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Holographic time-lapse of “cell budding”
PhaseHolographic Imaging
HoloMonitor® M4for careful discoveries
“One of the major challenges of pharmaceutical
sciences is identification of dormant variants of
drug-targeted cells that are highly tolerant to
treatment – persister cells. HoloMonitorM4 is
the ideal new tool that help us to identify and
study these rare cell populations”
Ed Luther, Northeastern University of Boston
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HoloMonitor M4 captures label-free holographic images (left),
making it independent of cell staining or labeling. Holographic
time-lapse cytometry allows adherent cells to be continuously
monitored and quantified before and after treatment, minute by
minute without affecting or disturbing them.
The M4 uses a low intensity, single wavelength, laser. As cells
are imaged unstained, they absorb no energy which can cause
phototoxicity. Adherent cells thus remain undis-turbed in their
usual cell culture vessels and medium during long-term
observations.
The incubator tolerant M4 is designed to withstand the
tem-perature and humidity inside an incubator, allowing the M4 to
monitor cell cultures in their optimal environment.
Unlike conventional microscopy, the M4 has the ability to
measure optical cell volume. By measuring cell volume and
morphology, individual cell health and state is continuously
monitored. Additionally, the cytometric software allows each cell
to be tracked both spatially and morphologically over time.
Unstained cells, imaged with phase contrast microscopy, are
notoriously difficult for a computer to automatically identify.
Images created by the M4 display the optical thickness of cells.
This dramatically simplifies cell identification. Cell volume
changes in a well defined way over time. At an unexpected change in
cell volume, e.g. caused by overlapping cells, the user is notified
to verify the automatic cell identification.
Time-lapse imaging is exceptionally straight forward. The M4
records a sequence of holograms. From this sequence, individ-ual
image frames are created, auto-focused and compiled into a
time-lapse movie. This process makes the M4 insensitive to focus
drift and allow images to be refocused after recording.
HoloMonitorquantitative imaging
Label-freetime-lapse cytometry
A cell cluster. In holographic images, color variation represent
variations in optical thickness.
“Time-lapse movies taken with Holo-
Monitor gives a totally new dimension
to our work and a new insight into drug
effects never monitored before”
Prof. Stina Oredsson, Lund University
Object beam
The hologram is recorded by an image senor
The image is created by a computer from the recorded
hologram
Laser diode
Sample
Microscope objective
The hologram is created by combining the object and reference
beam
Reference beam
Holographic microscopy
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Cell division can be analyzed step by step using the M4. At
first the typical contraction of the cell is seen, as it organizes
its chromatin. This is followed by an elongation of the cell with
two separate chromatin bars. Thereafter the cell sepa-rates into
the two daughter cells. Finally the daughter cells spread out to
their flat shape with asymmetrical protrusions.
Cell differentiation is iden-tified by quantifying
mor-phological parameters such as optical cell thickness and cell
area. Lipids are optical-ly dense. When they form, they emerge as
bright spots. In the right image, the lipid droplets have been
digitally colored yellow.
Record time-lapse
Cytometricsoftware
Cell viability is a measure of the health of a cell culture.
Besides cell number and confluence the HoloMonitor M4 quantifies a
multitude of cell parameters like area, optical volume, thickness
and irregularity. This allows cell viability to be assessed and
monitored directly in the cell culture vessel.
Cell motility has until now has been complicated to analyze.
Time-lapse sequences recorded by HoloMonitor M4 are easi-ly
analyzed for both random and migratory cell movement.
3T3LI cells differentiated into adipocytes
Analysisapplications
“HoloMonitor accele rates our research,
allowing us to perform projects we could
only dream about in the past”
Prof. Yunping Dai, China Agricultural University
Cell division
Cell growth
Mitosis
Post-mitosis cell expansion
Pre-mitosis cell contraction
TimeOpti
cal v
olum
eO
ptica
l thi
ckne
ss
Cell division
Mitosis
www.phiab.sefor additional information
View cells in 3D
Automatically identify cells
Automatically track cells
Analyze cells and export results
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For research use only. Not for use in diagnostic procedures.
HoloMonitor is a trademark of Phase Holographic Imaging PHI
ABScheelevägen 22, 223 63 Lund, Sweden | +46 46 38 60 80 |
[email protected] | www.phiab.se© 2014 Phase Holographic Imaging PHI AB
| All rights reserved
Sample stage Fixed or optional XY-stageLight source External
laser unit, 635 nmObjective 20×Lateral resolution 1 µmField of view
0.25 mm2
Working distance 0.5 – 2 mmDigital focusing range 1.5 mmMaximum
image rate 1 image/sReconstructed image size 1024 × 1024
pixelXY-stage travel range 70 × 50 mmDimensions (L × W × H) 250 ×
160 × 180 mmWeight 2.5 kg
Technical specification
Cells Mono-layer of adherent eukaryotic cellsCell culture
vessels T25, 6-well plate, IBIDI µ-slide, microscopy slide and 30
mm Petri dishComputer Windows 7/8 64-bit and 8GB RAMIncubator
Access port for cablingOperating temperature 10 – 40° COperating
humidity Max 95%
Requirements
Low voltage directive 2006/95/EC, Electromagnetic compatibility
2004/108/EC EN61010-1:2001, EN60825-1:2007
Norms and standards met
