All microscopic figures were obtained under a Leica DMi1 inverted phase contrast microscope (Leica Microsystems, Wetzlar, Germany), at 50× magnification.
Dmi1 inverted phase contrast microscope
Manufactured by Leica
Sourced in Germany, United States
The DMi1 is an inverted phase contrast microscope manufactured by Leica. It is designed for routine microscopy tasks, providing a stable and reliable platform for various applications. The DMi1 employs phase contrast technology to enhance the visibility of transparent specimens, making it suitable for a range of sample types.
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Lab products found in correlation
Pen strep, by Thermo Fisher Scientific (1 mentions)
Trypsin edta, by Thermo Fisher Scientific (1 mentions)
High glucose dmem, by Thermo Fisher Scientific (1 mentions)
25 cm2 flask, by Corning (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Gentamycin, by Thermo Fisher Scientific (1 mentions)
Trypsin edta, by Avantor (1 mentions)
Bioamf 3tm complete medium, by Sartorius (1 mentions)
7 protocols using dmi1 inverted phase contrast microscope
1
Inverted Phase Contrast Microscopy Imaging
2
Inverted Phase Contrast Microscopy Imaging
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All microscopic figures were obtained under a Leica DMi1 inverted phase contrast microscope (Leica Microsystems, Wetzlar, Germany), at 50× magnification.
3
HeLa Multicellular Tumor Spheroid Formation
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For the generation of HeLa multicellular tumor
spheroids (MTCS), 96-well Corning microplates with ultralow attachment
surface coating were used. Briefly, a single suspension of HeLa cells
at a density of 5 × 103 cells/well was prepared in
complete DMEM medium and dispensed into wells. The plates were covered
and transferred to incubator at 310 K with 5% CO2 atmosphere.
Within 3 days, uniform 200 μm diameter MTCS were formed from
cell suspension and were maintained under these conditions. At day
3, MTCS were incubated with tested agents (2 μM) for 6 h and
then irradiated with red light for 0.5 h. Treatments were then replaced
with fresh cell media and changed every 3 days by replacing 50% of
the media. The formation, integrity, diameter, and volume of the multicellular
tumor spheroids (MCTS) were monitored using a DMi1 inverted phase
contrast microscope (Leica Microsystems) over a span of 9 days.
spheroids (MTCS), 96-well Corning microplates with ultralow attachment
surface coating were used. Briefly, a single suspension of HeLa cells
at a density of 5 × 103 cells/well was prepared in
complete DMEM medium and dispensed into wells. The plates were covered
and transferred to incubator at 310 K with 5% CO2 atmosphere.
Within 3 days, uniform 200 μm diameter MTCS were formed from
cell suspension and were maintained under these conditions. At day
3, MTCS were incubated with tested agents (2 μM) for 6 h and
then irradiated with red light for 0.5 h. Treatments were then replaced
with fresh cell media and changed every 3 days by replacing 50% of
the media. The formation, integrity, diameter, and volume of the multicellular
tumor spheroids (MCTS) were monitored using a DMi1 inverted phase
contrast microscope (Leica Microsystems) over a span of 9 days.
4
Isolation and Preparation of hAF Cells
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The hAF cell samples were observed under DMi1 inverted phase contrast microscope (Leica Microsystems, USA). The cell samples in the 2nd passage were washed twice with sterile phosphate-buffered saline (PBS) (Amresco, Ohio, USA) and were trypsinized with 0.25% trypsin-EDTA. Subsequently, hAF cells were suspended in the basal growth medium (DMEM-high glucose with 10% FBS) and centrifuged at 2,035 g for 6 min at room temperature. After that, the supernatant was removed and the hAF cells were used in the experiments.
5
Multimodal Anti-cancer Treatment of MCTSs
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Briefly, a single suspension of A2780cis cells at a density of 5
× 103 cells/well in complete RPMI medium was dispatched
onto 96-well Corning microplates with ultralow attachment surface
coating. The plates were covered and transferred to an incubator at
310 K with a 5% CO2 atmosphere. After 3 days post-seeding,
uniform MCTSs were formed, which was confirmed using an inverted Zeiss
AXIO observer 7 microscope. Cell media of MCTSs was changed by replacing
50% with fresh cell media and allowed to grow for 3 days. On day 3,
MTCS was incubated with tested agents (10 μM) for 4 h and then
irradiated with 520 nm light for 1 h. Treatments were then carefully
removed, and fresh media was added. The integrity, radius, size, and
volume of the MCTSs were monitored using a DMi1 inverted phase contrast
microscope (Leica Microsystems) over 7 days. The radius of the tumorspheres
was measured using Fiji software, and the volume was calculated using
the following equation: V = 4/3πr3.
× 103 cells/well in complete RPMI medium was dispatched
onto 96-well Corning microplates with ultralow attachment surface
coating. The plates were covered and transferred to an incubator at
310 K with a 5% CO2 atmosphere. After 3 days post-seeding,
uniform MCTSs were formed, which was confirmed using an inverted Zeiss
AXIO observer 7 microscope. Cell media of MCTSs was changed by replacing
50% with fresh cell media and allowed to grow for 3 days. On day 3,
MTCS was incubated with tested agents (10 μM) for 4 h and then
irradiated with 520 nm light for 1 h. Treatments were then carefully
removed, and fresh media was added. The integrity, radius, size, and
volume of the MCTSs were monitored using a DMi1 inverted phase contrast
microscope (Leica Microsystems) over 7 days. The radius of the tumorspheres
was measured using Fiji software, and the volume was calculated using
the following equation: V = 4/3πr3.
6
HeLa Multicellular Tumor Spheroid Formation
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To generate HeLa multicellular tumor spheroids
(MCTS), 96-well Corning
microplates with an ultralow attachment surface coating were utilized.
The process involved preparing a single suspension of HeLa cells at
a density of 5 × 103 cells per well in complete DMEM
medium, which was then dispensed into the wells. The plates were covered
and placed in an incubator with a temperature of 37 °C and a
5% CO2 atmosphere. Within 3 days, uniform MCTS with a diameter
of 200 μm were formed from the cell suspension and maintained
under these conditions. On the first day of treatment, the MCTS were
treated withRe7 –Re9 and cisplatin
at their concentration of IC50. The media were changed
every 3 days by replacing 50% of the existing media. The formation,
integrity, diameter, and volume of the MCTS were monitored over a
span of 10 days using a DMi1 inverted phase contrast microscope (Leica
Microsystems). The volumes of the MCTS were calculated using the equation V = 4/3πr3, where “V″ represents volume and “r″ represents the radius of the MCTS measured with ImageJ software.
(MCTS), 96-well Corning
microplates with an ultralow attachment surface coating were utilized.
The process involved preparing a single suspension of HeLa cells at
a density of 5 × 103 cells per well in complete DMEM
medium, which was then dispensed into the wells. The plates were covered
and placed in an incubator with a temperature of 37 °C and a
5% CO2 atmosphere. Within 3 days, uniform MCTS with a diameter
of 200 μm were formed from the cell suspension and maintained
under these conditions. On the first day of treatment, the MCTS were
treated with
at their concentration of IC50. The media were changed
every 3 days by replacing 50% of the existing media. The formation,
integrity, diameter, and volume of the MCTS were monitored over a
span of 10 days using a DMi1 inverted phase contrast microscope (Leica
Microsystems). The volumes of the MCTS were calculated using the equation V = 4/3πr3, where “V″ represents volume and “r″ represents the radius of the MCTS measured with ImageJ software.
7
Isolation and Culture of hAF-MSCs
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The direct adherent method was used to separate hAF-MSCs [34 (link)]. In brief, hAF cells that were cultured in 25 cm2 flasks (Corning Incorporated, NY, USA) with expansion medium (BIOAMF-3TM Complete Medium) (Biological Industries, Kibbutz Beit Haemek, Israel) at 37 °C, 5% CO2 and 95% humidity were changed to culture with the basal growth medium comprised of Dulbecco's Modified Eagle Medium (DMEM)–high glucose (Gibco, USA) with a supplement of 10% fetal bovine serum (FBS) (Gibco, South America), gentamycin 40 mg/ml and Pen Strep (penicillin and streptomycin) 10,000 U/ml (Gibco, USA). The medium was changed every 3 days. After the cells reached 80% confluence, they were sub-cultured using 0.25% trypsin-EDTA (Gibco, USA). The cell samples that were collected from the 2nd passage were used in all of our experiments.
The hAF cell samples were observed under a DMi1 inverted phase contrast microscope (Leica Microsystems, USA). The cell samples that were collected from the 2nd passage were washed twice with sterile phosphate-buffered saline (PBS) (Amresco, Ohio, USA) and were trypsinized with 0.25% trypsin-EDTA. Subsequently, hAF cells were suspended in basal growth medium and centrifuged (C2 Series, Centurion Scientific Ltd, UK) at 2,035 g for 6 min at room temperature. After that, the supernatant was removed and the hAF cells were used in the experiments.
The hAF cell samples were observed under a DMi1 inverted phase contrast microscope (Leica Microsystems, USA). The cell samples that were collected from the 2nd passage were washed twice with sterile phosphate-buffered saline (PBS) (Amresco, Ohio, USA) and were trypsinized with 0.25% trypsin-EDTA. Subsequently, hAF cells were suspended in basal growth medium and centrifuged (C2 Series, Centurion Scientific Ltd, UK) at 2,035 g for 6 min at room temperature. After that, the supernatant was removed and the hAF cells were used in the experiments.
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