Generation of monoclonal antibodies and development of an immunofluorometric assay for the detection of CUZD1 in tissues and biological fluids

Sofia Farkona, Antoninus Soosaipillai, Panagiota Filippou, Dimitrios Korbakis, Stefano Serra, Felix Rückert, Eleftherios P. Diamandis, Ivan M. Blasutig

Research output: Contribution to journalArticle

Abstract

Background CUB and zona pellucida-like domain-containing protein 1 (CUZD1) was identified as a pancreas-specific protein and was proposed as a candidate biomarker for pancreatic related disorders. CUZD1 protein levels in tissues and biological fluids have not been extensively examined. The purpose of the present study was to generate specific antibodies targeting CUZD1 to assess CUZD1 expression within tissues and biological fluids. Methods Mouse monoclonal antibodies against CUZD1 were generated and used to perform immunohistochemical analyses and to develop a sensitive and specific enzyme-linked immunosorbent assay (ELISA). CUZD1 protein expression was assessed in various human tissue extracts and biological fluids and in gel filtration chromatography-derived fractions of pancreatic tissue extract, pancreatic juice and recombinant protein. Results Immunohistochemical staining of CUZD1 in pancreatic tissue showed that the protein is localized to the acinar cells and the lumen of the acini. Western blot analysis detected the protein in pancreatic tissue extract and pancreatic juice. The newly developed ELISA measured CUZD1 in high levels in pancreas and in much lower but detectable levels in several other tissues. In the biological fluids tested, CUZD1 expression was detected exclusively in pancreatic juice. The analysis of gel filtration chromatography-derived fractions of pancreatic tissue extract, pancreatic juice and recombinant CUZD1 suggested that the protein exists in high molecular weight protein complexes. Conclusion This study describes the development of tools targeting CUZD1 protein, its tissue expression pattern and levels in several biological fluids. These new tools will facilitate future investigations aiming to delineate the role of CUZD1 in physiology and pathobiology.

Original languageEnglish
Pages (from-to)1168-1174
Number of pages7
JournalClinical Biochemistry
Volume50
Issue number18
DOIs
Publication statusPublished - 1 Dec 2017

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Fluoroimmunoassay
Assays
Pancreatic Juice
Monoclonal Antibodies
Tissue
Pancreatic Extracts
Fluids
Proteins
Immunosorbents
Gel Chromatography
Pancreas
Chromatography
Enzyme-Linked Immunosorbent Assay
Zona Pellucida
Tissue Extracts
Gels
Acinar Cells
Protein Transport
Recombinant Proteins
Physiology

Cite this

Farkona, Sofia ; Soosaipillai, Antoninus ; Filippou, Panagiota ; Korbakis, Dimitrios ; Serra, Stefano ; Rückert, Felix ; Diamandis, Eleftherios P. ; Blasutig, Ivan M. / Generation of monoclonal antibodies and development of an immunofluorometric assay for the detection of CUZD1 in tissues and biological fluids. In: Clinical Biochemistry. 2017 ; Vol. 50, No. 18. pp. 1168-1174.
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Generation of monoclonal antibodies and development of an immunofluorometric assay for the detection of CUZD1 in tissues and biological fluids. / Farkona, Sofia; Soosaipillai, Antoninus; Filippou, Panagiota; Korbakis, Dimitrios; Serra, Stefano; Rückert, Felix; Diamandis, Eleftherios P.; Blasutig, Ivan M.

In: Clinical Biochemistry, Vol. 50, No. 18, 01.12.2017, p. 1168-1174.

Research output: Contribution to journalArticle

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T1 - Generation of monoclonal antibodies and development of an immunofluorometric assay for the detection of CUZD1 in tissues and biological fluids

AU - Farkona, Sofia

AU - Soosaipillai, Antoninus

AU - Filippou, Panagiota

AU - Korbakis, Dimitrios

AU - Serra, Stefano

AU - Rückert, Felix

AU - Diamandis, Eleftherios P.

AU - Blasutig, Ivan M.

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Y1 - 2017/12/1

N2 - Background CUB and zona pellucida-like domain-containing protein 1 (CUZD1) was identified as a pancreas-specific protein and was proposed as a candidate biomarker for pancreatic related disorders. CUZD1 protein levels in tissues and biological fluids have not been extensively examined. The purpose of the present study was to generate specific antibodies targeting CUZD1 to assess CUZD1 expression within tissues and biological fluids. Methods Mouse monoclonal antibodies against CUZD1 were generated and used to perform immunohistochemical analyses and to develop a sensitive and specific enzyme-linked immunosorbent assay (ELISA). CUZD1 protein expression was assessed in various human tissue extracts and biological fluids and in gel filtration chromatography-derived fractions of pancreatic tissue extract, pancreatic juice and recombinant protein. Results Immunohistochemical staining of CUZD1 in pancreatic tissue showed that the protein is localized to the acinar cells and the lumen of the acini. Western blot analysis detected the protein in pancreatic tissue extract and pancreatic juice. The newly developed ELISA measured CUZD1 in high levels in pancreas and in much lower but detectable levels in several other tissues. In the biological fluids tested, CUZD1 expression was detected exclusively in pancreatic juice. The analysis of gel filtration chromatography-derived fractions of pancreatic tissue extract, pancreatic juice and recombinant CUZD1 suggested that the protein exists in high molecular weight protein complexes. Conclusion This study describes the development of tools targeting CUZD1 protein, its tissue expression pattern and levels in several biological fluids. These new tools will facilitate future investigations aiming to delineate the role of CUZD1 in physiology and pathobiology.

AB - Background CUB and zona pellucida-like domain-containing protein 1 (CUZD1) was identified as a pancreas-specific protein and was proposed as a candidate biomarker for pancreatic related disorders. CUZD1 protein levels in tissues and biological fluids have not been extensively examined. The purpose of the present study was to generate specific antibodies targeting CUZD1 to assess CUZD1 expression within tissues and biological fluids. Methods Mouse monoclonal antibodies against CUZD1 were generated and used to perform immunohistochemical analyses and to develop a sensitive and specific enzyme-linked immunosorbent assay (ELISA). CUZD1 protein expression was assessed in various human tissue extracts and biological fluids and in gel filtration chromatography-derived fractions of pancreatic tissue extract, pancreatic juice and recombinant protein. Results Immunohistochemical staining of CUZD1 in pancreatic tissue showed that the protein is localized to the acinar cells and the lumen of the acini. Western blot analysis detected the protein in pancreatic tissue extract and pancreatic juice. The newly developed ELISA measured CUZD1 in high levels in pancreas and in much lower but detectable levels in several other tissues. In the biological fluids tested, CUZD1 expression was detected exclusively in pancreatic juice. The analysis of gel filtration chromatography-derived fractions of pancreatic tissue extract, pancreatic juice and recombinant CUZD1 suggested that the protein exists in high molecular weight protein complexes. Conclusion This study describes the development of tools targeting CUZD1 protein, its tissue expression pattern and levels in several biological fluids. These new tools will facilitate future investigations aiming to delineate the role of CUZD1 in physiology and pathobiology.

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