Reaching the Next Level of Sensitivity in Oligonucleotide Bioanalysis: A Hybridization-Based Method on Quanterix Simoa®

Simoa®, Single Molecule Array technology, is known and recognized as an ultra-sensitive immunoassay technology that can detect extremely low level of proteins and nucleic acids that traditional ligand binding assay cannot reach. Simoa® is well established in biomarker realm, where the gains in sensitivity have allowed, for example, the detection of biomarkers of brain injury and neurological diseases at a much earlier stage in more accessible matrices such as blood, serum or plasma, allowing more frequent and less invasive sampling. In the area of pharmacokinetic (PK) testing in the regulated bioanalytical laboratory however, Simoa® technology is still relatively new.
The power of single molecule detection technology brought a great opportunity to meet the challenge of detecting oligonucleotide therapeutics with the sensitivity required to fully evaluate their pharmacokinetics. In this presentation, a case study will be presented where a hybridization assay was developed on the Simoa®. Challenges faced during method establishment on this ultra-sensitive immunoassay technology will be discussed, such as labeling of paramagnetic beads, assay reproducibility and precision, stability of conjugated paramagnetic beads, along with approaches taken to overcome and/or mitigate. To evaluate the gains in sensitivity, the Simoa® assay will be compared to a more conventional hybridization assay approach using electrochemiluminescence (ECL) for detection of the same oligonucleotide therapeutic and the limits of quantitation achieved.