TY - JOUR
T1 - Improved native UV laser induced fluorescence detection for single cell analysis in poly(dimethylsiloxane) microfluidic devices
AU - Hellmich, Wibke
AU - Greif, Dominik
AU - Pelargus, Christoph
AU - Anselmetti, Dario
AU - Ros, Alexandra
N1 - Funding Information:
The authors acknowledge financial support from the Deutsche Forschungsgemeinschaft within the project “Microchip UV-LIF” (An-370/1-2). Sf9 cells were generously donated by Nickels Jensen and Prof. Karsten Niehaus from the Department of Genetics at Bielefeld University.
PY - 2006/10/20
Y1 - 2006/10/20
N2 - Single cell analytics is a key method in the framework of proteom research allowing analyses, which are not subjected to ensemble-averaging, cell-cycle or heterogeneous cell-population effects. Our previous studies on single cell analysis in poly(dimethylsiloxane) microfluidic devices with native label-free laser induced fluorescence detection [W. Hellmich, C. Pelargus, K. Leffhalm, A. Ros, D. Anselmetti, Electrophoresis 26 (2005) 3689] were extended in order to improve separation efficiency and detection sensitivity. Here, we particularly focus on the influence of poly(oxyethylene) based coatings on the separation performance. In addition, the influence on background fluorescence is studied by the variation of the incident laser power as well as the adaptation of the confocal volume to the microfluidic channel dimensions. Last but not least, the use of carbon black particles further enhanced the detection limit to 25 nM, thereby reaching the relevant concentration ranges necessary for the label-free detection of low abundant proteins in single cells. On the basis of these results, we demonstrate the first electropherogram from an individual Spodoptera frugiperda (Sf9) cell with native label-free UV-LIF detection in a microfluidic chip.
AB - Single cell analytics is a key method in the framework of proteom research allowing analyses, which are not subjected to ensemble-averaging, cell-cycle or heterogeneous cell-population effects. Our previous studies on single cell analysis in poly(dimethylsiloxane) microfluidic devices with native label-free laser induced fluorescence detection [W. Hellmich, C. Pelargus, K. Leffhalm, A. Ros, D. Anselmetti, Electrophoresis 26 (2005) 3689] were extended in order to improve separation efficiency and detection sensitivity. Here, we particularly focus on the influence of poly(oxyethylene) based coatings on the separation performance. In addition, the influence on background fluorescence is studied by the variation of the incident laser power as well as the adaptation of the confocal volume to the microfluidic channel dimensions. Last but not least, the use of carbon black particles further enhanced the detection limit to 25 nM, thereby reaching the relevant concentration ranges necessary for the label-free detection of low abundant proteins in single cells. On the basis of these results, we demonstrate the first electropherogram from an individual Spodoptera frugiperda (Sf9) cell with native label-free UV-LIF detection in a microfluidic chip.
KW - Microfluidic
KW - Native laser induced fluorescence
KW - Protein
KW - Single cell analysis
UR - http://www.scopus.com/inward/record.url?scp=33748955692&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=33748955692&partnerID=8YFLogxK
U2 - 10.1016/j.chroma.2006.06.008
DO - 10.1016/j.chroma.2006.06.008
M3 - Article
C2 - 16814305
AN - SCOPUS:33748955692
SN - 0021-9673
VL - 1130
SP - 195
EP - 200
JO - Journal of Chromatography A
JF - Journal of Chromatography A
IS - 2 SPEC. ISS.
ER -