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Table 1 The characteristics of the biomarkers with their corresponding isolation techniques, clinical applications and limitations

From: Integration of liquid biopsy and pharmacogenomics for precision therapy of EGFR mutant and resistant lung cancers

Biomarkers

Isolation technique

Clinical application

Limitations

References

Circulating tumor cells (CTCs)

Immunomagnetic enrichment

â–ª Prognosis

â–ª Treatment

â–ª Reproducibility

â–ª Sensitivity

â–ª CTC without epithelial marker could not be detected

â–ª Difficult to use with whole blood

â–ª Low purity of detected CTC

â–ª Cannot process whole blood

â–ª High detection cost

[86]

Immunomagnetic isolation

â–ª Diagnosis

â–ª Prognosis

â–ª Reliance on EpCAM and CK

â–ª Variation of expression of EpCAM and CK across cancers

â–ª Lack of selectivity

â–ª High detection cost

[87]

Magnetic beads

â–ª Prognosis

â–ª Treatment

â–ª Increased contamination with WBC

â–ª Requires more blood

[88]

Microfluidic positive immunocapture

(CTC-chip)

â–ª Prognosis

â–ª Diagnosis

â–ª Shear force might affect cell viability and attachment

â–ª Slow rate processing

â–ª Limited volume

[89]

Size based separation

(filter-based isolation)

â–ª Prognosis

â–ª Treatment

â–ª Prone to clogging

â–ª Requires high volume of blood

â–ª Sample may be adulterated

[90]

Density gradient separation

â–ª Prognosis

â–ª Treatment

â–ª Loss of large CTC and cell aggregates

â–ª Low purity

[91]

Inertial focusing

â–ª Prognosis

â–ª Diagnosis

â–ª Treatment

â–ª Morphological deformation of the captured cell

[92]

Single cell sequencing

â–ª Prognosis

â–ª Treatment

â–ª Poor reproducibility

â–ª False positives and false negatives

â–ª Allele deletion

â–ª Sequencing errors

[93]

Circulating tumor DNA (ctDNA)

Manual (DNA purification)

â–ª Detection

â–ª Prognosis

â–ª Treatment initiation and monitoring

â–ª Low accuracy and precision

[94]

Automated (ccfDNA purification)

â–ª Requires adequate control for downstream application

â–ª Only for use with plasma prepared from human whole blood samples collected in EDTA tube

â–ª Not for use in diagnostic procedures

ctRNA (miRNA, lncRNA)

Northern blot analysis

â–ª Diagnosis

â–ª mRNA degradation during electrophoresis

â–ª Low sensitivity

â–ª Detection with multiple probes is difficult

[95]

Microarray

â–ª Diagnosis

â–ª Therapeutic response prediction

â–ª Standardization and optimization

â–ª Low specificity

â–ª Low reproducibility

â–ª High cost of a single experiment

â–ª Unsuitable for clinical experiences

[96]

RT-qPCR

â–ª Diagnosis

â–ª Treatment initiation and

monitoring

â–ª Amplification bias

â–ª No template controls

â–ª Cannot perform multiple detection

[97]

Liquid chip technology

â–ª Diagnosis

â–ª Prognosis

â–ª Treatment

â–ª Resistance monitoring

â–ª In vivo validation

â–ª Difficult to scale up

[88]

Exosomes

Ultracentrifugation-based

â–ª Early diagnosis

â–ª Prognosis

â–ª Contamination and exosome loss

â–ª Low recovery

â–ª Laborious

[98]

Size-based

â–ª Diagnosis

â–ª Deformation of EVs

â–ª High risk of chip clogging

â–ª Long run time

[99]

Immunoaffinity capture-based

â–ª Diagnosis

â–ª Antibody cross reactivity

â–ª Possible detection of non-EV particles

â–ª Only exosomes with targeted proteins can be separated

â–ª Low yield

â–ª Tumor heterogeneity hinders immune recognition

â–ª Time consuming

â–ª Expensive

[98]

Microfluidics based

â–ª Diagnosis

â–ª Lack of standardization and method validation

â–ª Moderate to low sample capacity

[100]

Tumor-educated platelets (TEPs)

Spliced TEP mRNA

â–ª Diagnosis

â–ª Treatment monitoring

â–ª Complex isolation technique

â–ª Fragility of TEPs

[101, 102]

  1. Abbreviations: CTCs Circulating Tumor Cells, EpCAM Epithelial cell adhesion molecule, CK Cytokeratin, WBC White blood cell, ctDNA Circulating tumor DNA, ccfDNA Circulating cell free DNA, EDTA Ethylenediamine tetraacetic acid, ctRNA Circulating tumor RNA, miRNA MicroRNA, lncRNA Long non-coding RNA, mRNA Messenger RNA, RT-qPCR Quantitative real-time polymerase chain reaction, EVs Extracellular vesicles, TEPs Tumor-educated platelets