Advanced diseases diagnostic platform using cost-affordable functional biomaterials

Abstract

Background A rapid and accurate disease diagnosis based on identifying cancer and emerging infectious diseases is essential for preventing further infection, monitoring patients, and proper treatment. In particular, since individual symptom patterns are different depending on the severity of cancer mutations and infections, it is necessary to accurately diagnose diseases and proceed with treatment optimized for each patient. Recently, various diagnostic methods were reported for better diagnostics, but most of these techniques are time-consuming, complex, require large equipment, low sensitivity, and lack multiple-target detection specificity. To overcome these limitations, we reported sample preparation platform using cost-affordable functional biomaterials and advanced bio-optical sensing platform.

Method In Section 1, sample preparation platform using cost-affordable functional biomaterials, we reported a rapid and simple cfNAs sampling using bio-composite and microfluidic platform, and pathogen enrichment/NAs extraction using microfluidic platform. We analyzed a total of 366 clinical samples for sample preparation; cfDNA sampling using 13 blood plasma samples consisting of 3 colorectal and 10 pancreatic cancers in bio-composite platform, cfDNA sampling using 81 blood plasma samples consisting of 34 confirmed TB, 6 probable TB, and 41 not TB, and pathogen enrichment/DNAs extraction using 272 oral swab samples consisting of 128 TB and 144 not TB. In Section 2, advanced bio-optical sensing platform, we reported SMR biosensor based on an isothermal, label-free, one-step DNA/RNA amplification and detection, improved bio-optical sensing platform by combining CRISPR/dCas9, and novel arch-shaped multiple-target sensing. We analyzed a total of 61 clinical samples for bio-optical sensing; 35 blood plasma samples consisting of 16 Q fever and 19 other febrile diseases patients in SMR biosensor, 6 blood plasma samples consisting of 3 ST and 3 SFTS patients in CRISPR/dCas9-mediated biosensor, and 20 nasopharyngeal samples consisting of 11 MERS-CoV and 9 HCoV patients.

Result In sample preparation, the bio-composite platform showed high capture efficiency (86.78–90.26%) with genomic DNA and amplified DNA products (777, 525 and 150 bp). The bio-composite platform allowed the isolation of high purity and quantity cfDNA from the blood plasma of 13 cancer patients (3 colorectal and 10 pancreatic cancer samples) without requiring a lysis step and special equipment. The cfDNA sampling in microfluidic platform showed high sensitivity (80.0%) and specificity (78.1%) using the blood plasma of 34 confirmed TB, 6 probable TB, and 41 not TB. These results showed that the sensitivity of cfDNA is higher than acid-fast bacilli microscopy (31.6%), Xpert MTB/RIF (61.1%), and mycobacterial culture (65.8%) results. While the sensitivity and specificity of cfDNA were similar with those of IGRA (sensitivity 80.6% and specificity 71.4%), the combined sensitivity and specificity of the two assays was 94.4% and 64.3%, respectively, which can be used to rule out TB. The pathogen enrichment/DNA extraction showed high sensitivity (65.6%) and specificity (86.1%) using the oral swab of 128 TB and 144 not TB. These results showed that the sensitivity of DNA is higher than sputum-based Xpert MTB/RIF (sensitivity 43.4% and specificity 100%). In bio-optical sensing, the SMR biosensor showed high sensitivity (87.5%) and specificity (89.5%) using the blood plasma of 16 Q fever and 19 other febrile diseases within 10 min. By using CRISPR/dCas9-mediated biosensor, these results showed single molecule sensitivity for the detection of ST (0.54 aM) and SFTS (0.63 aM); this sensitivity is 100 times more sensitive than that of real-time PCR. The CRISPR/dCas9-mediated biosensor allowed the clearly distinguish between ST and SFTS in plasma samples within 20 min. The arch-shaped multiple-target biosensor showed high sensitivity with T7 RNA of MERS (10 copies), ZIKV (100 copies), EBOV (10 copies), and HCoV (10 copies) and simultaneously distinguished between Middle East respiratory syndrome and human coronavirus in clinical specimens within 20 min.

Conclusion In this study, advanced diseases diagnostic platform using cost-affordable functional biomaterials was proposed. This platform consists of two sections of sample preparation using bio-composite and microfluidic chip, and bio-optical sensing using SMR biosensor, CRISPR/dCas9-mediated biosensor, and arch-shaped multiple-target biosensor. We expect that these techniques can provide rapid, sensitive, and accurate diagnoses of cancer and emerging infectious diseases in clinical applications.