Naslov
Optimized protocol for whole-mount immunohistochemistry in zebrafish embryos (Danio rerio)

Autori
Karaica, Dean ; Mihaljević, Ivan ; Vujica, Lana ; Dragojević, Jelena ; Lončarić, Nora ; Bošnjak, Arvena ; Otten, Cecile ; Lončar, Jovica ; Smital, Tvrtko

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
Book of Abstracts of the Congress of the Croatian Society of Biochemistry and Molecular Biology / Dulić, Morana ; Sinčić, Nino ; Vrhovac Madunić, Ivana - Zagreb : Hrvatsko Društvo za Biotehnologiju, 2022, 90-90

Skup
Congress of the Croatian Society of Biochemistry and Molecular Biology: From Science to Knowledge (HDBMB22)

Mjesto i datum
Brela, Hrvatska, 28.09.2022. - 01.10.2022

Vrsta sudjelovanja
Poster

Vrsta recenzije
Recenziran

Ključne riječi
whole-mount ; immunohistochemistry ; zebrafish ; optimization

Sažetak
In the last couple of decades, the use of zebrafish model in biomedical and toxicological research is exponentially increasing. Many studies investigate the physiological/ecotoxicological function of different proteins localized across different zebrafish organs/tissues. Whole-mount preparations coupled with fluorescence microscopy are widely used for this task. To date there is no single approach for conducting whole-mount immunohistochemistry in zebrafish embryos. In this study we used zebrafish embryos (96 hpf) to determine key parameters in preparation of zebrafish embryos for the use in whole-mount immunohistochemistry using two antibodies detecting Na+/K+ATPase and ZN-8 membrane proteins. Two fixatives (4% paraformaldehyde/4% PFA and Dents fixative) and three different incubation temperatures (37 °C/30 min, 70 °C/15 min, 95 °C/5 min) were tested to optimize the whole-mount immunohistochemistry approach. Results showed that the use of Dents fixative compared to 4% PFA fixative enables better binding of the tested primary antibodies and reveals the known localization of Na+/K+ATPase and ZN-8 proteins in zebrafish embryos. However, the use of different incubation temperatures was not crucial for the binding of the tested antibodies. Overall, this work presents a unified approach for whole-mount immunohistochemistry that can be used to test optimal conditions for localization of proteins in zebrafish embryos.

Izvorni jezik
Engleski

Znanstvena područja
Biologija, Temeljne medicinske znanosti

POVEZANOST RADA