| DOI | Resolve DOI: https://doi.org/10.1109/JPROC.2003.820551 |
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| Author | Search for: Ocvirk, Gregor; Search for: Salimi-Moosavi, Hossein; Search for: Szarka, Rod J.; Search for: Arriaga, Edgar A.; Search for: Andersson, Per E.; Search for: Smith, Richard; Search for: Dovichi, Norman J.; Search for: Harrison, D. Jed |
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| Format | Text, Article |
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| Subject | microfluidics; galactosidase; lab on a chip; single-cell analysis |
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| Abstract | The use of microfluidic glass chips for continuous single-cell lysis and assay of internal β-Galactosidase (β-Gal) content is described. Cells were transported single file toward a Y-shaped mixing junction at which lytic agents were introduced by suction. Flow velocities of ∼100 and ∼40 μm/s were used under protein denaturing [35 mM sodium dodecylsulfate (SDS)] and nondenaturing (0.1% Triton X-100) conditions, respectively. Complete and reproducible lysis of individual cells on-chip occurred within 30 s using Triton X-100 and 2 s when using SDS. Optimal concentrations of lysis and enzyme substrate reagents were determined using microtitre plate and chip-based procedures. Fluorescence peaks, due to the enzymatic product fluorescein mono-β-D-galactopyranoside (FMG), were detected downstream of the mixing and cell lysis point for the reaction of β-Gal with 200 μM of the fluorogenic substrate fluorescein-di-β-D-galactopyranoside (FDG). FMG fluorescence was observed from cells preincubated with FDG off-chip then subsequently lysed on-chip with SDS. Unincubated cells were mixed on-chip with both FDG and Triton X-100, each individual cell generating FMG fluorescence downstream of the mixing point detected within 2 min of mixing. In contrast, viable cells incubated with FDG required 1 h or more in order to generate significant signal in a flow cytometer. |
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| Publication date | 2004-01 |
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| In | |
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| NRC publication | This is a non-NRC publication"Non-NRC publications" are publications authored by NRC employees prior to their employment by NRC. |
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| NPARC number | 12329152 |
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| Export citation | Export as RIS |
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| Report a correction | Report a correction (opens in a new tab) |
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| Record identifier | 04ca6679-9724-466f-92df-59747c08dea4 |
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| Record created | 2009-09-10 |
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| Record modified | 2020-04-17 |
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