Automated microdroplet platform for sample manipulation and polymerase chain reaction.
We present a fully automated system performing continuous sampling, reagent mixing, and polymerase chain reaction (PCR) in microdroplets transported in immiscible oil. Sample preparation and analysis are totally automated, using an original injection method from a modified 96-well plate layered with three superimposed liquid layers and in-capillary laser-induced fluorescence endpoint detection. The process is continuous, allowing sample droplets to be carried uninterruptedly into the reaction zone while new drops are aspirated from the sample plate. Reproducible amplification, negligible cross-contamination, and detection of low sample concentrations were demonstrated on numerous consecutive sample drops. The system, which opens the route to strong reagents and labor savings in high-throughput applications, was validated on the clinically relevant quantification of progesterone receptor gene expression in human breast cancer cell lines.
Chabert
Max
M
Laboratoire Physicochimie-Curie, UMR 168 IC/CNRS, Institut Curie Section de Recherche, 26, rue d'Ulm, 75005 Paris, France.
Dorfman
Kevin D
KD
de Cremoux
Patricia
P
Roeraade
Johan
J
Viovy
Jean-Louis
JL
D001331:Automation; D001943:Breast Neoplasms / Q000235:genetics / Q000473:pathology; D002460:Cell Line; D005260:Female; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D007834:Lasers; D008832:Microchemistry / Q000379:methods*; D009821:Oils / Q000737:chemistry; D016133:Polymerase Chain Reaction; D011980:Receptors, Progesterone / Q000378:metabolism; D015203:Reproducibility of Results; D013050:Spectrometry, Fluorescence / Q000379:methods; D013997:Time Factors
D009821:Oils; D011980:Receptors, Progesterone