| A device is developed that combines single molecule detection with microfluidic manipulation to obtain absolute quantification of proteins within a single cell. It utilizes microvalve design and a picoliter-sized reaction chamber to capture and lyse a cell and then labels its contents with specific fluorescent probes. Capillary electrophoresis separation is employed to enable simultaneous analysis of multiple targets. The use of cylindrical optics in fluorescent detection allows high efficiency single molecule counting in a microfluidic channel with the dimension of micrometers. This device can determine the absolute copy number of proteins at very low concentrations, that is, one to several thousand copies per cell, without amplification procedures. Using this device, we demonstrated the analysis of phycobiliprotein complexes in the cyanobacteria, Synechococcus sp. PCC 7942. We are able to quantify the phycobilisome core subcomplexes when the cyanobacteria grow under nitrogen depleted conditions. Marked cell to cell variations are observed and rare cells that show incomplete degradation of phycobilisome are identified. |
