Non-SELEX method for aptamer selection against β-casomorphin-7 peptide.

Area of Science:

• Biotechnology
• Molecular Biology
• Biosensor Development

Background:

• β-casomorphin-7 (BCM-7) peptide detection is crucial for understanding certain health conditions.
• Conventional SELEX for aptamer selection is time-consuming and complex.
• Development of rapid and efficient aptamer selection methods is needed.

Purpose of the Study:

• To develop and evaluate a non-systematic evolution of ligands by exponential enrichment (non-SELEX) method for selecting BCM-7 specific aptamers.
• To assess the efficacy of selected aptamers in detecting BCM-7 peptide in human urine samples.
• To explore the potential of these aptamers in developing biosensors for autistic patient diagnostics.

Main Methods:

• Employed a modified non-SELEX approach without aptamer amplification or counterselection, including a negative selection round.
• Performed aptamer selection in a single day, identifying 16 aptamers after 5 rounds via Sanger sequencing.
• Utilized in silico analysis, gel electrophoresis, and colorimetric assays on magnetic beads and membranes for aptamer screening and validation.

Main Results:

• Identified Sequence ID no. 3 as a high-performing aptamer for BCM-7 detection, outperforming previously selected SELEX aptamers.
• Demonstrated successful detection of low amounts of BCM-7 spiked in human urine samples across various sensing platforms.
• Validated the non-SELEX method as an efficient and time-saving process for aptamer selection.

Conclusions:

• The non-SELEX method provides a rapid and effective strategy for aptamer selection.
• Selected aptamers, particularly Sequence ID no. 3, show significant potential for developing sensitive BCM-7 detection biosensors.
• These aptamers could aid in the diagnosis of conditions like autism by detecting BCM-7 peptide in urine.