Unique molecular indices (UMI) are a great help in this research, says Labarrière. UMIs are molecular barcodes that are ligated to the ends of nucleic acid fragments prior to sequencing. Tagging DNA and RNA with UMIs before any amplification takes place allows reads to be assigned to individual molecules, allowing for a computational correction of amplification bias and sequencing errors. “With UMI we can detect low-frequency variants, for instance, to identify T-cell receptor (TCR) repertoire variations in melanoma patients responding to immunotherapy with anti-PD-1, and to characterize immune cell subsets associated with therapeutic response.”
She uses UMIs from the QIAseq Immune Repertoire RNA Library Kit and the QIAseq UPX3' Targeted RNA Panel to eliminate PCR bias and to improve accuracy in variant detection and estimation of the allelic fraction and allele frequency, saying that she gets a “more accurate quantification of the transcript count and reduced technical noise” this way.
Accuracy is important as the immune system for every patient is unique and needs to be genetically defined. There are more than 10,000 different types of T-cells found in blood, all of which have different functions. Of these 10,000-plus T-cells, one must be identified that will react to a tumor’s specific genetic profile—comparable to finding a needle in the proverbial haystack.