The Gel Microdrop (GMD) Growth Assay is a new, improved method for evaluating the effect of antibiotics, cytotoxic agents and growth factors on cell growth. This unique, single cell assay combines microencapsulation technology with flow cytometry. In the GMD Growth Assay, single cells are encapsulated in small agarose GMDs (20-50 µm diameter) which function using miniaturized Petri dishes. Clonogenic growth or growth inhibition can be quantified at the individual cell level. Encapsulation thus permits rapid flow cytometric analysis of microcolonies forming within the GMDs which is not possible with unencapsulated cells. Drug resistant sub-populations can be rapidly detected, reducing the time needed for analyzing clinical samples. The GMD Growth Assay has been used to perform fungal susceptibility testing in 6 hr compared to 48 hr using conventional broth dilution methods. Assay time for mycobacterial susceptibility testing has been shortened to 10 days compared to 32 days for conventional culture methods. Infectious samples can be analyzed by fixing cells in GMDs before analysis by flow cytometry. An additional advantage of the GMD Growth Assay is that using FACS, cells of interest can be isolated and recovered for cloning or gene analysis, automating drug discovery applications.
This schematic representation depicts the growth of cells in GMDs (top row) and the corresponding fluorescence profiles (bottom row). Using the CellSys 100™ Microdrop Maker, single cells are encapsulated in CelGel™ Encapsulation Matrix, a unique, agarose-based material, incubated with antibiotics, cytotoxic agents or growth factors, and stained with fluorescent markers. The first panel shows GMDs occupied with individual cells. The second and third panels show microcolony formation within GMDs occupied with resistant subpopulations. Sensitive cells will not grow in the presence of an inhibitory concentration of the compound of interest. Microcolony formation is accompanied by a proportional increase in fluorescence intensity. If desired, microcolony-containing GMDs can be isolated by FACS and colony forming units recovered for further analysis.
General GMD Growth Assay Protocol
Encapsulate cells in CelGel™ Encapsulation Matrix.
Culture in the presence of compound of interest.
Stain with fluorescent marker.
Measure colony growth using flow cytometry or select cells using FACS.
GMD versus Conventional Susceptibility Assays
| Cell Type | GMD Assay | Conventional Assay |
| Mycobacteria | 10 days | 32 days (a) |
| Fungi | 4-6 hours | 24-48 hours (b) |
(a) Massachusetts State Laboratory Procedure
(primary culture, AccuProbe® identification, BACTEC™
analysis)
(b) NCCLS Broth Dilution Assay
The GMD Growth Assay decreases the time required for susceptibility testing and reduces the overall time for diagnosis and drug susceptibility determination.
| Growth Assay Benefits | Growth Assay Applications |
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Critical Factors for the GMD Growth Assay
You will need a strong project champion to develop support for this unique system in your organization.
Please note, since both leases require review by legal counsel, start this process early in order to ensure rapid delivery of your GMD system.
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© 1999, One Cell Systems, Inc.
