Development of ELISA Systems for Measurement of Human Tumor Necrosis Factor-Alpha (TNF-a)

Ayşegül YÜCEL; Cemalettin AYBAY

Article Details

Development of ELISA Systems for Measurement of Human Tumor Necrosis Factor-Alpha (TNF-a)

Authors:

Ayşegül YÜCEL | Cemalettin AYBAY

Publisher:

TUBITAK

Journal:

Turkish Journal of Medical Sciences 2005 Vol.35 Issue 6, pp.385-393

Document Type:

Article

Publication Date:

5/2005

Document Language:

Turkish

Document ID (Org No.):

2000756299

Frequency:

Bimonthly

eISSN:

1303-6165

pISSN:

1300-0144

Keywords:

TNF-a
ELISA

Abstract:

 

The aim of this study was to develop a human tumor necrosis factor alpha (hTNF-a) ELISA system because specific and sensitive measurement of low levels of circulating TNF-a is very important for enlightening the immunopathologial mechanisms associated with TNF-a. Monoclonal antibodies 6A4c and 8A6 were produced against hTNF-a and were used as the capture antibody and the tracer antibody respectively for the first hTNF-a ELISA system (6A4c/biotin-8A6). Murine polyclonal IgG was used as the tracer antibody in the second hTNF-a ELISA system (6A4c/biotin-polyclonal IgG). Both systems could detect both recombinant hTNF-a and native hTNF-a. The detection limits defined as minimal concentration of hTNF-a were less than 4 pg/ml for the first and less than 12 pg/ml for the second ELISA systems. 6A4c/biotin-8A6 system resulted in some non-specific reactions to some extent with human sera; however, 6A4c/biotin-polyclonal IgG system produced acceptable background levels with human sera. A prominent inhibitory effect of TNF receptors-I and -II did not occur in any of the ELISA systems at physiological concentrations. Two different types of ELISA systems with high sensitivity and specificity were developed to measure hTNF-a level both in human serum and cell culture supernatant.

 

The aim of this study was to develop a human tumor necrosis factor alpha (hTNF-a) ELISA system because specific and sensitive measurement of low levels of circulating TNF-a is very important for enlightening the immunopathologial mechanisms associated with TNF-a. Monoclonal antibodies 6A4c and 8A6 were produced against hTNF-a and were used as the capture antibody and the tracer antibody respectively for the first hTNF-a ELISA system (6A4c/biotin-8A6). Murine polyclonal IgG was used as the tracer antibody in the second hTNF-a ELISA system (6A4c/biotin-polyclonal IgG). Both systems could detect both recombinant hTNF-a and native hTNF-a. The detection limits defined as minimal concentration of hTNF-a were less than 4 pg/ml for the first and less than 12 pg/ml for the second ELISA systems. 6A4c/biotin-8A6 system resulted in some non-specific reactions to some extent with human sera; however, 6A4c/biotin-polyclonal IgG system produced acceptable background levels with human sera. A prominent inhibitory effect of TNF receptors-I and -II did not occur in any of the ELISA systems at physiological concentrations. Two different types of ELISA systems with high sensitivity and specificity were developed to measure hTNF-a level both in human serum and cell culture supernatant.

Article Details

Development of ELISA Systems for Measurement of Human Tumor Necrosis Factor-Alpha (TNF-a)

Authors: Ayşegül YÜCEL | Cemalettin AYBAY

Publisher: TUBITAK

Journal: Turkish Journal of Medical Sciences 2005 Vol.35 Issue 6, pp.385-393

Document Type: Article

Publication Date: 5/2005

Document Language: Turkish

Document ID (Org No.): 2000756299

Frequency: Bimonthly

eISSN: 1303-6165

pISSN: 1300-0144

TNF-a

ELISA

Issues

Authors:

Ayşegül YÜCEL | Cemalettin AYBAY

Publisher:

TUBITAK

Journal:

Turkish Journal of Medical Sciences 2005 Vol.35 Issue 6, pp.385-393

Document Type:

Article

Publication Date:

5/2005

Document Language:

Turkish

Document ID (Org No.):

2000756299

Frequency:

Bimonthly

eISSN:

1303-6165

pISSN:

1300-0144