Dear friend Souad Hamraoui

The Alamar Blue cell viability test is a commonly used assay for assessing cell viability by measuring metabolic activity. It is based on resazurin, a compound that upon entering living cells is reduced to resorufin. The resulting fluorescence is read on a plate reader or fluorescence spectrophotometer.

According to Thermo Fisher Scientific, the Alamar Blue cell viability reagent can be used to assess cell viability by simply adding the 10X, ready-to-use solution to mammalian or bacterial cells in culture media. There is no requirement to aspirate media from cells or place cells in minimal media. Consequently, alamarBlue® reagent can easily be used in a single tube or microtiter plate format in a “no-wash” fashion.

Regarding your question about whether the compound reduced by the cells gets out of the gel into the culture medium, I couldn't find any information on that topic specifically. However, according to Allevi3D, AlamarBlue is a fast assay that can measure cell viability and it is based on resazurin which enters living cells and is reduced to resorufin. Therefore, it seems that if the cells are alive and metabolically active enough to reduce resazurin to resorufin, then it should be able to do so regardless of whether it's encapsulated in an alginate-based hydrogel or not.

I hope this helps! Let me know if you have any other questions.

Source:

(1) alamarBlue® Cell Viability Reagent - Thermo Fisher Scientific.

(2) AlamarBlue – Cell Viability Assay for 3D Cell Culture.

(3) Multiple applications of Alamar Blue as an indicator of metabolic .... https://pubmed.ncbi.nlm.nih.gov/23112716/.

(4) alamarBlue Assays for Cell Viability - Thermo Fisher Scientific. https://www.thermofisher.com/us/en/home/life-science/cell-analysis/fluorescence-microplate-assays/microplate-assays-cell-viability/alamarblue-assay-cell-viability.html.

(5) alamarBlue Assays for Cell Viability Protocol, for Microplates. https://www.thermofisher.com/us/en/home/references/protocols/cell-and-tissue-analysis/protocols/alamarblue-cell-viability-reagent-for-microplates-protocol.html.

hi Souad Hamraoui

Dr. Kaushik Shandilya has explained the context of Alamar blue usage. I will just say that due to their non-toxic nature and non-interference with hydrogel material, Alamar blue remains the best dye to study hydrogel-encapsulated cell viability.

We have used Alamar blue in study cell viability of MSCs encapsulated in sodium alginate. After staining with dye on day 1, we washed the beads and again incubated for further. like this we maintained cells for 5 days and measured the reduced resazurin directly proportional to cell viability.

Article Encapsulated human mesenchymal stem cells (eMSCs) as a novel...

Regards

Saurabh

This is very helpful, Thank you very much for your answers Saurabh Mandal & Dr. Kaushik Shandilya