Simultaneous Multianalyte Detection with a Nanometer-Scale Pore

Abstract

It was recently shown that naturally occurring, genetically engineered or chemically modified channels can be used to detect analytes in solution. We demonstrate here that the overall range of analytes that can be detected by single nanometer-scale pores is expanded using a potentially simpler system. Instead of attaching recognition elements to a channel, they are covalently linked to polymers that otherwise thread through a nanometer-scale pore. Because the rate of unbound polymer entering the pore is proportional to its concentration in the bulk, the binding of analyte to the polymer alters the latter's ability to thread through the pore, and the signal that results from individual polymer translocation is unique to the polymer type; the method permits multianalyte detection and quantitation. We demonstrate here that two different proteins can be simultaneously detected with this technique.

Probing single nanometer-scale pores with polymeric molecular rulers

“…This and previous work demonstrated that the lifetimes of polynucleotide transits are typically 1-10 s per base. 40,59,63,64 …”

“…40 These events were digitally recorded and the lifetime, amplitude, and pattern of the ssDNA-induced current blockades were measured. [59][60][61] A. Membrane and channel formation Solvent-free planar lipid bilayer membranes 62 were formed from diphytanoyl phosphatidylcholine ͑Avanti Polar Lipids, Inc., Alabaster, AL͒ in pentane ͑Burdick and Jackson, Muskegon, MI͒. The membranes were made on 50-120 m diameter holes in a 17 m thick Teflon partition that separates two identical Teflon chambers.…”

“…57,59,65 Here, we provide more detailed experimental results that show how different length polymers can act as semiquantitative "molecular rulers" to determine how far the polymer must thread into the pore before it is committed to transport through the channel.…”

Theory of DNA electrophoresis (∼ 1999 –2002 ½)

“…Indeed, the target DNA binds to its complementary sequence and takes longer to translocate than the other chains. Another analysis method has been implemented by Kasianowicz' group [12]. In this case, DNA sequences are first designed to bind with a given analyte and are then allowed to translocate.…”

“…Therefore, in practice, this is a superb equation to use. An example of mobility-size data fitting using the new mobility relation (12). Double-stranded DNA fragments in the 200 bp to 48 kbp range were used.…”