Aptamer-based trapping of phytosphingosine in urine samples

Area of Science:

• Biotechnology
• Analytical Chemistry
• Molecular Diagnostics

Background:

• Quantifying small molecules like metabolites for clinical diagnostics typically relies on instrumental methods (e.g., LC-MS) or bioanalytical techniques.
• Aptamers, as selective receptors, offer an alternative or complementary approach to antibodies for small molecule detection.

Purpose of the Study:

• To generate aptamers with high affinity for the medically relevant metabolite phytosphingosine.
• To demonstrate the feasibility of the 'just in time'-Selection approach for selecting aptamers against small molecules.
• To showcase the application of aptamers for affinity enrichment of phytosphingosine.

Main Methods:

• Generation of aptamers targeting phytosphingosine using the established 'just in time'-Selection protocol.
• Immobilization of selected aptamers onto magnetic particles for affinity-based separation.
• Application of aptamer-functionalized magnetic particles for enrichment of phytosphingosine in selection buffer and urine samples.

Main Results:

• Successfully generated aptamers with dissociation constants in the low nanomolar range for phytosphingosine.
• Achieved significant enrichment factors for phytosphingosine: up to 9-fold in selection buffer and 4-fold in urine samples.
• Demonstrated the proof-of-concept for extending the 'just in time'-Selection method to small molecule aptamer selection.

Conclusions:

• The 'just in time'-Selection approach is effective for generating high-affinity aptamers against small molecules like phytosphingosine.
• Aptamers can be utilized for efficient affinity enrichment, improving the pre-analytical processing of samples for metabolite analysis.
• This methodology holds potential for adaptation to other small molecule targets in diagnostics and bioanalysis.