TY - JOUR A1 - Nayak, S A1 - Sashidhar, R A1 - Bhat, K T1 - Development of an immunoanalytical method for the detection of β- and γ- Crystallins and anti-crystallin antibodies. A molecular biomarker for cataract Y1 - 2002/1/1 JF - Indian Journal of Ophthalmology JO - Indian J Ophthalmol SP - 41 EP - 48 VL - 50 IS - 1 UR - https://journals.lww.com/ijo/pages/default.aspx/article.asp?issn=0301-4738;year=2002;volume=50;issue=1;spage=41;epage=48;aulast=Nayak N2 - 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 ER -