Not to be Overlooked: Cytotoxicity Testing of Biomaterials and Medical Devices

May 5, 2023

Biomaterials are defined as a material intended to interface with biological systems to evaluate, treat, augment, or replace any tissue, organ, or function in the body. Medical devices incorporate these biomaterials into their design for use in the body. One of the most important tests that must be performed during the development of a biomaterial is the laboratory-based assessment of its cytotoxicity status. Cytotoxicity refers to whether the biomaterial will cause mammalian cell death due to leaching of toxic substances or from direct contact. It is critical to perform this test because if not, the biomaterial may cause mammalian cell death and can lead to massive inflammatory responses occurring and rejection of the device.

The cytotoxicity test involves growing mammalian cells and exposing these cells to the biomaterial. This exposure can take numerous approaches:

  • Direct contact, the biomaterial is laid directly on the mammalian cells.
  • Indirect contact, the biomaterial is placed on a filter in contact with mammalian cells and any cytotoxic substances pass through the filter and potentially impact the cells.
  • Extract testing, extracts are eluted from the biomaterial and these extracts are tested against a mammalian cell culture.

Once the requisite incubation is complete, the physical biomaterial is removed, and the mammalian cells are stained using a viability stain to calculate how many cells remain alive after exposure to the biomaterial. Figure 1 demonstrates the appearance of the cells after staining with a viability stain.

Figure 1: Fibroblasts attached to a polycaprolactone scaffold and stained with a viability stain. Red cells are designated as dead, whereas the green cells are considered alive. As can be observed from this image, the majority of the cells are alive.

The analysis of these cells within the Biohubx common laboratory are performed using a Biotek Cytation C10 which allows for the selection of certain search criteria in your cells, for example, the red and green colours associated with the viability stain. Once these search criteria are specified, the Cytation C10 will automatically scan your cells for these criteria and produce data for analysis.

Regardless of whichever methodology is used, the cut off point for these types of tests is that the total amount of cells alive after exposure to the biomaterial or extract should be above 70%. Any value below this is considered potentially cytotoxic. If a biomaterial is considered to be potentially cytotoxic, additional testing is required.

These tests follow the guidelines laid out in ISO 10993-5. Biohubx is capable of performing these tests to these standards and the production of reports. Please contact Dr. Steven Jones ( for further information.