Human Phosphatidylinositol-binding clathrin assembly protein (PICALM) ELISA Kit

Dreyling et al. (1996) identified a breakpoint on chromosome 11 involving the previously unknown gene, CALM. CALM has a very high homology to the murine clathrin assembly protein ap3. Mao et al. (2001) determined the crystal structure of the N-terminal domain (the NAP domain) of the Drosophila Lap protein, which is homologous to human PICALM. The structure revealed a novel fold consisting of a large double layer of sheets of 10 alpha helices and a unique site for binding phosphoinositides.This motif is found in other proteins predicted to have domains of similar structure (e.g., Huntingtin-interacting protein-1) . The structure is in part similar to the epsin N-terminal (ENTH) domain, but epsin lacks the phosphatidylinositol-4,5-bisphosphate-binding site.

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Human Phosphatidylinositol-binding clathrin assembly protein (PICALM) ELISA Kit has high sensitivity and excellent specificity for detection of Human PICALM. No significant cross-reactivity or interference between Human PICALM and analogues was observed.

This Human Phosphatidylinositol-binding clathrin assembly protein (PICALM) ELISA Kit employs a two-site sandwich ELISA to quantitate PICALM in samples. An antibody specific for PICALM has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and anyPICALM present is bound by the immobilized antibody. After removing any unbound substances, a biotin-conjugated antibody specific for PICALM is added to the wells. After washing, Streptavidin conjugated Horseradish Peroxidase (HRP) is added to the wells. Following a wash to remove any unbound avidin-enzyme reagent, a substrate solution is added to the wells and color develops in proportion to the amount of PICALM bound in the initial step. The color development is stopped and the intensity of the color is measured.

• Do not mix components from different kit lots or use reagents beyond the kit expiration date.• Allow all reagents to warm to room temperature for at least 30 minutes before opening.• Pre-rinse the pipet tip with reagent, use fresh pipet tips for each sample, standard and reagent to avoid contamination.• Unused wells must be kept desiccated at 4 °C in the sealed bag provided.• Mix Thoroughly is very important for the result. It is recommended using low frequency oscillator or slight hand shaking every 10 minutes.• It is recommended that all samples and standards be assayed in duplicate or triplicate.