DOI: 10.5061/dryad.k98sf7mdd
title: Data for: Sensitive and specific detection of tumor-derived exosomes using nanopore-crystal microchips

description: Tumor-derived exosomes (tExos) have emerged as promising circulating
 biomarkers for early cancer diagnosis. However, the sensitivity and
 specificity of existing assays often limit the clinical translation of
 tExos. In this study, we present a highly versatile microfluidic platform,
 termed the nanopore-crystal microchip (NC-Chip), for specific isolation
 and ultrasensitive detection of tExos in as little as 0.5 μL of plasma
 samples from pancreatic cancer (PC) patients. The NC-Chip incorporates a
 herringbone-patterned hierarchical porous hydrogel scaffold, enabling
 fluid manipulation, size exclusion, immunoaffinity capture, and signal
 amplification. These integrated features significantly enhance the
 sensitivity and specificity of tExos assays in complex clinical scenarios.
 Utilizing an eight-protein signature, the NC-Chip can distinguish patients
 with pancreatitis and non-metastatic PC with 100% accuracy in the training
 cohort and 94.9% accuracy in the validation cohort. This platform is
 sensitive, specific, inexpensive, and only needs small-volume samples,
 offering a powerful exosome-based liquid biopsy tool for early PC
 diagnosis.
description: The mean, SD, and LOD were computed utilizing established
 standard formulas. A student’s t-test with two tails was employed to
 assess significance. The intensities of individual protein markers
 measured by the NC-Chip were normalized using Min-Max. The normalized
 intensities of eight protein markers were weighted together to create the
 PC signature by LDA. The nonparametric, two-tailed Mann-Whitney
 <em>U</em> test for binary classification was used to
 determine <em>P</em> values for pairwise comparisons. With a
 post-hoc Dunn’s test for pairwise multiple comparisons, a Kruskal-Wallis
 one-way ANOVA was used to determine the overall and group pair
 <em>P</em> values for ternary classification. To evaluate the
 AUC, sensitivity, specificity, and accuracy of PC diagnosis, ROC analyses
 were developed for individual markers or marker combinations. The
 discriminant function model was initially constructed using data from the
 training cohort, and then it was applied to classify patients in the
 validation cohort. The validation was conducted using non-blinded samples.
 The validation cohort’s sensitivity, specificity, and accuracy were
 evaluated after the optimal cutoff values were selected based on the
 training cohort using Youden’s index. Using OriginPro 2018, GraphPad Prism
 (v.8.0), and R software (version 4.1.2), all statistical analyses were
 performed with 95% confidence intervals (<em>P </em>&lt;
 0.05).
description: # README:Sensitive and specific detection of plasma tumor-derived exosomes
 using nanopore-crystal microchips Description of the data and file
 structure (1) Fig.3e-Fluorescence_enhancement_factor.csv This document is
 the enhancement factor of fluorescent FDG, Cyanine3, and Tex-red inside
 Flat-Chip, Solid-Chip, and NC-Chip. Before use, the FDG solution at FDG:
 β-Gal ratio of 1:1 was created and reacted for 30 min at 37 °C in the
 dark. The enhancement factor was calculated by ∆E,exp = (IE−Ix)/(I0−Ix),
 where IE represents the intensity on the NC-Chip or Solid-Chip on the
 herringbone pattern, I0 represents the intensity on the Flat-Chip channel,
 and Ix represents the background correction. The FDG represents
 fluorescein-di-β-d-galactopyranoside. The β-Gal represents
 β-galactosidase. The NC-Chip represents nanopore-crystal microchip. The
 Solid-Chip represents solid herringbone chip. The Flat-Chip represents an
 assay chip on clean glass slides. (2)
 Fig.3f-Simulated_reflection_spectra.csv This document is the simulated
 reflection spectra of NC-Chip excited by plane light. The N represents the
 layer of nanopore-crystal structure. (3)
 Fig.3h-Simulated_fluorescence_emission_power_curves.csv This document is
 the simulated fluorescence emission power curves of a dipole on a glass
 substrate (Flat chip) and nanopore-crystal structure (NC-Chip). A single
 dipole was employed to simulate the generated fluorescence of FDG. (4)
 Fig.4d-Capture_efficiency_of_anti-CD81-based_NC-Chip_and_Solid-Chip.csv
 This document is the comparison capture efficiency of anti-CD81-based
 NC-Chip and Solid-Chip using DiO-stained PANC-1 Exo (1.0×109 particles
 mL-1) in PBS and extracellular vesicles-depleted plasma (100-fold
 dilution). The anti-CD81-based NC-Chip and Solid-Chip represents the
 NC-Chip and Solid-Chip that was modified by CD81 monoclonal antibodies.
 The DiO represents 3,3'-dioctadecyloxacarbocyanine perchlorate. The
 PANC-1 Exo represents PANC-1 cell-derived exosomes. (5)
 Fig.4e-Detectionand_Solid-Chip_dynamic_range_for_quantifying_NCL_expression on_PANC-1_Exo.csv This document is the detection dynamic range for quantifying nucleolin (NCL) expression on PANC-1 Exo by the NC-Chip and Solid-Chip. The c represents the concentration of exosomes. (6) Fig.4f-NC-Chip_for_detection_of_NCL_expression.csv This document is the NC-Chip for detection of NCL expression on hTERT-HPNE cell-derived exosomes (hTERT-HPNE Exo) and PANC-1 Exo (1.0 × 109 particles mL-1) spiked in both PBS and 100-fold diluted extracellular vesicles (EV)-depleted plasma. Control experiment for PBS or 100-fold diluted EV-depleted plasma. (7) Fig.4g-Calibration_curves_for_detection_of_NCL_expression.csv This document is the calibration curves for detecting the NCL expression on PANC-1 Exo spiked in PBS and 100-fold diluted EV-depleted plasma. (8) Fig.4h-Radar_plot_showing \_expression_of *eight* proteins_from_one_normal_and \_seven_tumor_cell_lines.csv This document is the radar plot showing expression of eight proteins from one normal hTERT-HPNE- and seven tumor PC cell lines (CFPAC-1-, PANC-1-, MIAPaCa-2-, SW1990, Panc 08.13, PaTu 8988t, and BxPC-3)-derived exosomes. (9) Fig.4i-1-Correlation_of_expression_of_eight_proteins_on_unpurified_Exo.csv This document is the correlation of expression of eight proteins on unpurified Exo derived from PANC-1-, MIAPaCa-2-, and SW1990 cell lines by NC-Chip. The unpurified Exo represents a cell-depleted culture medium. (10) Fig.4i-2-Correlation_of_expression_of_eight_proteins_on_UC-purified_Exo.csv This document is the correlation of expression of eight proteins on ultracentrifugation (UC)-purified Exo derived from PANC-1-, MIAPaCa-2-, and SW1990 cell lines by NC-Chip. (11) Fig.4j-NC-Chip_assessment_of_tExos_in_PR_control_and_two_PC_patient_plasma.csv This document is the NC-Chip assessment of eight protein expression levels on Tumor-derived exosomes (tExos) directly from one pancreatitis (PR) control and two pancreatic cancer (PC) patient plasma samples. (12) Fig.5-Integrative_phenotyping_of_pancreatic_cancer_exosomes_in_a_training \_cohort.csv This document is the integrative phenotyping of pancreatic cancer exosomes in a training cohort. A training set of 28 age-matched healthy donors (HD), 26 PR, 40 non-metastatic pancreatic cancer (nPC), and 25 metastatic pancreatic cancer (mPC). After removing the background, the signal intensities for each sample were averaged over three measurements and then normalized using min-max. The NA represents represents the word "information not available." (13) Fig.6-Validation_of_the_NC-Chip_for_PC_diagnosis.csv This document is the validation of the NC-Chip for PC diagnosis. A validation set of 11 HD, 26 PR, 33 nPC, and 20 mPC participants. After removing the background, the signal intensities for each sample were averaged over three measurements and then normalized using min-max.