Large dynamic range digital nanodot gradients of biomolecules made by low-cost nanocontact printing for cell haptotaxis.

A novel method is introduced for ultrahigh throughput and ultralow cost patterning of biomolecules with nanometer resolution and novel 2D digital nanodot gradients (DNGs) with mathematically defined slopes are created. The technique is based on lift-off nanocontact printing while using high-resolution photopolymer stamps that are rapidly produced at a low cost through double replication from Si originals. Printed patterns with 100 nm features are shown. DNGs with varying spacing between the dots and a record dynamic range of 4400 are produced; 64 unique DNGs, each with hundreds of thousands of dots, are inked and printed in 5.5 min. The adhesive response and haptotaxis of C2C12 myoblast cells on DNGs demonstrated their biofunctionality. The great flexibility in pattern design, the massive parallel ability, the ultra low cost, and the extreme ease of polymer lift-off nanocontact printing will facilitate its use for various biological and medical applications.

Assessment of sphingosine-1-phosphate receptor expression and associated intracellular signaling cascades in primary cells of the human central nervous system.

Measuring the effects of sphingosine-1-phosphate (S1P) receptor modulators on human primary neural cells is of particular interest given the recent application of these central nervous system-accessible agents to the treatment of neurodegenerative diseases, such as multiple sclerosis. Issues to consider in experimental studies include the ability of some of these modulators to bind multiple S1P receptor subtypes simultaneously, the nonspecificity of commercially available S1P receptor antibodies, and activation of multiple intracellular signaling cascades by a given S1P receptor. Here, we discuss how to assay S1P receptor expression and activation using multiple agonists/antagonists, by linking the results of real-time reverse transcriptase polymerase chain reaction with the assessment of intracellular signaling derived from Western blot analyses.