P–769 FTIR spectroscopy analysis reveals differences between human embryo culture media composition by type of formulation and by manufacturer
Authors: Ribeiro I , Pires F , Calado C R C , Gallard. Molina M 
Published: August 2021

Abstract

Study question
Can we detect the variation in different commercial human embryo culture media composition by Fourier-transform infrared (FTIR) spectroscopic analysis?

Summary answer
The spectra reveals distint features that allow distinguishing between continuous and sequential media, as well as between manufacturers of the same media type. What is known already: Manufacturers do not fully disclose commercially available culture media formulations. For this reason, it is important to gain insight into de differences between the available formulations, in order to understand how they might be linked with efficacy and safety ART parameters. Fourier Transform Infra-Red (FTIR) can be used for this purpose.

Study design, size, duration
Culture media samples (1 mL) were collected from local IVF laboratories, and stored frozen at –20 °C. Five repeats of 25 µL aliquots from the samples were analysed using FTIR spectroscopy to acquire the whole molecular fingerprint of each culture media.

Participants/materials, setting, methods
Three continuous (SAGE 1-STEP, Origio, G-TL, Vitrolife, and GERI, Genea) and four sequential (G1 PLUS, Vitrolife; Sequential Cleav with phenolred, Origio; G2 PLUS, Vitrolife and Sequential Blast with phenolred, Origio) media were analysed. Different pre-processing methods (atmospheric and baseline correction, normalizations, and derivatives) were carried out to minimize physical artefacts while highlighting chemical features. To compare the spectra of different media, multivariate analysis, as principal component analysis (PCA) and hierarchical cluster analysis (HCA) were employed.

Main results and the role of chance
The whole molecular fingerprint of all media analysed showed a similar pattern, revealing that, overall, the composition is very similar. However, PCA and HCA analysis revealed that significant differences exist, both between media type (continuous vs. sequential), and between different manufacturers within the same media type. Average linkage clustering using Spearman’s rank correlation confirms the similarities between the continuous and the sequential formulations. An analysis focusing the fingerprint region of the spectra (900 – 1700 cm^–1), also revealed variability between manufacturer, between media type (continuous vs. sequential) and of stage-specific media (cleavage vs. blastocyst). For instance, GERI media visually appeared to have distinct peaks compared to all other media, which was confirmed later through multivariate statistical analysis.

Limitations, reasons for caution
FTIR spectroscopy does not allow for a direct identification of the analytes present in the culture media, we can only infer the functional groups, but that are common on diverse biomolecules.
Wider implications of the findings: FTIR analysis reveals differences between different media, such as cleavage and blastocyst specific, sequential and continuous, or manufacturer’s formulations. The increased resolution of the FTIR profile proves to be a powerful tool for analysing human embryo media, and could be used to establish correlations with media clinical performance and safety.

Trial registration number
Not applicable