Molecular Analysis using a Magnetic Sifter and Nanowell System

摘要

A method for identification of circulating tumor cells (CTCs) in a blood sample uses magnetic enrichment and a nanowell assay. The CTCs are magnetically labeled with cancer cell markers conjugated to magnetic nanoparticles and then separated by passing the blood sample through a magnetic sifter. The enriched CTCs are then loaded into a microfluidic single-cell molecular assay comprising an array of 25,600 or more nanowells, each containing at most a single one of the CTCs. Using multiple fluorescent gene markers, simultaneous multiple-color multiplexed gene expression of the CTCs is performed, preferably using RT-PCR. Images of fluorescence signals from individual nanowells are analyzed to identify CTCs.

主权项

1. A method for identification of circulating tumor cells (CTCs), the method comprising:
magnetically labeling the CTCs in a blood sample with cancer cell markers conjugated to magnetic nanoparticles; separating the magnetically labeled CTCs by passing the blood sample through a magnetic sifter during application of an external magnetic field; collecting the separated magnetically labeled CTCs to produce enriched CTCs; loading the enriched CTCs into a microfluidic single-cell molecular assay comprising an array of 25,600 or more nanowells, where each of the nanowells is adapted to contain at most a single one of the CTCs; performing multiple simultaneous multiple-color multiplexed gene expression of the CTCs using the microfluidic single-cell molecular assay and multiple fluorescent gene markers; imaging the array of nanowells using fluorescence signal acquisition from individual nanowells, producing images of the array of nanowells; analyzing the images using a signal processor to identify CTCs based on the concurrent expression of two or more genes.