Measuring antimicrobial peptide activity on epithelial surfaces in cell culture.

Measuring antimicrobial peptide activity on epithelial surfaces in cell culture.
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DOI:
10.1007/978-1-60761-594-1_23
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发表时间:
2010
期刊:
Methods in molecular biology (Clifton, N.J.)
影响因子:
--
通讯作者:
McMahon, Laura
McMahon, Laura
中科院分区:
其他
文献类型:
--
作者:
Diamond, Gill;Yim, Sunghan;Rigo, Isaura;McMahon, Laura

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为了更准确地评估上皮细胞衍生的抗微生物肽在其天然环境中的活性和作用,必须在相关条件下在表面进行测定。为了实现这一点,我们利用三维培养的气道和牙龈上皮细胞,这是生长在气液界面。在这些条件下,可以用已知引起抗微生物肽基因表达增加的多种因子对培养物进行攻击。然后可以通过直接在细胞表面上或使用分泌到培养物顶端表面上的流体定量抗菌活性来评估这种诱导的功能相关性。肽的相对贡献也可以通过将分泌的流体与特异性抑制性抗体预孵育来测量。因此,一个相对便宜的体外模型可用于评估抗菌肽在粘膜上皮中的作用。
To more accurately assess the activity and role of epithelial-cell derived antimicrobial peptides in their native settings, it is essential to perform assays at the surfaces under relevant conditions. In order to carry this out, we utilize 3-dimensional cultures of airway and gingival epithelium, which are grown at an air-liquid interface. Under these conditions, the cultures can be subjected to challenge with a variety of factors known to cause an increase in antimicrobial peptide gene expression. The functional relevance of this induction can then be assessed by quantifying antibacterial activity either directly on the surface of the cells or using the fluid secreted onto the apical surface of the cultures. The relative contribution of the peptides can also be measured by pre-incubation of the secreted fluid with specific inhibitory antibodies. Thus, a relatively inexpensive in vitro model can be used to evaluate the role of antimicrobial peptides in mucosal epithelium.
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