Abstract
Current interest in novel compounds with dermal repair potential has led to the development of a rapid and accurate assay to screen large numbers of samples for newly synthesized tropoelastin in the dermis of natural and photo aged skin. Herein we report the development of a competitive enzyme‐linked immunosorbent assay (ELISA) for rodent tropoelastin using commercially available reagents. Utilizing a polyclonal antibody to mouse tropoelastin and rat lung alpha elastin (Elastin Products Company, Inc) we were able to quantify tropoelastin production in the supernatants of cultured skin and lysate of dermal punch biopsies. Briefly, micro titer plates were coated with antibody (10 micro g/ml) in carbonate buffer (pH 9.6) overnight at 4oC, washed and blocked with 1% BSA in PBS for 1 hour to reduce non‐specific binding. Equal volumes of samples or elastin (standards) and biotinylated elastin were simultaneously added to wells and incubated for 2 hours at room temperature. Plates were washed to remove unbound antigen and Streptavidin‐HRP followed by OPD substrate was used for detection. The range of detection of elastin standard curve was 0.1–200 micro g/ml with a coefficient of variation of < 3%. This assay will prove a valuable tool in determining the ability of novel compounds to increase dermal tropoelastin production and subsequently increase dermal elasticity. Work supported by NIH RO1 HL07097
Published Version
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