Development of an immunoanalytical method for the detection of β- and γ- Crystallins and anti-crystallin antibodies. A molecular biomarker for cataract
Purpose: To develop and evaluate an immunoanalytical method for the detection of β - and g-crystallins and anti-crystallin antibodies. Materials and Methods: Beta and g-crystallins isolated from rat lens were used as immunogens to raise polyclonal antibodies in rabbits. Antibody capt...
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| Format: | Article |
| Language: | English |
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Wolters Kluwer Medknow Publications
2002-01-01
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| Series: | Indian Journal of Ophthalmology |
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| Online Access: | http://www.ijo.in/article.asp?issn=0301-4738;year=2002;volume=50;issue=1;spage=41;epage=48;aulast=Nayak |
| _version_ | 1818148737551171584 |
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| author | Nayak Sujatha Sashidhar R Bhat Seetharam |
| author_facet | Nayak Sujatha Sashidhar R Bhat Seetharam |
| author_sort | Nayak Sujatha |
| collection | DOAJ |
| description | Purpose: To develop and evaluate an immunoanalytical method for the detection of β - and g-crystallins and anti-crystallin antibodies. Materials and Methods: Beta and g-crystallins isolated from rat lens were used as immunogens to raise polyclonal antibodies in rabbits. Antibody capture assay and western blot analysis showed that the antibodies to β - and g-crystallins were specific. An indirect competitive enzyme linked immunosorbent assay (ELISA) developed to quantitate β - and g-crystallin showed an IC50 value of 70 ng and 65 ng, respectively, based on regression analysis. Spiking studies with purified β -crystallin antibodies showed that 33 ng of the purified antibody gave an absorbance of 1.1 at 450 nm, indicating the sensitivity of the method. Results: Antibodies to β - and g-crystallins were not detected in serum samples of the cataractous CFY/NIN rats (used as an animal model for induction of experimental cataract by feeding high galactose diet). However, the cataractous rat serum samples effectively displaced β - and g-crystallin antibodies, indicating that these crystallins leak during cataract formation. The concentration of β - and g-crystallins in the rat serum, as analysed by indirect competitive ELISA, was found to be in the range of 17.6 - 81.6 mg/µl and 12.4- 19.6 µg/ml, respectively. Conclusions: The methodology developed in the present study may find application as a biochemical tool in molecular epidemiology of cataract |
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| institution | Directory Open Access Journal |
| issn | 0301-4738 |
| language | English |
| last_indexed | 2024-12-11T12:55:54Z |
| publishDate | 2002-01-01 |
| publisher | Wolters Kluwer Medknow Publications |
| record_format | Article |
| series | Indian Journal of Ophthalmology |
| spelling | doaj.art-73b08ea054d341c5899543ffc1cbb6a22022-12-22T01:06:35ZengWolters Kluwer Medknow PublicationsIndian Journal of Ophthalmology0301-47382002-01-015014148Development of an immunoanalytical method for the detection of β- and γ- Crystallins and anti-crystallin antibodies. A molecular biomarker for cataractNayak SujathaSashidhar RBhat SeetharamPurpose: To develop and evaluate an immunoanalytical method for the detection of β - and g-crystallins and anti-crystallin antibodies. Materials and Methods: Beta and g-crystallins isolated from rat lens were used as immunogens to raise polyclonal antibodies in rabbits. Antibody capture assay and western blot analysis showed that the antibodies to β - and g-crystallins were specific. An indirect competitive enzyme linked immunosorbent assay (ELISA) developed to quantitate β - and g-crystallin showed an IC50 value of 70 ng and 65 ng, respectively, based on regression analysis. Spiking studies with purified β -crystallin antibodies showed that 33 ng of the purified antibody gave an absorbance of 1.1 at 450 nm, indicating the sensitivity of the method. Results: Antibodies to β - and g-crystallins were not detected in serum samples of the cataractous CFY/NIN rats (used as an animal model for induction of experimental cataract by feeding high galactose diet). However, the cataractous rat serum samples effectively displaced β - and g-crystallin antibodies, indicating that these crystallins leak during cataract formation. The concentration of β - and g-crystallins in the rat serum, as analysed by indirect competitive ELISA, was found to be in the range of 17.6 - 81.6 mg/µl and 12.4- 19.6 µg/ml, respectively. Conclusions: The methodology developed in the present study may find application as a biochemical tool in molecular epidemiology of cataracthttp://www.ijo.in/article.asp?issn=0301-4738;year=2002;volume=50;issue=1;spage=41;epage=48;aulast=NayakCataractanti-crystallin antibodyβ -crystallinsg-crystallins |
| spellingShingle | Nayak Sujatha Sashidhar R Bhat Seetharam Development of an immunoanalytical method for the detection of β- and γ- Crystallins and anti-crystallin antibodies. A molecular biomarker for cataract Indian Journal of Ophthalmology Cataract anti-crystallin antibody β -crystallins g-crystallins |
| title | Development of an immunoanalytical method for the detection of β- and γ- Crystallins and anti-crystallin antibodies. A molecular biomarker for cataract |
| title_full | Development of an immunoanalytical method for the detection of β- and γ- Crystallins and anti-crystallin antibodies. A molecular biomarker for cataract |
| title_fullStr | Development of an immunoanalytical method for the detection of β- and γ- Crystallins and anti-crystallin antibodies. A molecular biomarker for cataract |
| title_full_unstemmed | Development of an immunoanalytical method for the detection of β- and γ- Crystallins and anti-crystallin antibodies. A molecular biomarker for cataract |
| title_short | Development of an immunoanalytical method for the detection of β- and γ- Crystallins and anti-crystallin antibodies. A molecular biomarker for cataract |
| title_sort | development of an immunoanalytical method for the detection of x0026 946 and x0026 947 crystallins and anti crystallin antibodies a molecular biomarker for cataract |
| topic | Cataract anti-crystallin antibody β -crystallins g-crystallins |
| url | http://www.ijo.in/article.asp?issn=0301-4738;year=2002;volume=50;issue=1;spage=41;epage=48;aulast=Nayak |
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