<abstract xmlns="http://www.w3.org/1999/xhtml">

<sec><h3>Introduction</h3>
<p>Development of an effective, suitable, and reliable method for both the isolation and culturing of melanocytes is crucial for studies on pathomechanisms of skin diseases originating from melanocyte disorders. In this study, we have investigated the influence of geneticin (G418), a substance used for melanocyte selection, in the view of the frequency of presence of cells such as keratinocytes and fibroblasts, widely known as contaminators of melanocyte-originating cell cultures.</p>
</sec>
<sec><h3>Materials and Methods</h3>
<p>Study was conducted on primary, freshly isolated melanocytes, keratinocytes, fibroblasts, and melanoma cells as well as on commercially available melanoma cell lines MeWo, G-361, and A375. Cells were cultured in different culture media supplemented with various concentrations of geneticin ranging from 0.05 to 1 mg/mL. Cell viability, proliferation rate and detection of apoptotic/necrotic cells was assessed.</p>
</sec>
<sec><h3>Results</h3>
<p>Choice of culture media supplemented with various concentrations of geneticin (0.05 mg/mL, 0.1 mg/mL, 0.5 mg/mL and 1 mg/mL) strongly affect viability of melanocytes, fibroblasts, and keratinocytes. Selective culture media without FBS facilitate the process of melanocytes and melanoma pure cell culture, yet without geneticin supplementation are insufficient for complete eradication of fibroblast contamination from cell culture.</p>
</sec>
<sec><h3>Conclusions</h3>
<p>In this study we provide, for the first time, the dose-response action of keratinocytes and fibroblasts upon geneticin stimulation in different culture media and show that a low concentration (0.05 mg/mL) of geneticin added to the selective culture media may be safely implemented to facilitate the production of melanocyte and melanoma cell cultures that are free from frequent cell contaminants.</p>
</sec>
</abstract>

Introduction
Development of an effective, suitable, and reliable method for both the isolation and culturing of melanocytes is crucial for studies on pathomechanisms of skin diseases originating from melanocyte disorders. In this study, we have investigated the influence of geneticin (G418), a substance used for melanocyte selection, in the view of the frequency of presence of cells such as keratinocytes and fibroblasts, widely known as contaminators of melanocyte-originating cell cultures.

Materials and Methods
Study was conducted on primary, freshly isolated melanocytes, keratinocytes, fibroblasts, and melanoma cells as well as on commercially available melanoma cell lines MeWo, G-361, and A375. Cells were cultured in different culture media supplemented with various concentrations of geneticin ranging from 0.05 to 1 mg/mL. Cell viability, proliferation rate and detection of apoptotic/necrotic cells was assessed.

Results
Choice of culture media supplemented with various concentrations of geneticin (0.05 mg/mL, 0.1 mg/mL, 0.5 mg/mL and 1 mg/mL) strongly affect viability of melanocytes, fibroblasts, and keratinocytes. Selective culture media without FBS facilitate the process of melanocytes and melanoma pure cell culture, yet without geneticin supplementation are insufficient for complete eradication of fibroblast contamination from cell culture.

Conclusions
In this study we provide, for the first time, the dose-response action of keratinocytes and fibroblasts upon geneticin stimulation in different culture media and show that a low concentration (0.05 mg/mL) of geneticin added to the selective culture media may be safely implemented to facilitate the production of melanocyte and melanoma cell cultures that are free from frequent cell contaminants.

Role of geneticin in isolation and culturing of skin melanocytes and melanoma cells

National Institute of Cardiology, Stefan Cardinal Wyszyński State Research Institute

Laboratory of Biomolecular Interactions Studies

Chair of Drug and Cosmetics Biotechnology, Faculty of Chemistry

Department of Plastic and Reconstructive Surgery

Hirszfeld Institute of Immunology and Experimental Therapy

Polish Academy of Sciences, Laboratory of Virology

Postępy Higieny i Medycyny Doświadczalnej

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

Introduction
Development of an effective, suitable, and reliable method for both the isolation and culturing of melanocytes is crucial for studies on...

Role of geneticin in isolation and culturing of skin melanocytes and melanoma cells

Article Category: Original Study

Pubblicato online: 14 set 2023

Pagine: 72 - 81

Ricevuto: 12 mar 2023

Accettato: 06 lug 2023

DOI: https://doi.org/10.2478/ahem-2023-0014

Parole chiave
melanocytes, fibroblasts, keratinocytes, melanoma, geneticin, cell culture contamination

© 2023 Aneta Ścieżyńska et al., published by Sciendo

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

Role of geneticin in isolation and culturing of skin melanocytes and melanoma cells

Article Category: Original Study

Pubblicato online: 14 set 2023

Pagine: 72 - 81

Ricevuto: 12 mar 2023

Accettato: 06 lug 2023

DOI: https://doi.org/10.2478/ahem-2023-0014

Parole chiavemelanocytes, fibroblasts, keratinocytes, melanoma, geneticin, cell culture contamination

© 2023 Aneta Ścieżyńska et al., published by Sciendo

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

Parole chiave
melanocytes, fibroblasts, keratinocytes, melanoma, geneticin, cell culture contamination