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and quantitative results (e.g., determination of minimum inhibi-
tory concentration) [ 1 , 13 ], although they may not accurately pre-
dict the results of many clinical samples [ 14 ]. Additionally, new
emerging mechanisms of bacterial resistance require continuous
revision of the adequacy of each AST method [ 15 , 16 ].
We developed an automated digital, time-lapse, bright field
imaging system (oCelloScope, BioSense Solutions ApS, Farum,
Denmark) for microbiological research allowing rapid AST (Fig. 1 ).
It allows fast monitoring of up to 96 bacteria–antibiotic combina-
tions in a time as quick as 2 min 19 s when a single image per well
is acquired. The oCelloScope supports several types of samples and
containers including microscope slides and microtiter plates up to
96 wells. It is well suited for liquid samples such as bacterial cul-
tures and clinical isolates such as urine [ 17 ] and blood [ 18 ] sam-
ples. Moreover, the oCelloScope also showed promising preliminary
results for solid cultures of a single bacterial colony growing on a
semitransparent medium. The miniaturized imaging system com-
prises a digital camera, an illumination unit, and a lens where the
optical axis is tilted 6.25° relative to the horizontal plane of the
stage (Fig. 2 ). Such tilt allows scanning of volumes (Fig. 2 ) and
formation of an image Z-stack containing the best-focus image, as
well as the adjacent out-of-focus images (which contain progres-
sively more out-of-focus bacteria, the further the distance from the
best-focus position). When all the bacteria are sedimented in a
microwell, they are all in focus in the best-focus image (Fig. 2 ). An
optical resolution of 1.3 μm is achieved through a proprietary lens
system and a monochrome 5 megapixel complementary metal
oxide semiconductor (CMOS) camera chip (5.6 mm length and
4.2 mm height) with a focus depth of ~10 μm and an optical mag-
nification factor of 4. The dimensions of the oCelloScope
(45 × 26 × 25 cm) allow portability and operation in standard
laboratory incubators for thorough control of temperature (20–
40 °C) and humidity (20–93%). The oCelloScope comprises
UniExplorer software (BioSense Solutions ApS) for both setting
the experiment and data analysis. The software communicates with
the instrument via an Ethernet connection, which can favorably
reduce the time required in the laboratory when working with haz-
ardous microorganisms. The UniExplorer software generates time-
lapse videos of the best-focus images acquired over time and
performs image-based analysis for online and offline quantification
of several morphological parameters [ 18 – 22 ] and microbial growth
and inhibition over time [ 23 , 24 ]. The output data can also be
exported to Microsoft Excel or in the CSV format for further
processing.
AST using the oCelloScope showed a statistically significant
antibiotic effect within 6 min for Escherichia coli monocultures and
within 30 min in complex samples from pigs with catheter-
associated urinary tract infections [ 19 ]. Additional investigationsChiara Canali et al.