README file
Data Set Title: Dataset of the article "Combining circulating tumor cell and circulating cell free DNA analyses enhances liquid biopsy sensitivity in detecting high grade serous tubo-ovarian carcinoma”
Data Set Author: 
Ivana Kurelac (Department of Medical and Surgical Sciences, University of Bologna), ORCID 0000-0002-8364-9985; 
Data Set Contributors: 
Beatrice Cavina (Department of Medical and Surgical Sciences, University of Bologna)
Dario De Biase (Department of Pharmacy and Biotechnology, University of Bologna)
Data Set Contact Person: 
Ivana Kurelac (Department of Medical and Surgical Sciences, University of Bologna), ORCID 0000-0002-8364-9985; email: ivana.kurelac@unibo.it  
Data Set License: This data set is distributed under a Creative Commons Attribution 4.0 International (CC BY 4.0) license, https://creativecommons.org/licenses/by/4.0/.
Publication Year: 2025
Project Info: The research leading to the results found in this dataset received funding from MORFEO (MORphological biomarkers For Early diagnosis in Oncology), funded by Italian Ministry of University and Research, “Progetti di Rilevante Interesse Nazionale” funding programme - anno 2017 (PRIN 2017), ID 2017N7R2CJ. 
Data set Contents
The data set consists of:

•	4 compressed folders 
“Cavina_2025_raw_data_qRT-PCR_analyses.zip” (that contains 5 .xlsx files)  
“Cavina_2025_raw_data_dPCR.zip” (that contains 3 .xlsx files)  
“Cavina_2025_raw_data_Spiked_SKOV3_GFP_cells.zip” (that contains 14 .tiff images)
“Cavina_2025_raw_data_Sanger_sequencing.zip” (that contains 26 .ab1 files)

•	1 README file in .rtf format 
“Cavina_2025_raw_data_README.rtf”

Data set Documentation
Abstract
Liquid biopsy is a promising strategy for detecting and monitoring neoplastic diseases, with circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA) being the most common objects of investigation. Most laboratories are focusing on these biomarkers separately and simultaneous detection has not yet been attempted in high grade serous tubo-ovarian carcinoma (HGSOC). This dataset comprises raw data obtained in the study in which we assess whether tandem CTC/ctDNA approach increases HGSOC detection efficiency. We developed specific liquid biopsy assays, all based on nucleic acid analyses. For CTC detection, we tested gene expression assays and TP53 next-generation sequencing (NGS), whereas the latter was also applied for ctDNA detection. The assays were then verified on two HGSOC cohorts and respective controls, demonstrating the complementary value of simultaneous CTC and cfDNA analysis in HGSOC. Specifically, this dataset contains (i) raw data obtained from all quantitative Real-Time PCR and digital PCR analyses, (ii) Sanger sequencing of patient germ-line tissue to confirm tumor-specific mutations, and (iii) microscopic images of the Parsortix-enriched ovarian cancer cells spiked in the healthy donor blood sample.

Content of the files: 
-	The compressed folder “Cavina_2025_raw_data_qRT-PCR_analyses.zip” contains 5 .xlsx files:
1.	File “validation_SYBR.xlsx” containing qRT-PCR results obtained by Applied Biosystems sds7500fast from screening 21 genes in HGSOC patient derived biopsies (n=8, T1-T8), and healthy donor derived PBMC samples (n=3, CNTR1-CNTR3). The file contains 9 sheets, entitled “Results1” to “Results9”. Each sheet reports exported results file retrieved from the instrument, which includes general experiment information (Block Type, Chemistry, Experiment Run End Time, Instrument Type, Passive Reference), and the following data listed in columns: Well, Sample Name, Target Name, Reporter, Ct, Ct Threshold, Tm1, Tm2, Tm3. 
Results1: raw data from samples T1-T2-T3-T4-CNTR1-CNTR2-CNTR3 for the targets EPCAM, ERCC1, KRT18, KRT19, PPIC, ERBB2, with respective technical duplicate and blank (BLK). 
Results2: raw data from samples T1-T2-T3-T4-CNTR1-CNTR2-CNTR3 for the targets AGR2, LAMB1, MAL2, PRAME, SERPINE2, TUSC3, with respective technical duplicate and blank (BLK).  
Results3: raw data from samples T1-T2-T3-T4-CNTR1-CNTR2-CNTR3 for the targets CDH2, GPX8, MUC1, with respective technical duplicate and blank (BLK). 
Results4: raw data from samples T1-T2-T3-T4-T5-T6-T7-T8-CNTR1-CNTR2-CNTR3 for the targets WT1, TIFF1, with respective technical duplicate and blank (BLK). 
Results5: raw data from samples T1-T2-T3-T4-T5-T6-T7-T8-CNTR1-CNTR2-CNTR3 for the targets CDH3, PGR, with respective technical duplicate and blank (BLK). 
Results6: raw data from samples T1-T2-T3-T4-T5-T6-T7-T8-CNTR1-CNTR2-CNTR3 for the targets ERBB3, TIMP1, with respective technical duplicate and blank (BLK). 
Results7: raw data from samples T5-T6-T7-T8 for the targets ERBB2, GPX8, LAMB1, MAL2, with respective technical duplicate and blank (BLK). 
Results8: raw data from samples T5-T6-T7-T8 for the targets TUSC3, PRAME, AGR2, SERPINE2, CDH2, MUC1, EPCAM, ERCC1, KRT18, with respective technical duplicate and blank (BLK). 
Results9: raw data from samples T5-T6-T7-T8 for the targets KRT19, PPIC, with respective technical duplicate and blank (BLK).
2.	File “validation_controls_TaqMan.xlsx” containing qRT-PCR results obtained by Applied Biosystems sds7500fast from screening 12 genes in healthy donor derived PBMC samples (n=10, CNTR4-CNTR13). The file contains 12 sheets, entitled with the respective gene analyzed: GPX8, WT1, PRAME, CDH3, MUC1, ERBB3, AGR2, LAMB1, PGR, MAL2, ERBB2, EPCAM. Each sheet reports exported results file retrieved from the instrument, which includes general experiment information (Block Type, Chemistry, Experiment Run End Time, Instrument Type, Passive Reference), and the following data listed in columns: Sample Name, Target Name, Task, Reporter, Quencher, Ct, Ct Threshold. The column cDNA was added to indicate which type of sample has been used, native cDNA or pre-amplified (PA). The data are from samples CNTR4-13, with respective technical duplicate, blank (B), and positive control for each sheet. 
3.	File “OC_cells_TaqMan.xlsx” containing qRT-PCR results obtained by Applied Biosystems sds7500fast from screening AGR2, WT1 and GPX8 genes in OC cell lines, namely SKOV3, OV90, and OC314. The Results sheet reports exported results file retrieved from the instrument, which includes general experiment information (Block Type, Chemistry, Experiment Run End Time, Instrument Type, Passive Reference), and the following data listed in columns: Sample Name, Target Name, Task, Reporter, Quencher, Ct, Ct Threshold. The data are presented with respective technical duplicate and blank.
4.	File “Spiking_TaqMan.xlsx” containing qRT-PCR results obtained by Applied Biosystems sds7500fast from screening AGR2, WT1 and GPX8 genes in spiked samples with known number of SKOV3 cells and enriched or not with Parsortix or CD45 negative magnetic assisted cell sorting. The file contains 2 sheets, entitled with the respective gene analyzed, WT1 and GPX8_AGR2. Each sheet reports exported results file retrieved from the instrument, which includes general experiment information (Block Type, Chemistry, Experiment Run End Time, Instrument Type, Passive Reference), and the following data listed in columns: Sample Name, Target Name, Task, Reporter, Quencher, Ct, Ct Threshold. The column cDNA was added to indicate which type of sample has been used, native cDNA or pre-amplified (PA). The data are from samples mimicking liquid biopsies by spiking known ovarian cancer cell numbers (0, 5, 50 or 500k) in healthy donor peripheral blood, namely: NotEnriched_0SKOV, NotEnriched_5SKOV, NotEnriched_50SKOV, NotEnriched_500kSKOV, Parsortix_0SKOV, Parsortix_5SKOV, Parsortix_50SKOV, Parsortix_500kSKOV, CD45sorting_0SKOV, CD45sorting_5SKOV, CD45sorting_50SKOV, CD45sorting_500kSKOV, with respective technical duplicate and blank (B) for each sheet.
5.	File “patients_TaqMan.xlsx” containing qRT-PCR results obtained by Applied Biosystems sds7500fast from screening AGR2, WT1 and GPX8 genes in HGSOC patient derived liquid biopsy (HGSOC-1 to HGSOC-15). The Results sheet reports exported results file retrieved from the instrument, which includes general experiment information (Block Type, Chemistry, Experiment Run End Time, Instrument Type, Passive Reference), and the following data listed in columns: Well, Sample Name, Target Name, Task, Reporter, Quencher, Ct, Ct Threshold. The data are presented with respective technical duplicate, blank of pre-amplification (b PA), blank of retro transcription (b RT), RNA not retro transcribed (RT-), qRT-PCR blank (b qRT-PCR), and positive control (SKOV3).

-	The compressed folder “Cavina_2025_raw_data_dPCR.zip” contains 3 .xlsx files:
1.	File “SKOV3.xlsx” containing dPCR results obtained by Absolute Q 22120661 from screening AGR2, WT1 and GPX8 genes in SKOV3. The file contains 2 sheets, entitled with the respective gene analyzed, WT1 and GPX8_AGR2. Each sheet reports exported results retrieved from the instrument, which the following data listed in columns: Run, Instrument, Plate, Sample, Index, Reject, ROX, FAM_gene name, FAM_gene name_pos, VIC_gene name, VIC_gene name_pos (sheet GPX8_AGR2). 
2.	File “Parsortix.xlsx” containing dPCR results obtained by Absolute Q 22120661 from screening AGR2, WT1 and GPX8 genes in spiked samples with known number of SKOV3 cells and enriched with Parsortix. The file contains 8 sheets, entitled with the sample name and respective gene analyzed: “500k_Parsortix_WT1”, “500k_Parsortix_GPX8_AGR2”, “50_Parsortix_WT1”, “50_Parsortix_GPX8_AGR2”, “5_Parsortix_WT1”, “5_Parsortix_GPX8_AGR2”, “0_Parsortix_WT1”, and “0_Parsortix_GPX8_AGR2”, where 0, 50 and 500k indicate the number of cancer cells spiked into healthy donor peripheral blood. Each sheet reports exported results retrieved from the instrument, which the following data listed in columns: Run, Instrument, Plate, Sample, Index, Reject, ROX, FAM_gene name, FAM_gene name_pos, VIC_gene name, VIC_gene name_pos. 
3.	File “CD45_neg_selection.xlsx” containing dPCR results obtained by Absolute Q 22120661 from screening AGR2, WT1 and GPX8 genes in spiked samples with known number of SKOV3 cells and enriched with CD45 negative sorting. The file contains 8 sheets, entitled with the sample name and respective gene analyzed: “500k_CD45_WT1”, “500k_CD45_GPX8_AGR2”, “50_CD45_WT1”, “50_CD45_GPX8_AGR2”, “5_CD45_WT1”, “5_CD45_GPX8_AGR2”, “0_CD45_WT1”, and “0_CD45_GPX8_AGR2”, where 0, 50 and 500k indicate the number of cancer cells spiked into healthy donor peripheral blood. Each sheet reports exported results retrieved from the instrument, which the following data listed in columns: Run, Instrument, Plate, Sample, Index, Reject, ROX, FAM_gene name, FAM_gene name_pos, VIC_gene name, VIC_gene name_pos. 

-	The compressed folder “Cavina_2025_raw_data_Spiked_SKOV3_GFP_cells.zip” contains 14 .tiff images obtained by EVOS M5000 Imaging System of 5 SKOV3 spiked sample enriched with Parsortix (SKOV3GFP), and of SKOV3 cells in flask (files “SKOV3_GFP.tiff” and “SKOV3_TRANS.tiff”). For 5 SKOV3 spiked sample 4 events were captured in phase contrast (“SKOV3GFP_ID event_TRANS.tiff” files), GFP channel (“SKOV3GFP_ID event_GFP.tiff” files), and the merge (“SKOV3GFP_ID event.tiff” files).
-	The compressed folder “Cavina_2025_raw_data_Sanger_sequencing.zip” with 26 .ab1 files obtained by ABI PRISM 3730 capillary sequencer Applied Biosystems from sequencing TP53 specific exons in HGSOC patient derived PBMC (n=15). For each sequence the file name reports the HGSOC patient, forward (fw) or revers (rev) strand sequenced, 96-well position, RUN name. For HGSOC-1, -2, -3, -7, -8, -22, -23, -25, -26, -27, -28 both strands are reported. For HGSOC-16 and -30 only forward sequence, whereas for HGSOC-6 and -29 only the reverse strand has been analyzed. 

Notes 
•	The data included in this dataset are presented in the following Biorxiv preprint: 
Cavina, B., Corrà, S., Coadă, C. A., De Luise, M., Lemma, S., Coluccelli, S., De Leo, A., Di Costanzo, S., Mezzapesa, F., Girolimetti, G., De Iaco, P., Porcelli, A. M., Perrone, A. M., de Biase, D., Gasparre, G., & Kurelac, I. (2025). Combining circulating tumor cell and circulating cell free DNA analyses enhances liquid biopsy sensitivity in detecting high grade serous tubo-ovarian carcinoma. bioRxiv, 2025.10.13.681971. https://doi.org/10.1101/2025.10.13.681971
•	The related next generation sequencing data are available at https://www.ncbi.nlm.nih.gov/bioproject/PRJNA1338131.