Table 2.

Summary of pros and cons of MSI testing in tissue and liquid biopsies.

Type	Advantages	Disadvantages
Tissue biopsy	Clinically validated; Gold standard for MSI detection (IHC and PCR); Potential CRC prognosis based on MSI status (especially during early stages of CRC); Allow prediction of the outcomes/overall survival of MSI-related CRC patients; Potential management decisions and prognostication; Allow distinct clinicopathological features of MSI-related CRC.	Not incorporated into the routine analysis; Invasive with high risk; Inability to interrogate full tumor load’s heterogeneity; Difficulties in repeated sampling; Lack of viable tissue and not routinely available; Unavailability/infeasibility in certain patients; Requirement of an adequate amount of tissue with a high concentration of cancer cells (paired with normal tissues) for PCR-based MSI detection; The need of specialized laboratory equipment (PCR) and professional expertise (e.g., pathologist) (IHC); Low sensitive for samples with low proportions of cancer cells; Impractical for periodic/real-time monitoring of cancer progression and treatment response.
Liquid biopsy	Rapid detection; High specificity; Non-invasive procedures and minimal risk; High concordance rate with tissue biopsy-based detection; Potential to monitor genetic heterogeneity; Ability to capture the mutational landscape of CRC patients; Capable of capturing TMB; Identification of rare MSI frameshift alleles; High repeatability and easily reproducible; Potential to enhance utility of tumor detection assays to help direct clinicians beyond targeted therapies to include immunotherapies; Possible to perform continuous follow up examinations.	Still in early development without established clinical practice rules; Lack of sophisticated multicenter clinical validation studies and regulatory guidelines; Unstandardized laboratory procedures; Limited diagnostic accuracy and sensitivity; Low detection rate of mutations; Inability to reflect ITH; Great technical and bioinformatics challenges (inefficient molecular capture, sequencing, mapping, variant calling, error correction at MSI loci); Low tumor fraction in circulation; High level of technical noise due to polymerase slippage; Low signal-to-noise ratio (presence of contaminating non-tumor cells such as hematopoietic cells, immune cells and blood stromal cells); Risk of false-positive and false-negative results; Microenvironment changes may influence the release or the amount of biological materials.

CRC = colorectal cancer, IHC = immunohistochemistry, ITH = intratumoral heterogeneity, MSI = microsatellite instability, PCR = polymerase chain reaction, TMB = tumor mutation burden.