Development of an Enzyme-Linked Immunosorbent Assay (ELISA) for the Quantification of ARID1A in Tissue Lysates

ARID1A is a subunit of the mammalian SWI/SNF complex, which is thought to regulate gene expression through restructuring chromatin structures. Its gene ARID1A is frequently mutated and ARID1A levels are lowered in several human cancers, especially gynecologic ones. A functional ARID1A loss may have...

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Bibliographic Details
Published in:Cancers 2023-08, Vol.15 (16), p.4096
Main Authors: Hinsberger, Manuel, Becker-Kettern, Julia, Jürgens-Wemheuer, Wiebke M, Oertel, Joachim, Schulz-Schaeffer, Walter J
Format: Article
Language:English
Subjects:
Amino acids
Analysis
Antigens
Apoptosis
Cell culture
Chromatin
Chromatin remodeling
Connective tissue
Enzyme-linked immunosorbent assay
Enzymes
Estrogens
Fractionation
Gene expression
Lysates
Mutation
Ovaries
Proteins
SWI/SNF complex
Tissue culture
Tumorigenesis
Tumors
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Summary:ARID1A is a subunit of the mammalian SWI/SNF complex, which is thought to regulate gene expression through restructuring chromatin structures. Its gene ARID1A is frequently mutated and ARID1A levels are lowered in several human cancers, especially gynecologic ones. A functional ARID1A loss may have prognostic or predictive value in terms of therapeutic strategies but has not been proposed based on a quantitative method. Hardly any literature is available on ARID1A levels in tumor samples. We developed an indirect enzyme-linked immunosorbent assay (ELISA) for ARID1A based on the current EMA and FDA criteria. We demonstrated that our ELISA provides the objective, accurate, and precise quantification of ARID1A concentrations in recombinant protein solutions, cell culture standards, and tissue lysates of tumors. A standard curve analysis yielded a ‘goodness of fit’ of R2 = 0.99. Standards measured on several plates and days achieved an inter-assay accuracy of 90.26% and an inter-assay precision with a coefficient of variation of 4.53%. When tumor lysates were prepared and measured multiple times, our method had an inter-assay precision with a coefficient of variation of 11.78%. We believe that our suggested method ensures a high reproducibility and can be used for a high sample throughput to determine the ARID1A concentration in different tumor entities. The application of our ELISA on various tumor and control tissues will allow us to explore whether quantitative ARID1A measurements in tumor samples are of predictive value.
ISSN:2072-6694
2072-6694
DOI:10.3390/cancers15164096