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The term liquid biopsy (LB) refers to the study of circulating tumor cells, circulating tumors nucleic acids free of cells or contained in exosomes, and information about platelets associated with tumors. LB can be performed in different biofluids and allows the limitations of tissue biopsy to be overcome offering possibilities of tumor identification reflecting in real time tumor heterogeneity. In addition, LB allows screening and early detection of cancer, real-time monitoring of therapy, stratification and therapeutic intervention, a therapeutic target and resistance mechanism, and a risk of metastatic relapse.
Molecular Detection Techniques | Properties |
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quantitative polymerase chain reaction (qPCR) |
qPCR, is a laboratory technique of molecular biology based on the polymerase chain reaction (PCR). qPCR is regarded as the ‘gold standard’ in the quantitative analysis of nucleic acids, be it DNA, RNA or micro-RNA molecules. qPCR has high sensitivity, robustness, good reproducibility, broad dynamic quantification range, and very importantly, affordability. |
Safe-Sequencing System (Safe-SeqS) |
Safe-SeqS is a Unique Molecular Identifier (UMI) approach to detect rare variants. Safe-SeqS assigns a UMI to each template molecule and amplifies each uniquely tagged template molecule to create UMI families. The abundance of each UMI can be used to distinguish between rare mutations and technical errors, as well as to correct for PCR amplification bias |
CAncer Personalized Profiling by deep Sequencing (CAPP-Seq) |
CAPP-Seq is a next-generation sequencing based method used to quantify circulating DNA in cancer. CAPP-Seq is an economical and ultrasensitive method and could be routinely applied clinically to detect and monitor diverse malignancies, thus facilitating personalized cancer therapy. |
Digital PCR (dPCR) |
dPCR is a modification of the qPCR method that can be employed to quantify precisely defined nucleic acid targets. The technique is based on the concept of limiting dilutions, which involves the partitioning of a PCR reaction into multiple sub-reactions such that each sub-reaction either contains none or one or more DNA targets. Following thermal cycling, reactions are classified as either positive (target detected) or negative (no target detected), hence providing the basis for a digital output format. By determining the proportion of empty partitions, Poissonian statistics can be applied and the initial number of target molecules present can be estimated. |
Copy number alterations (CNAs) |
CNA are somatic changes in chromosome structure that result in gains or losses in copies of DNA sections in somatic tissue, and are prevalent in many cancers. CNA has facilitated the discovery of tumor suppressor genes and oncogenes. Microarray-based CNA assays designed to detect these chromosomes copy number alterations on a high-resolution, genome-wide scale have emerged as a key technology in the genomic era. |
Whole-genome sequencing (WGS) |
WGS is a comprehensive method for analyzing entire genomes. Whole genome sequencing WGS has revolutionized the biosciences and proven to be essential and invaluable to the identification of gene functions and their involvement in disease. The feasibility of WGS analysis is under the support of next generation sequencing (NGS) technologies, which require substantial computational and biomedical resources to acquire and analyze large and complex sequence data. |
Whole-exome sequencing (WES) |
WES is a genomic technique for sequencing all of the protein-coding regions of genes in a genome (known as the exome). WES provides coverage of more than 95% of the exons, which harbor the majority of the large genetic variants and single nucleotide polymorphisms (SNPs) associated with human disease phenotypes. WES strategy starts by narrowing down the details of variants to be studied by filtering against databases such as HapMap, from the approximately 3.5 million SNPs identified in the human genome project. This focus enables a simpler way for discovery and validation of causative genes and common and rare variants. |
Tagged-Amplicon deep sequencing (TAm-Seq) |
TAM-Seq allows targeted sequencing of entire genes to detect mutations in ctDNA. TAM-Seq is based on a multiplex pre-amplification of tiling short amplicons with target-specific primers and initial eminent of the target regions is performed followed by a selective amplification in individual (singleplex) PCRs in order to exclude non-specific products |