Quantifying picomoles of analyte from less than 100 live bacteria: A novel method with a buffering hydrogel as an electrochemical cell.
Quantifying picomoles of analyte from less than 100 live bacteria: A novel method with a buffering hydrogel as an electrochemical cell.
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DOI：
10.1016/j.electacta.2023.143527
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发表时间：
2023-11
影响因子：
6.6
通讯作者：
Bronson Samel-Garloff;Subir Goswami;Ankan Ghosh;Jens Kreth;Dipankar Koley
中科院分区：
文献类型：
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作者：
Bronson Samel-Garloff;Subir Goswami;Ankan Ghosh;Jens Kreth;Dipankar Koley
细菌细胞化学环境的微环境变化可能会对生物膜的生态学产生深远的影响。然而，从极少量的细菌中定量分析物的皮摩尔总量是一个分析挑战。在这里，我们提供了一种新的微升体积的水凝胶为基础的电化学电池平台，适合库仑测量过氧化氢（H2 O2）产生的少于100个细胞的血链球菌，健康的口腔微生物组的相关成员。在水凝胶的聚合物结构中引入了一个吗啉部分，以在生物pH下创造一个受控的微环境。我们使用一种新的缓冲水凝胶的方法计算了这种水凝胶在pH 7.2-6.2范围内的缓冲容量为0.257±0.135 mol HNO 3 mol ME A× Δ pH。在pH值为7.0-3.0的范围内，H2 O2传感器的灵敏度没有变化，可用于H2 O2对S.血液不依赖于它们产生的任何酸。该新平台能够测量不到100个细菌细胞产生的22.7±3.5 pmol H2 O2，这在大型溶液分析中是无法实现的。这些发现表明，这是一个合适的平台，用于定量来自亚毫克生物样品的代谢物，甚至可能适用于直接分析原始生物膜样品，几乎没有样品预处理。
Microenvironmental changes in the chemical surrounding of bacterial cells might have a profound impact on the ecology of biofilms. However, quantifying total amount of picomoles of analyte from a miniscule number of bacteria is an analytical challenge. Here we provide a novel microliter volume hydrogel based electrochemical cell platform suitable of coulometrically measuring hydrogen peroxide (H 2 O 2) produced by less than 100 cells of Streptococcus sanguinis, a relevant member of the healthy oral microbiome. A morpholine moiety was incorporated into the polymer structure of the hydrogel to create a controlled microenvironment at biological pH. We calculated the buffering capacity of this hydrogel as 0.257±0.135 m o l H N O 3 m o l M E A× Δ p H over the pH range of 7.2–6.2 by using a novel method designed for buffering hydrogels. The H 2 O 2 sensors coated in microliter volume of buffering hydrogel showed no change in sensitivity within the pH range of 7.0–3.0, allowing for H 2 O 2 measurements of S. sanguinis independent of any acid they produce. The novel platform was able to measure down to 22.7±3.5 pmol H 2 O 2 produced by less than 100 bacterial cells, which would otherwise not be attainable in large solution-based assays. These findings indicate that this is a suitable platform for quantifying metabolites from sub-milligram biological samples and may even be suitable for direct analysis of raw biofilms samples with little to no sample pretreatment.