| DOI | Trouver le DOI : https://doi.org/10.1109/JPROC.2003.820551 |
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| Auteur | Rechercher : Ocvirk, Gregor; Rechercher : Salimi-Moosavi, Hossein; Rechercher : Szarka, Rod J.; Rechercher : Arriaga, Edgar A.; Rechercher : Andersson, Per E.; Rechercher : Smith, Richard; Rechercher : Dovichi, Norman J.; Rechercher : Harrison, D. Jed |
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| Format | Texte, Article |
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| Sujet | microfluidics; galactosidase; lab on a chip; single-cell analysis |
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| Résumé | 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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| Date de publication | 2004-01 |
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| Dans | |
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| Publication du CNRC | Cette publication n’est pas du CNRCCe sont des publications qui ont été rédigées par un auteur du CNRC, mais avant que celui-ci soit employé du CNRC. |
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| Numéro NPARC | 12329152 |
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| Exporter la notice | Exporter en format RIS |
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| Signaler une correction | Signaler une correction (s'ouvre dans un nouvel onglet) |
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| Identificateur de l’enregistrement | 04ca6679-9724-466f-92df-59747c08dea4 |
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| Enregistrement créé | 2009-09-10 |
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| Enregistrement modifié | 2020-04-17 |
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