FGF21 responsive reporter gene cell line

An assay method for detecting and quantitating an FGF21 antibody in a test sample, said method comprising:

• (i) providing a test sample;
• (ii) providing a first mammalian cell and a second mammalian cell, wherein said first mammalian cell and said second mammalian cell are independently selected from the group consisting of HEK293, Jurkat, K652 and U937, comprising:
• (1) a first heterologous polynucleotide comprising a heterologous cis-acting regulatory sequence operably linked to a downstream promoter sequence, wherein said promoter sequence is operably linked to an open reading frame encoding a first reporter protein,
• (2) a second heterologous polynucleotide encoding a chimeric transcription factor, wherein said chimeric transcription factor comprises a trans-activation domain of ternary complex factor (TCF) Elk-1 fused to a heterologous DNA binding domain capable of binding to said cis-acting regulatory sequence, wherein the DNA binding domain does not exist in mammalian cells, and
• (3) a cell surface bound heterodimeric receptor protein comprising a tyrosine kinase fibroblast growth factor receptor 1c (FGFR1c) and a beta-Klotho protein,
• wherein said cell comprises a heterologous polynucleotide comprising promoter operably linked to a down-stream open reading frame encoding the beta-Klotho protein and from which the beta-Klotho protein is expressed, and wherein the open reading frame in (iii) comprises the sequence set forth in SEQ ID NO.:
• 1;
• (iii) contacting the first mammalian cell with the test sample and subsequently with FGF21;
• (iv) contacting the second mammalian cell with FGF21;
• (v) determining the activity of the first reporter protein in the first mammalian cell and determining the activity of the first reporter protein in the second mammalian cell;
• and
• (vi) providing a ratio between the first reporter protein activity of the first mammalian cell and the second mammalian cell, wherein, if the ratio is less than one, then FGF21 antibodies are present in the test sample.

An assay method for detecting and quantitating an FGF21 antibody present in a test sample, said method comprising:

• (i) providing a test sample,
• (ii) providing a first mammalian cell and a second mammalian cell, wherein said first mammalian cell and said second mammalian cell are independently selected from the group consisting of HEK293, Jurkat, K652 and U937, comprising:
• (1) a first heterologous polynucleotide comprising a heterologous cis-acting regulatory sequence operably linked to a downstream promoter sequence, wherein said promoter sequence is operably linked to an open reading frame encoding a first reporter protein,
• (2) a second heterologous polynucleotide encoding a chimeric transcription factor, wherein said chimeric transcription factor comprises a trans-activation domain of ternary complex factor (TCF) Elk-1 fused to a heterologous DNA binding domain capable of binding to said cis-acting regulatory sequence, wherein the DNA binding domain does not exist in mammalian cells,
• (3) a cell surface bound heterodimeric receptor protein comprising a tyrosine kinase fibroblast growth factor receptor 1c (FGFR1c) and a beta-Klotho protein,
• wherein said cell comprises a heterologous polynucleotide comprising promoter operably linked to a down-stream open reading frame encoding the beta-Klotho protein and from which the beta-Klotho protein is expressed, and wherein the open reading frame in (iii) comprises the sequence set forth in SEQ ID NO:
• 1, and
• (4) a third heterologous polynucleotide encoding a promoter sequence that is operably linked to an open reading frame encoding a second reporter protein;
• (iii) contacting the first mammalian cell with the test sample and subsequently with FGF21;
• (iv) contacting the second mammalian cell with FGF21;
• (v) determining the activity of the first and second reporter proteins in the first mammalian cell and determining the activity of the first and second reporter proteins in the second mammalian cell;
• (vi) providing a first ratio between the reporter activity of the first and second reporter proteins in the first mammalian cell providing a second ratio between the reporter activity of the first and second reporter proteins in the second mammalian cell;
• and
• (vii) providing a third ratio between first and second ratio of step (vi), wherein, if the ratio is less than one, then FGF21 antibodies are present in the test sample, wherein the assay method is without interference from normal human serum.