Application of enzyme-linked immunosorbent assay to detect antimicrobial peptides in human intestinal lumen

IF 1.6 4区 医学 Q4 BIOCHEMICAL RESEARCH METHODS
Julie S. Hong , Abrar Shamim , Hussein Atta , Eric B. Nonnecke , Sarah Merl , Satyajit Patwardhan , Elin Manell , Esad Gunes , Philip Jordache , Bryan Chen , Wuyuan Lu , Bo Shen , Beatrice Dionigi , Ravi P. Kiran , Megan Sykes , Emmanuel Zorn , Charles L. Bevins , Joshua Weiner
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引用次数: 0

Abstract

Intestinal transplantation is the definitive treatment for intestinal failure. However, tissue rejection and graft-versus-host disease are relatively common complications, necessitating aggressive immunosuppression that can itself pose further complications. Tracking intraluminal markers in ileal effluent from standard ileostomies may present a noninvasive and sensitive way to detect developing pathology within the intestinal graft. This would be an improvement compared to current assessments, which are limited by poor sensitivity and specificity, contributing to under or over-immunosuppression, respectively, and by the need for invasive biopsies. Herein, we report an approach to reproducibly analyze ileal fluid obtained through stoma sampling for antimicrobial peptide/protein concentrations, reasoning that these molecules may provide an assessment of intestinal homeostasis and levels of intestinal inflammation over time. Concentrations of lysozyme (LYZ), myeloperoxidase (MPO), calprotectin (S100A8/A9) and β-defensin 2 (DEFB2) were assessed using adaptations of commercially available enzyme-linked immunosorbent assays (ELISAs). The concentration of α-defensin 5 (DEFA5) was assessed using a newly developed sandwich ELISA. Our data support that with proper preparation of ileal effluent specimens, precise and replicable determination of antimicrobial peptide/protein concentrations can be achieved for each of these target molecules via ELISA. This approach may prove to be reliable as a clinically useful assessment of intestinal homeostasis over time for patients with ileostomies.

应用酶联免疫吸附试验检测人体肠腔中的抗菌肽
肠道移植是治疗肠道功能衰竭的最终方法。然而,组织排斥反应和移植物抗宿主疾病是相对常见的并发症,因此必须采取积极的免疫抑制措施,而免疫抑制本身又会带来更多并发症。跟踪标准回肠造口术中回肠流出物中的腔内标记物可能是检测肠移植物内病变发展的一种无创、灵敏的方法。与目前的评估方法相比,这种方法的灵敏度和特异性都很差,分别导致免疫抑制不足或过度,而且需要进行侵入性活检,因此这种方法是一种进步。在此,我们报告了一种可重复分析通过造口取样获得的回肠液中抗菌肽/蛋白质浓度的方法,认为这些分子可以评估肠道稳态和肠道炎症随时间变化的水平。使用市售的酶联免疫吸附测定法(ELISA)对溶菌酶(LYZ)、髓过氧化物酶(MPO)、钙粘蛋白(S100A8/A9)和β-防御素 2(DEFB2)的浓度进行了评估。α-防御素 5 (DEFA5) 的浓度是通过一种新开发的夹心酶联免疫吸附测定法进行评估的。我们的数据证明,只要正确制备回肠流出物标本,就能通过 ELISA 法精确、可重复地测定每种目标分子的抗菌肽/蛋白质浓度。这种方法可能被证明是可靠的,可作为回肠造口患者肠道长期稳态的临床有用评估方法。
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来源期刊
CiteScore
4.10
自引率
0.00%
发文量
120
审稿时长
3 months
期刊介绍: The Journal of Immunological Methods is devoted to covering techniques for: (1) Quantitating and detecting antibodies and/or antigens. (2) Purifying immunoglobulins, lymphokines and other molecules of the immune system. (3) Isolating antigens and other substances important in immunological processes. (4) Labelling antigens and antibodies. (5) Localizing antigens and/or antibodies in tissues and cells. (6) Detecting, and fractionating immunocompetent cells. (7) Assaying for cellular immunity. (8) Documenting cell-cell interactions. (9) Initiating immunity and unresponsiveness. (10) Transplanting tissues. (11) Studying items closely related to immunity such as complement, reticuloendothelial system and others. (12) Molecular techniques for studying immune cells and their receptors. (13) Imaging of the immune system. (14) Methods for production or their fragments in eukaryotic and prokaryotic cells. In addition the journal will publish articles on novel methods for analysing the organization, structure and expression of genes for immunologically important molecules such as immunoglobulins, T cell receptors and accessory molecules involved in antigen recognition, processing and presentation. Submitted full length manuscripts should describe new methods of broad applicability to immunology and not simply the application of an established method to a particular substance - although papers describing such applications may be considered for publication as a short Technical Note. Review articles will also be published by the Journal of Immunological Methods. In general these manuscripts are by solicitation however anyone interested in submitting a review can contact the Reviews Editor and provide an outline of the proposed review.
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