Lundkvist A, Björsten S, Niklasson B, Ahlborg N
Department of Defense Microbiology, Swedish Institute for Infectious Disease Control, Stockholm.
Clin Diagn Lab Immunol. 1995 Jan;2(1):82-6. doi: 10.1128/cdli.2.1.82-86.1995.
The complete amino acid sequence of the Puumala (PUU) virus nucleocapsid protein (N), deduced from the genome of the prototype strain Sotkamo, was synthesized as decapeptides with 5-amino-acid overlaps. By use of the PEPSCAN method, 86 peptides were examined for reactivity with sera from serologically confirmed nephropathia epidemica (NE) patients and 11 PUU virus N-specific bank vole monoclonal antibodies. The human sera showed reactivity with several different regions, while only one of the monoclonal antibodies reacted with one single peptide. Sequences were selected by this PEPSCAN analysis of human antibody reactivities, and five 15-amino-acid peptides were synthesized and evaluated as antigens by an enzyme-linked immunosorbent assay (ELISA). Peptide-reactive antibodies of the immunoglobulin M (IgM) class were measured in serum samples drawn from patients with acute NE. In comparison with the results of a mu-capture IgM ELISA using native PUU virus antigen, only a few serum samples were found positive (sensitivity, 2 to 10%). Interestingly, when antibodies of the IgG class were measured, the sensitivities of the five peptide ELISAs were found to be 79, 46, 2, 100, and 40%, respectively, as compared with the sensitivity of an IgG ELISA based on native viral antigen. The IgG reactivities of sequentially drawn sera from NE patients with the two peptides giving the highest assay sensitivities were analyzed and compared with their reactivities with native viral antigen. All patients had detectable anti-peptide IgG in the acute-phase sample, which, however, had totally declined in samples drawn after 2 years. The opposite pattern was seen with native viral antigen, in which case all patients showed the highest levels of specific IgG after 2 years. The results suggest the presence of epitopes specific for the acute IgG response.
从普马拉（PUU）病毒原型株索特卡莫基因组推导出来的核衣壳蛋白（N）的完整氨基酸序列，被合成为具有5个氨基酸重叠的十肽。通过使用PEPSCAN方法，检测了86种肽与血清学确诊的流行性肾病（NE）患者血清以及11种PUU病毒N特异性田鼠单克隆抗体的反应性。人血清与几个不同区域有反应，而只有一种单克隆抗体与一个单一肽发生反应。通过对人抗体反应性的这种PEPSCAN分析选择序列，并合成了5种15个氨基酸的肽，并通过酶联免疫吸附测定（ELISA）评估其作为抗原的性能。在急性NE患者采集的血清样本中检测免疫球蛋白M（IgM）类的肽反应性抗体。与使用天然PUU病毒抗原的μ捕获IgM ELISA结果相比，仅发现少数血清样本呈阳性（敏感性为2%至10%）。有趣的是，当检测IgG类抗体时，与基于天然病毒抗原的IgG ELISA敏感性相比，5种肽ELISA的敏感性分别为79%、46%、2%、100%和40%。分析了NE患者连续采集的血清与两种测定敏感性最高的肽的IgG反应性，并将其与它们与天然病毒抗原的反应性进行比较。所有患者在急性期样本中均检测到抗肽IgG，但在2年后采集的样本中其已完全下降。天然病毒抗原则呈现相反的模式，在这种情况下，所有患者在2年后显示出最高水平的特异性IgG。结果表明存在针对急性IgG反应的表位。
