Single cell analysis in full body quartz glass chips with native UV laser-induced fluorescence detection.
In order to investigate the individual and inhomogenous cellular response, e.g. to external stimuli, single cell analysis is mandatory and may provide new cognitions in proteomics as well as in other fields of systems biology in the future. Here, we report on novel chip architectures for single cell analysis based on full body quartz glass microfluidic chips (QG chips) that extend our previous studies in polydimethylsiloxane (PDMS) chips, and enhance the detection sensitivity of native UV laser-induced fluorescence (UV-LIF) detection. Detection of a 10nM tryptophan solution with an S/N ratio of 11.9, which gives a theoretical limit of detection of 2.5 nM (with S/N=3), was possible. With these optimizations the three proteins alpha-chymotrypsinogen A, ovalbumin and catalase each at a concentration of 100 microg/mL (equal to 4 microM, 0.4 microM and 2.2 microM) were injected electrokinetically and could be separated with nearly baseline resolution. Furthermore, fluorescence spectra (excitation wavelength, lambda(ex) = 266 nm) clearly demonstrate the favourable properties like the very high UV transparency and the nearly vanishing background fluorescence of the QG chips as compared to PDMS chips and to PDMS quartz window (PQW) chips. Finally we exploit the improved sensitivity for single cell electropherograms of Spodoptera frugiperda (Sf9) insect cells.
Greif
Dominik
D
Experimental Biophysics and Applied Nanoscience, Physics Department, Bielefeld University, Universitaetsstrasse 25, 33615 Bielefeld, NRW, Germany. dgreif@physik.uni-bielefeld.de
Galla
Lukas
L
Ros
Alexandra
A
Anselmetti
Dario
D
D000818:Animals; D002374:Catalase / Q000032:analysis / Q000302:isolation & purification; D002460:Cell Line; D002477:Cells / Q000737:chemistry*; D002919:Chymotrypsinogen / Q000032:analysis / Q000302:isolation & purification; D004586:Electrophoresis; D007834:Lasers; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation / Q000379:methods; D008856:Microscopy, Fluorescence / Q000379:methods; D010047:Ovalbumin / Q000032:analysis / Q000302:isolation & purification; D011506:Proteins / Q000032:analysis* / Q000302:isolation & purification; D011791:Quartz; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence / Q000379:methods; D014364:Tryptophan / Q000032:analysis; D014466:Ultraviolet Rays
D011506:Proteins; D011791:Quartz; D014364:Tryptophan; D010047:Ovalbumin; D002919:Chymotrypsinogen; D002374:Catalase