Barcode tagging of human oocytes and embryos to prevent mix-ups in assisted reproduction technologies.
OBJECTIVE
Is the attachment of biofunctionalized polysilicon barcodes to the outer surface of the zona pellucida an effective approach for the direct tagging and identification of human oocytes and embryos during assisted reproduction technologies (ARTs)?


CONCLUSIONS
The direct tagging system based on lectin-biofunctionalized polysilicon barcodes of micrometric dimensions is simple, safe and highly efficient, allowing the identification of human oocytes and embryos during the various procedures typically conducted during an assisted reproduction cycle.


BACKGROUND
Measures to prevent mismatching errors (mix-ups) of the reproductive samples are currently in place in fertility clinics, but none of them are totally effective and several mix-up cases have been reported worldwide. Using a mouse model, our group has previously developed an effective direct embryo tagging system which does not interfere with the in vitro and in vivo development of the tagged embryos. This system has now been tested in human oocytes and embryos.


METHODS
Fresh immature and mature fertilization-failed oocytes (n = 21) and cryopreserved day 1 embryos produced by in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) (n = 205) were donated by patients (n = 76) undergoing ARTs. In vitro development rates, embryo quality and post-vitrification survival were compared between tagged (n = 106) and non-tagged (control) embryos (n = 99). Barcode retention and identification rates were also calculated, both for embryos and for oocytes subjected to a simulated ICSI and parthenogenetic activation. Experiments were conducted from January 2012 to January 2013.


METHODS
Barcodes were fabricated in polysilicon and biofunctionalizated with wheat germ agglutinin lectin. Embryos were tagged with 10 barcodes and cultured in vitro until the blastocyst stage, when they were either differentially stained with propidium iodide and Hoechst or vitrified using the Cryotop method. Embryo quality was also analyzed by embryo grading and time-lapse monitoring. Injected oocytes were parthenogenetically activated using ionomycin and 6-dimethylaminopurine.


RESULTS
Blastocyst development rates of tagged (27/58) and non-tagged embryos (24/51) were equivalent, and no significant differences in the timing of key morphokinetic parameters and the number of inner cell mass cells were detected between the two groups (tagged: 24.7 ± 2.5; non-tagged: 22.3 ± 1.9), indicating that preimplantation embryo potential and quality are not affected by the barcodes. Similarly, re-expansion rates of vitrified-warmed tagged (19/21) and non-tagged (16/19) blastocysts were similar. Global identification rates of 96.9 and 89.5% were obtained in fresh (mean barcode retention: 9.22 ± 0.13) and vitrified-warmed (mean barcode retention: 7.79 ± 0.35) tagged embryos, respectively, when simulating an automatic barcode reading process, though these rates were increased to 100% just by rotating the embryos during barcode reading. Only one of the oocytes lost one barcode during intracytoplasmic injection (100% identification rate) and all oocytes retained all the barcodes after parthenogenetic activation.


CONCLUSIONS
Although the direct embryo tagging system developed is effective, it only allows the identification and traceability of oocytes destined for ICSI and embryos. Thus, the traceability of all reproductive samples (oocytes destined for IVF and sperm) is not yet ensured.


CONCLUSIONS
The direct embryo tagging system developed here provides fertility clinics with a novel tool to reduce the risk of mix-ups in human ARTs. The system can also be useful in research studies that require the individual identification of oocytes or embryos and their individual tracking.


BACKGROUND
This study was supported by the Sociedad Española de Fertilidad, the Spanish Ministry of Education and Science (TEC2011-29140-C03) and the Generalitat de Catalunya (2009SGR-00282 and 2009SGR-00158). The authors do not have any competing interests.
Departament de Biologia Cel·lular, Fisiologia i Immunologia, Facultat de Biociències, Universitat Autònoma de Barcelona, Bellaterra 08193, Spain.
Sergi
S
Novo
Carme
C
Nogués
Oriol
O
Penon
Leonardo
L
Barrios
Josep
J
Santaló
Rodrigo
R
Gómez-Martínez
Jaume
J
Esteve
Abdelhamid
A
Errachid
José Antonio
JA
Plaza
Lluïsa
L
Pérez-García
Elena
E
Ibáñez
D001755:Blastocyst; D004624:Embryo Transfer; D004622:Embryo, Mammalian / Q000378:metabolism*; D047108:Embryonic Development; D006801:Humans; D009865:Oocytes / Q000166:cytology*; D027724:Reproductive Techniques, Assisted / Q000592:standards*; D012825:Silicon / Q000378:metabolism; D058989:Vitrification; D014909:Wheat Germ Agglutinins / Q000378:metabolism; D015044:Zona Pellucida / Q000378:metabolism
D014909:Wheat Germ Agglutinins; D012825:Silicon
embryo identification; polysilicon barcodes; traceability