Isolation of an Aptamer that Binds Specifically to E. coli

Abstract

Escherichia coli is a bacterial species found ubiquitously in the intestinal flora of animals, although pathogenic variants cause major public health problems. Aptamers are short oligonucleotides that bind to targets with high affinity and specificity, and have great potential for use in diagnostics and therapy. We used cell-based Systematic Evolution of Ligands by EXponential enrichment (cell-SELEX) to isolate four single stranded DNA (ssDNA) aptamers that bind strongly to E. coli cells (ATCC generic strain 25922), with Kd values in the nanomolar range. Fluorescently labeled aptamers label the surface of E. coli cells, as viewed by fluorescent microscopy. Specificity tests with twelve different bacterial species showed that one of the aptamers–called P12-31—is highly specific for E. coli. Importantly, this aptamer binds to Meningitis/sepsis associated E. coli (MNEC) clinical isolates, and is the first aptamer described with potential for use in the diagnosis of MNEC-borne pathologies.

High Efficiency Binding Aptamers for a Wide Range of Bacterial Sepsis Agents

“…For the aptamer binding assay we used the method as previously described by Marton et al [13]. Bacterial cells were washed twice with phosphate-buffered saline (PBS) (137 mM NaCl and 0.05% BSA).…”

Aptamer-Based Biosensing with a Cationic AIEgen

“…[1,2] As each aptamer assumes distinct secondary and tertiary conformations, they possess high binding affinity and specificity to a wide variety of ligands, including metal ions, [3][4][5] small molecules, [6][7][8] peptides, [9,10] proteins, [11][12][13] and even microorganisms. [14][15][16] Compared with monoclonal antibodies, aptamers offer similar binding profiles with many additional advantages such as inexpensive synthesis, better chemical and thermal stability, reusability, improved tissue penetration, and low immunogenicity. [17] Indeed, quite a few aptamer-based fluorescent sensors have been described.…”

“…[17] Indeed, quite a few aptamer-based fluorescent sensors have been described. [8,16,[18][19][20][21] However, these sensors generally incorporate conventional dyes that are vulnerable to fluorescence quenching when they form aggregates at overly high concentration. This phenomenon, known as 'aggregationcaused quenching' (ACQ), is a notorious issue that restricts the usefulness of conventional dyes in biomolecular applications.…”

Rapid Selection of Single-Stranded DNA Aptamers Binding Staphylococcus Epidermidis in Platelet Concentrates

“…A method called Systematic Evolution of Ligands by Exponential (SELEX) [15] has been used to select ssDNA aptamers that bind to bacteria of different species [16,17]. For example, whole-cell SELEX was conducted to select and characterize aptamers binding to Salmonella typhimurium, E. coli and S. aureus [3,1820]. The combined use of multiple aptamers increased sensitivity for detection of whole S. aureus cells [20].…”