Department of Infection Biology, London School of Hygiene and Tropical Medicine, London, United Kingdom.
Department of Physiology and Biophysics, University of California, Irvine, Irvine, CA, United States of America.
PLoS One. 2022 Aug 29;17(8):e0273106. doi: 10.1371/journal.pone.0273106. eCollection 2022.
The evaluation of protein antigens as putative serologic biomarkers of infection has increasingly shifted to high-throughput, multiplex approaches such as the protein microarray. In vitro transcription/translation (IVTT) systems-a similarly high-throughput protein expression method-are already widely utilised in the production of protein microarrays, though purified recombinant proteins derived from more traditional whole cell based expression systems also play an important role in biomarker characterisation. Here we have performed a side-by-side comparison of antigen-matched protein targets from an IVTT and purified recombinant system, on the same protein microarray. The magnitude and range of antibody responses to purified recombinants was found to be greater than that of IVTT proteins, and responses between targets from different expression systems did not clearly correlate. However, responses between amino acid sequence-matched targets from each expression system were more closely correlated. Despite the lack of a clear correlation between antigen-matched targets produced in each expression system, our data indicate that protein microarrays produced using either method can be used confidently, in a context dependent manner, though care should be taken when comparing data derived from contrasting approaches.
作为感染的潜在血清学生物标志物的蛋白质抗原的评估已经越来越多地转向高通量、多重方法,如蛋白质微阵列。体外转录/翻译 (IVTT) 系统 - 一种同样高通量的蛋白质表达方法 - 已经广泛用于蛋白质微阵列的生产中,尽管来自更传统的全细胞表达系统的纯化重组蛋白在生物标志物表征中也发挥着重要作用。在这里,我们在同一蛋白质微阵列上对来自 IVTT 和纯化重组系统的匹配抗原的蛋白质靶标进行了并排比较。发现对纯化重组体的抗体反应的幅度和范围大于 IVTT 蛋白,并且来自不同表达系统的靶标之间的反应没有明显相关。然而,来自每个表达系统的氨基酸序列匹配靶标之间的反应更为密切相关。尽管在每个表达系统中产生的匹配抗原靶标之间没有明显的相关性,但我们的数据表明,使用两种方法之一制备的蛋白质微阵列可以在依赖于上下文的方式下被自信地使用,尽管在比较来自不同方法的数据时应小心谨慎。