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Accessing the Cytotoxicity and Cell Response to Biomaterials

Anabela Baptista Paula*1,2,3,4, Mafalda Laranjo*2,3,4,5, Ana Sofia Coelho1,2,3,4, Ana Margarida Abrantes2,3,4,5, Ana Cristina Gonçalves2,3,4,6, Ana Bela Sarmento-Ribeiro2,3,4,6, Manuel Marques Ferreira2,3,4,7, Maria Filomena Botelho2,3,4,5, Carlos Miguel Marto1,2,3,4,8, Eunice Carrilho1,2,3,4
* These authors contributed equally

Abstract

Biomaterials contact directly or indirectly with the human tissues, making it important to evaluate its cytotoxicity. This evaluation can be performed by several methods, but a high discrepancy exists between the approaches used, compromising the reproducibility and the comparison among the obtained results. In this paper, we propose a protocol to evaluate biomaterial cytotoxicity using soluble extracts, which we use for dental biomaterials. The extract preparation is detailed, from pellet production to its extraction in a culture medium. The biomaterial cytotoxicity evaluation is based the following: metabolic activity using the MTT assay, cell viability using the Sulphorhodamine B (SBR) assay, cell death profile by flow cytometry, and cell morphology using May-Grünwald Giemsa. Additional to cytotoxicity evaluation, a protocol to evaluate cell function is described based on the expression of specific markers assessed by immunocytochemistry and PCR. This protocol provides a comprehensive guide for biomaterials cytotoxicity and cellular effects evaluation, using the extracts methodology, in a reproducible and robust manner.

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