 - 4·9抗原组各100微克与Freund完全佐剂一起分别注射到小鼠体内。此外，还检测了部分pl4·0抗原的保护作用。所有抗原制剂都能产生显著的保护作用。用高效液相色谱（HPLC）进一步分析pl 4.0。在（2-18）（脂肪族）柱上，用65%乙腈等比洗脱经FRIEP检测的两个尖峰抗原。两个抗原峰在FRIEP上表现出完全的交叉反应性，并在220 nm处被吸收。氨基酸分析显示，各HPLC峰的组成没有可检测到的差异。每个峰均以天冬氨酸（13mol %）和谷氨酸（18mol %）为主。该抗原不含大量芳香族氨基酸，在206nm处被强吸收。ppl 4.0和ppl 9.0抗原均未被PAS反应阳性染色。
The soluble portion of a large particle fraction which was derived from the muscle larva ofT. spiraliswas subjected to molecular sizing column chromatography using Sephacryl S‐200. Five major peaks of 280 nm absorbing material were obtained. Analysis by immunoelectrophoresis revealed that each peak contained antigens, with the majority of them occurring in peaks 3,4 and 5. Preliminary studies indicated that peak 4 (mol. wt range 20 000‐10 000) contained protection‐inducing antigens. Crossed‐immunoelectrophoretic and single‐dimension electrophoretic analysis of peak 4 revealed a minimum of 10 antigens, while analytical isoelectric focusing demonstrated the presence of proteins with widely different pl, ranging from 4·0 to 9·0. Peak 4 was fractionated by preparative flatbed isoeletric focusing (PIEF) using two gradients: one from 3·5 to 9·5 and the other from 3·5 to 5·5. Fused rocket immunoelectrophoretic (FRIEP) analysis of both runs indicated that several antigens were separated from the others: one at pl 4·0 and the other at pl 9·0. The remaining antigens focused between pl 4·3 and 4·9. One hundred micrograms of whole peak 4, pl 9·0 antigen and the group of antigens at pl 4·3–4·9 were each separately injected, along with Freund's complete adjuvant, into mice. In addition, a portion of the pl 4·0 antigen was also assayed for protection. All antigenic preparations induced significant levels of protection. The pl 4·0 was further analysed on high‐performance liquid chromatography (HPLC). Two sharp peaks of antigen, as detected by FRIEP, were eluted isocratically with 65% acetonitrile from a (2–18 (aliphatic) column. Both peaks of antigen showed complete cross‐reactivity on FRIEP and absorbed at 220 nm. Amino acid analysis of each HPLC peak revealed no detectable differences in composition. Each peak contained a predominance of aspartic (13 mol%) and glutamic (18 mol%) acid. This antigen did not contain significant quantities of aromatic amino acids, and absorbed strongly at 206 nm. Neither the pl 4·0 or pl 9·0 antigen stained positively with the PAS reaction.