Determination of desmosine in bronchoalveolar lavage fluids by time-resolved fluoroimmunoassay
Urinary excretion of desmosine has been reported to be increased in patients with pulmonary fibrosis; however, several investigators have pointed out that measuring urinary desmosine is not a very useful indicator of lung wall destruction. We developed a sensitive time-resolved fluoroimmunoassay (TR...
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| Published in | Clinica Chimica Acta Vol. 387; no. 1-2; pp. 113 - 119 |
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| Main Authors | , , , , |
| Format | Journal Article |
| Language | English |
| Published |
Netherlands
Elsevier B.V
01.01.2008
Elsevier BV |
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| Online Access | Get full text |
| ISSN | 0009-8981 1873-3492 |
| DOI | 10.1016/j.cca.2007.09.015 |
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| Abstract | Urinary excretion of desmosine has been reported to be increased in patients with pulmonary fibrosis; however, several investigators have pointed out that measuring urinary desmosine is not a very useful indicator of lung wall destruction. We developed a sensitive time-resolved fluoroimmunoassay (TR-FIA) to identify trace amounts of desmosine in bronchoalveolar lavage fluid (BALF), and applied this method to analyze BALF samples from healthy subjects and patients with interstitial lung diseases.
In the proposed TR-FIA, a polystyrene strip was coated with desmosine-conjugated gelatin. The strip was then incubated with rabbit anti-desmosine antibody and the test solution. The desmosine bound to the solid phase and free desmosine in the sample or standard solution were allowed to compete to bind to the anti-desmosine. The solid-phase antibody was detected by Eu-complex conjugated anti-rabbit IgG.
The detectable limit of desmosine was 50 fmol/ml in the TR-FIA developed in this study. TR-FIA showed low cross-reactivity against amino acids. BALF desmosine levels were significantly higher in patients with idiopathic pulmonary fibrosis and sarcoidosis compared with healthy subjects.
Desmosine levels in BALF may be useful to investigate lung disease. |
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| AbstractList | Urinary excretion of desmosine has been reported to be increased in patients with pulmonary fibrosis; however, several investigators have pointed out that measuring urinary desmosine is not a very useful indicator of lung wall destruction. We developed a sensitive time resolved fluoroimmunoassay (TR-FIA) to identify trace amounts of desmosine in bronchoalveolar lavage fluid (BALF), and applied this method to analyse BALF samples from healthy subjects and patients with interstitial lung diseases.
In the proposed TR-FIA, a polystyrene strip was coated with desmosine-conjugated gelatin. The strip was then incubated with rabbit anti-desmosine antibody and the test solution. The desmosine bound to the solid phase and free desmosine in the sample or standard solution were allowed to compete to bind to the anti-desmosine. The solid-phase antibody was detected by Eu-complex conjugated anti-rabbit IgG.
The detectable limit of desmosine was 50 fmol/ml in the TR-FIA developed in this study. TR-FIA showed low cross-reactivity against amino acids. BALF desmosine levels were significantly higher in patients with idiopathic fibrosis and sarcoidosis compared with healthy subjects.
Desmosine levels in BALF may be useful to investigate lung disease. Urinary excretion of desmosine has been reported to be increased in patients with pulmonary fibrosis; however, several investigators have pointed out that measuring urinary desmosine is not a very useful indicator of lung wall destruction. We developed a sensitive time resolved fluoroimmunoassay (TR-FIA) to identify trace amounts of desmosine in bronchoalveolar lavage fluid (BALF), and applied this method to analyse BALF samples from healthy subjects and patients with interstitial lung diseases.BACKGROUNDUrinary excretion of desmosine has been reported to be increased in patients with pulmonary fibrosis; however, several investigators have pointed out that measuring urinary desmosine is not a very useful indicator of lung wall destruction. We developed a sensitive time resolved fluoroimmunoassay (TR-FIA) to identify trace amounts of desmosine in bronchoalveolar lavage fluid (BALF), and applied this method to analyse BALF samples from healthy subjects and patients with interstitial lung diseases.In the proposed TR-FIA, a polystyrene strip was coated with desmosine-conjugated gelatin. The strip was then incubated with rabbit anti-desmosine antibody and the test solution. The desmosine bound to the solid phase and free desmosine in the sample or standard solution were allowed to compete to bind to the anti-desmosine. The solid-phase antibody was detected by Eu-complex conjugated anti-rabbit IgG.METHODSIn the proposed TR-FIA, a polystyrene strip was coated with desmosine-conjugated gelatin. The strip was then incubated with rabbit anti-desmosine antibody and the test solution. The desmosine bound to the solid phase and free desmosine in the sample or standard solution were allowed to compete to bind to the anti-desmosine. The solid-phase antibody was detected by Eu-complex conjugated anti-rabbit IgG.The detectable limit of desmosine was 50 fmol/ml in the TR-FIA developed in this study. TR-FIA showed low cross-reactivity against amino acids. BALF desmosine levels were significantly higher in patients with idiopathic fibrosis and sarcoidosis compared with healthy subjects.RESULTSThe detectable limit of desmosine was 50 fmol/ml in the TR-FIA developed in this study. TR-FIA showed low cross-reactivity against amino acids. BALF desmosine levels were significantly higher in patients with idiopathic fibrosis and sarcoidosis compared with healthy subjects.Desmosine levels in BALF may be useful to investigate lung disease.CONCLUSIONSDesmosine levels in BALF may be useful to investigate lung disease. Urinary excretion of desmosine has been reported to be increased in patients with pulmonary fibrosis; however, several investigators have pointed out that measuring urinary desmosine is not a very useful indicator of lung wall destruction. We developed a sensitive time-resolved fluoroimmunoassay (TR-FIA) to identify trace amounts of desmosine in bronchoalveolar lavage fluid (BALF), and applied this method to analyze BALF samples from healthy subjects and patients with interstitial lung diseases. In the proposed TR-FIA, a polystyrene strip was coated with desmosine-conjugated gelatin. The strip was then incubated with rabbit anti-desmosine antibody and the test solution. The desmosine bound to the solid phase and free desmosine in the sample or standard solution were allowed to compete to bind to the anti-desmosine. The solid-phase antibody was detected by Eu-complex conjugated anti-rabbit IgG. The detectable limit of desmosine was 50 fmol/ml in the TR-FIA developed in this study. TR-FIA showed low cross-reactivity against amino acids. BALF desmosine levels were significantly higher in patients with idiopathic pulmonary fibrosis and sarcoidosis compared with healthy subjects. Desmosine levels in BALF may be useful to investigate lung disease. |
| Author | Saito, Yoshihiro Sato, Takaji Chikuma, Masahiko Nagai, Sonoko Kajikuri, Takashi |
| Author_xml | – sequence: 1 givenname: Takaji surname: Sato fullname: Sato, Takaji email: sato@gly.oups.ac.jp organization: Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki 569-1094, Japan – sequence: 2 givenname: Takashi surname: Kajikuri fullname: Kajikuri, Takashi organization: Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki 569-1094, Japan – sequence: 3 givenname: Yoshihiro surname: Saito fullname: Saito, Yoshihiro organization: Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki 569-1094, Japan – sequence: 4 givenname: Masahiko surname: Chikuma fullname: Chikuma, Masahiko organization: Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki 569-1094, Japan – sequence: 5 givenname: Sonoko surname: Nagai fullname: Nagai, Sonoko organization: Department of Respiratory Medicine, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto, 606-8507, Japan |
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| Keywords | PBS Time-resolved fluoroimmunoassay Desmosine BALF Bronchoalveolar lavage fluid Interstitial lung disease IPF TR-FIA |
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| SubjectTerms | Bronchoalveolar Lavage Fluid Bronchoalveolar Lavage Fluid - chemistry Desmosine Desmosine - analysis Fluorescent Antibody Technique Fluorescent Antibody Technique - methods Humans Hydrolysis Interstitial lung disease Reproducibility of Results Sensitivity and Specificity Time-resolved fluoroimmunoassay |
| Title | Determination of desmosine in bronchoalveolar lavage fluids by time-resolved fluoroimmunoassay |
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