ELISA solid phase: stability and binding characteristics.
Stability of solid-phase antibody (SP-Ab) was investigated using polystyrene surface coated, by simple physical adsorption, with a rabbit antibody against the beta subunit of human chorionic gonadotropin. SP-Ab stored wet in 0.1 M Tris-HCl buffer, pH 7.2, or in 10% ammonium sulphate solution in the Tris buffer did not lose any activity upto the entire period of the study, i.e., 95 days at 6 degrees C and room temperature and 53 days at 40 degrees C. However, SP-Ab lost some activity upon drying and decrease in activity continued when air-dried SP-Ab was stored at any of these temperatures. Secondary coating of SP-Ab with sucrose or lactose before drying offered only partial protection to the activity of SP-Ab. Using radiolabeled antibody it was found that the loss of activity was not caused by desorption of the antibody; rather it could be the result of surface denaturation of the molecules. Binding of the antibody to polystyrene was so strong that week-long storage of SP-Ab in 6 M urea or 0.1% acetic acid could not bring about desorption by more than 1.8%. It is recommended that SP-Ab should be stored in a buffer and should not be allowed to dry.
A A
AA
Ansari
N S
NS
Hattikudur
S R
SR
Joshi
M A
MA
Medeira
D000327:Adsorption; D000818:Animals; D000906:Antibodies / Q000276:immunology*; D006063:Chorionic Gonadotropin / Q000032:analysis; D018997:Chorionic Gonadotropin, beta Subunit, Human; D003890:Desiccation; D004797:Enzyme-Linked Immunosorbent Assay / Q000295:instrumentation*; D006801:Humans; D007785:Lactose; D010446:Peptide Fragments / Q000032:analysis; D011137:Polystyrenes; D011309:Preservation, Biological / Q000379:methods*; D011489:Protein Denaturation; D011817:Rabbits; D013395:Sucrose; D013696:Temperature; D013997:Time Factors
D000906:Antibodies; D006063:Chorionic Gonadotropin; D018997:Chorionic Gonadotropin, beta Subunit, Human; D010446:Peptide Fragments; D011137:Polystyrenes; D013395:Sucrose; D007785:Lactose