Overexpression and ELISA-based detection of asprosin in cultured cells and mice

Ila Mishra; Atul R. Chopra

doi:10.1016/j.xpro.2022.101762

Overexpression and ELISA-based detection of asprosin in cultured cells and mice

Ila Mishra
, Atul R. Chopra

Producción científica: Article › revisión exhaustiva

3 Citas (Scopus)

Resumen

The unreliability of commercial recombinant asprosin preparations and variability between asprosin detection assays have proven to be a bottleneck in experimental interpretation. This protocol describes the use of viral vectors and expression plasmid for overexpression and secretion of human asprosin to achieve sustained elevation of asprosin protein in mice and HEK293T cells without using recombinant proteins. This protocol also includes a sandwich ELISA using anti-asprosin monoclonal antibodies for detection of asprosin in media from cultured cells and in plasma of mice. For complete details on the use and execution of this protocol, please refer to Duerrschmid et al. (2017), Mishra et al. (2021), and Mishra et al. (2022).

Idioma original	English
Número de artículo	101762
Publicación	STAR Protocols
Volumen	3
N.º	4
DOI	https://doi.org/10.1016/j.xpro.2022.101762
Estado	Published - dic 16 2022

Nota bibliográfica

Publisher Copyright:
© 2022 The Author(s)

Financiación

This work was supported by the NIDDK ( DK102529 , DK118290 ). We thank members of the Chopra lab for helpful suggestions and critical reading of the manuscript. We thank Zhiqiang An for rabbit monoclonal anti-asprosin antibody development. Summary figure created using Biorender.com . This work was supported by the NIDDK (DK102529, DK118290). We thank members of the Chopra lab for helpful suggestions and critical reading of the manuscript. We thank Zhiqiang An for rabbit monoclonal anti-asprosin antibody development. Summary figure created using Biorender.com. Investigation, methodology, and writing, I.M.; conceptualization, resources, supervision, funding acquisition, and writing – review and editing, A.R.C. A.R.C. has been awarded asprosin-related patents and is a co-founder, director, and officer of Vizigen, Inc. and Aceragen, Inc. and holds equity in both companies.

Financiadores	Número del financiador
A.R.C. A.R.C.
National Institute of Diabetes and Digestive and Kidney Diseases	DK118290, DK102529
National Institute of Diabetes and Digestive and Kidney Diseases

ASJC Scopus subject areas

General Neuroscience
General Biochemistry, Genetics and Molecular Biology
General Immunology and Microbiology

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abstract = "The unreliability of commercial recombinant asprosin preparations and variability between asprosin detection assays have proven to be a bottleneck in experimental interpretation. This protocol describes the use of viral vectors and expression plasmid for overexpression and secretion of human asprosin to achieve sustained elevation of asprosin protein in mice and HEK293T cells without using recombinant proteins. This protocol also includes a sandwich ELISA using anti-asprosin monoclonal antibodies for detection of asprosin in media from cultured cells and in plasma of mice. For complete details on the use and execution of this protocol, please refer to Duerrschmid et al. (2017), Mishra et al. (2021), and Mishra et al. (2022).",

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Overexpression and ELISA-based detection of asprosin in cultured cells and mice

Resumen

Nota bibliográfica

Financiación

ASJC Scopus subject areas

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