We present an analytical study of peak mode isotachophoresis (ITP), and provide closed form solutions for sample distribution and electric field, as well as for leading-, trailing-, and counter-ion concentration profiles. Importantly, the solution we present is valid not only for the case of fully ionized species, but also for systems of weak electrolytes which better represent real buffer systems and for multivalent analytes such as proteins and DNA. The model reveals two major scales which govern the electric field and buffer distributions, and an additional length scale governing analyte distribution. Using well-controlled experiments, and numerical simulations, we verify and validate the model and highlight its key merits as well as its limitations. We demonstrate the use of the model for determining the peak concentration of focused sample based on known buffer and analyte properties, and show it differs significantly from commonly used approximations based on the interface width alone. We further apply our model for studying reactions between multiple species having different effective mobilities yet co-focused at a single ITP interface. We find a closed form expression for an effective-on rate which depends on reactants distributions, and derive the conditions for optimizing such reactions. Interestingly, the model reveals that maximum reaction rate is not necessarily obtained when the concentration profiles of the reacting species perfectly overlap. In addition to the exact solutions, we derive throughout several closed form engineering approximations which are based on elementary functions and are simple to implement, yet maintain the interplay between the important scales. Both the exact and approximate solutions provide insight into sample focusing and can be used to design and optimize ITP-based assays.

We present a novel microfluidic assay for continuous and quantitative detection of bacteria in water. We leverage isotachophoresis (ITP), an electrophoretic focusing technique, to create a stationary high concentration zone of fluorescently labeled antimicrobial peptides (AMPs) in a microfluidic channel. The tested water sample flows continuously through this high concentration AMPs reaction zone; any bacteria present in the sample is simultaneously labeled by, and separated from, the high concentration AMPs. The labeled bacteria continue into the downstream pure-buffer zone where the fluorescence signal is monitored, providing a direct quantitative measurement of the original bacterial concentration in the sample. We present the principles of the technique, demonstrate its applicability for quantitative detection of E. coli as well as its stability over a 1 h monitoring time, and provide a simple model for predicting its performance at different operating conditions. The method could be potentially expanded for use with other types of probes and provide continuous analysis and monitoring of water samples at the point of need.

We present a novel assay for rapid and high sensitivity detection of nucleic acids without amplification. Utilizing the neutral backbone of peptide nucleic acids (PNA), our method is based on the design of low electrophoretic mobility PNA probes, which do not focus under isotachophoresis (ITP) unless bound to their target sequence. Thus, background noise associated with free probes is entirely eliminated, significantly improving the signal-to-noise ratio while maintaining a simple single-step assay requiring no amplification steps. We provide a detailed analytical model and experimentally demonstrate the ability to detect targets as short as 17 nucleotides (nt) and a limit of detection of 100 fM with a dynamic range of 5 decades. We also demonstrate that the assay can be successfully implemented for detection of DNA in human serum without loss of signal. The assay requires 15 min to complete, and it could potentially be used in applications where rapid and highly sensitive amplification-free detection of nucleic acids is desired.