Role of Non-invasive Preimplantation Genetic Testing-Aneuploidy Using Spent Culture Media and its Concordance with Trophectoderm Biopsy: A Proof of Concept and Validation Study
Background: While trophectoderm (TE) biopsy with next-generation sequencing (NGS) remains the gold standard for preimplantation genetic testing for aneuploidy (PGT-A), the discovery of cell-free DNA (cfDNA) in spent culture media (SCM) has sparked interest in non-invasive PGT-A (NiPGT-A) as a potent...
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| Format: | Article |
| Language: | English |
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Wolters Kluwer Medknow Publications
2025-04-01
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| Series: | Journal of Human Reproductive Sciences |
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| Online Access: | https://journals.lww.com/10.4103/jhrs.jhrs_192_24 |
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| author | Neeta Singh Ankita Sethi Lata Rani Monika Saini Ritu Gupta Neena Malhotra Reeta Mahey |
| author_facet | Neeta Singh Ankita Sethi Lata Rani Monika Saini Ritu Gupta Neena Malhotra Reeta Mahey |
| author_sort | Neeta Singh |
| collection | DOAJ |
| description | Background:
While trophectoderm (TE) biopsy with next-generation sequencing (NGS) remains the gold standard for preimplantation genetic testing for aneuploidy (PGT-A), the discovery of cell-free DNA (cfDNA) in spent culture media (SCM) has sparked interest in non-invasive PGT-A (NiPGT-A) as a potential alternative.
Aim:
The study was conducted to assess the feasibility of cell cfDNA from SCM as a tool for NiPGT-A in patients undergoing IVF for advanced age, repeated implantation failure or severe male factor infertility.
Settings and Design:
This is a prospective study where a total of 44 embryos having TE biopsy for aneuploidy testing and their respective SCM collected at day 5/6 were analysed.
Materials and Methods:
All aneuploid blastocysts (WB) were subjected to DNA extraction and amplification using Sureplex DNA amplification system followed by library preparation using VeriSeq™ PGS Library Prep kit and sequencing on MiSeq (Illumina, California, USA).
Statistical Analysis Used:
Copy Number Variation visualisation and analysis were carried out using BlueFuse Multi Software (Illumina). The statistical data were analysed by STATA version 14.
Results:
Informative results were obtained in 36/44 (81.2%) SCM samples. The reads were analysable in 26 paired (SCM and TE biopsy) samples. Concordant NGS results for both TE biopsy and SCM sample were obtained in 17/26 (65.38%) embryos. The per chromosome concordance rate was 85.13% (487/572) and the sex chromosome concordance rate was 73% (19/26). The sensitivity and specificity of NiPGT-A were 66.6% and 60%, respectively. On comparing the ploidy concordance rate, poor morphology embryos had better, but not statistically significant concordance rate (83.33%) as compared to good morphology embryos (50%, P = 0.16). Although not significant, day 6 embryos had better per chromosome as well as sex chromosome concordance rate as compared to day 5 embryos.
Conclusion:
Aneuploidy testing using cf DNA in SCM is a promising technique but needs more research on larger cohort size to improve the sensitivity, specificity and concordance rate. |
| format | Article |
| id | doaj-art-00bfcbc186904892bb63f1d4728b75e5 |
| institution | DOAJ |
| issn | 0974-1208 1998-4766 |
| language | English |
| publishDate | 2025-04-01 |
| publisher | Wolters Kluwer Medknow Publications |
| record_format | Article |
| series | Journal of Human Reproductive Sciences |
| spelling | doaj-art-00bfcbc186904892bb63f1d4728b75e52025-08-20T02:43:27ZengWolters Kluwer Medknow PublicationsJournal of Human Reproductive Sciences0974-12081998-47662025-04-01182707910.4103/jhrs.jhrs_192_24Role of Non-invasive Preimplantation Genetic Testing-Aneuploidy Using Spent Culture Media and its Concordance with Trophectoderm Biopsy: A Proof of Concept and Validation StudyNeeta SinghAnkita SethiLata RaniMonika SainiRitu GuptaNeena MalhotraReeta MaheyBackground: While trophectoderm (TE) biopsy with next-generation sequencing (NGS) remains the gold standard for preimplantation genetic testing for aneuploidy (PGT-A), the discovery of cell-free DNA (cfDNA) in spent culture media (SCM) has sparked interest in non-invasive PGT-A (NiPGT-A) as a potential alternative. Aim: The study was conducted to assess the feasibility of cell cfDNA from SCM as a tool for NiPGT-A in patients undergoing IVF for advanced age, repeated implantation failure or severe male factor infertility. Settings and Design: This is a prospective study where a total of 44 embryos having TE biopsy for aneuploidy testing and their respective SCM collected at day 5/6 were analysed. Materials and Methods: All aneuploid blastocysts (WB) were subjected to DNA extraction and amplification using Sureplex DNA amplification system followed by library preparation using VeriSeq™ PGS Library Prep kit and sequencing on MiSeq (Illumina, California, USA). Statistical Analysis Used: Copy Number Variation visualisation and analysis were carried out using BlueFuse Multi Software (Illumina). The statistical data were analysed by STATA version 14. Results: Informative results were obtained in 36/44 (81.2%) SCM samples. The reads were analysable in 26 paired (SCM and TE biopsy) samples. Concordant NGS results for both TE biopsy and SCM sample were obtained in 17/26 (65.38%) embryos. The per chromosome concordance rate was 85.13% (487/572) and the sex chromosome concordance rate was 73% (19/26). The sensitivity and specificity of NiPGT-A were 66.6% and 60%, respectively. On comparing the ploidy concordance rate, poor morphology embryos had better, but not statistically significant concordance rate (83.33%) as compared to good morphology embryos (50%, P = 0.16). Although not significant, day 6 embryos had better per chromosome as well as sex chromosome concordance rate as compared to day 5 embryos. Conclusion: Aneuploidy testing using cf DNA in SCM is a promising technique but needs more research on larger cohort size to improve the sensitivity, specificity and concordance rate.https://journals.lww.com/10.4103/jhrs.jhrs_192_24cell-free dnanext-generation sequencingnon-invasive preimplantation genetic testingspent culture mediawhole genome amplification |
| spellingShingle | Neeta Singh Ankita Sethi Lata Rani Monika Saini Ritu Gupta Neena Malhotra Reeta Mahey Role of Non-invasive Preimplantation Genetic Testing-Aneuploidy Using Spent Culture Media and its Concordance with Trophectoderm Biopsy: A Proof of Concept and Validation Study Journal of Human Reproductive Sciences cell-free dna next-generation sequencing non-invasive preimplantation genetic testing spent culture media whole genome amplification |
| title | Role of Non-invasive Preimplantation Genetic Testing-Aneuploidy Using Spent Culture Media and its Concordance with Trophectoderm Biopsy: A Proof of Concept and Validation Study |
| title_full | Role of Non-invasive Preimplantation Genetic Testing-Aneuploidy Using Spent Culture Media and its Concordance with Trophectoderm Biopsy: A Proof of Concept and Validation Study |
| title_fullStr | Role of Non-invasive Preimplantation Genetic Testing-Aneuploidy Using Spent Culture Media and its Concordance with Trophectoderm Biopsy: A Proof of Concept and Validation Study |
| title_full_unstemmed | Role of Non-invasive Preimplantation Genetic Testing-Aneuploidy Using Spent Culture Media and its Concordance with Trophectoderm Biopsy: A Proof of Concept and Validation Study |
| title_short | Role of Non-invasive Preimplantation Genetic Testing-Aneuploidy Using Spent Culture Media and its Concordance with Trophectoderm Biopsy: A Proof of Concept and Validation Study |
| title_sort | role of non invasive preimplantation genetic testing aneuploidy using spent culture media and its concordance with trophectoderm biopsy a proof of concept and validation study |
| topic | cell-free dna next-generation sequencing non-invasive preimplantation genetic testing spent culture media whole genome amplification |
| url | https://journals.lww.com/10.4103/jhrs.jhrs_192_24 |
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