Microchip electrophoresis separation of a panel of preterm birth biomarkers

Abstract

Preterm birth (PTB) is responsible for over one million infant deaths annually worldwide. Often, the first and only indication of PTB risk is the onset of early labor. Thus, there is an urgent need for an early PTB risk diagnostic that is inexpensive, reliable, and robust. Here, we describe the development of a microchip electrophoresis (μCE) method for separating a mixture of six PTB protein and peptide biomarkers present in maternal blood serum. μCE devices were photografted with a poly(ethylene glycol) diacrylate surface coating to regulate EOF and reduce nonspecific analyte adsorption. Separation conditions including buffer pH, buffer concentration, and applied electric field were varied to improve biomarker peak resolution while minimizing deleterious effects like Joule heating. In this way, it was possible to separate six PTB biomarkers, the first μCE separation of this biomarker panel. LODs were also measured for each of the six PTB biomarkers. In the future, this μCE separation can be integrated with upstream maternal blood serum sample preparation steps to yield a complete PTB risk diagnosis microdevice.

Analysis of thrombin‐antithrombin complex formation using microchip electrophoresis and mass spectrometry

“…Fabrication of “T‐shaped” devices was performed as in previous papers . In summary, we used standard photolithography and etching techniques to create silicon wafer templates.…”

“…Before use, the microchip channels were passivated. Channels were filled with a 4% poly(ethylene glycol) diacrylate, 1% benzoin methyl ether, and 50:50 v/v water/methanol solution and placed under a UV lamp (Uvitron, West Springfield, MA) for 12 min. The channels were then vacuumed out and flushed with HEPES buffer.…”

“…The molar ratios of fluorescent dye to biomarkers were P2 3:2, Fer 30:1, LF 15:1, CRF 3:2, and TNF 20:1. Cutoff filters were used corresponding to their respective molecular weights .…”

“…An automated, integrated microfluidic device could be developed to extract the biomarkers from blood serum, fluorescently label them, and separate them using microchip electrophoresis (µCE). Each of these three steps has been demonstrated separately (but not in a fully integrated platform) for select PTB biomarkers .…”

“…Six of the nine PTB risk biomarkers have previously been separated using µCE . One of the three missing biomarkers in this prior separation is TAT, which was not included because it is not available commercially.…”