Incucyte Imagelock (Sartorius; #BA-04857) plates were coated with 0.1 mg/mL Matrigel (Corning; #354234) and incubated at 37 °C overnight. Cells were seeded the following day (4 × 104/well for U2OS, HOS and 143B, 4.5 × 104/well for LN-229, 5 × 104/well for MDA-MB-231) and incubated for 4–6 h until cells were completely adhered and confluent. The monolayer was wounded as above, and the plate was chilled on ice for 5 min. Culture medium was removed and replaced with 50 μL of ice-cold 2 mg/mL Matrigel diluted in culture medium. To allow the Matrigel to solidify, the plate was incubated at 37 °C for 30 min before the addition of 100 μL of culture medium to each well. Phase contrast images were taken every hour using the Incucyte S3 live-cell analysis instrument. The relative wound density was measured by the Incucyte analysis software (versions 2020B and 2021C).
Imagelock
Manufactured by Sartorius
Sourced in United States
ImageLock is a laboratory equipment product designed for image-based analysis and processing. It provides core functionality for capturing, managing, and analyzing digital images within a controlled laboratory environment.
Automatically generated - may contain errors
Lab products found in correlation
Woundmaker, by Sartorius (6 mentions)
Incucyte zoom, by Sartorius (5 mentions)
Matrigel, by Corning (3 mentions)
96 pin woundmaker, by Sartorius (3 mentions)
Incucyte, by Sartorius (3 mentions)
Incucyte zoom system, by Sartorius (2 mentions)
Woundmaker device, by Sartorius (2 mentions)
Incucyte zoom kinetic imaging system, by Sartorius (1 mentions)
Incucyte system, by Sartorius (1 mentions)
Matrigel basement membrane matrix, by Corning (1 mentions)
96 well plate woundmaker tool, by Sartorius (1 mentions)
Incucyte software system, by Sartorius (1 mentions)
Mitomycin c, by Merck Group (1 mentions)
Wound maker pin tool, by Sartorius (1 mentions)
Integrated cell migration analysis module, by Sartorius (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
Matrigel matrix, by Corning (1 mentions)
Fetal calf serum (fcs), by Thermo Fisher Scientific (1 mentions)
Glutamine, by Thermo Fisher Scientific (1 mentions)
Incucyte kinetic imaging system, by Sartorius (1 mentions)
26 protocols using imagelock
1
Wound Healing Assay with Matrigel
2
Real-Time Monitoring of Receptor Binding
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HEK293S cells stably expressing human CGRP or AMY1 receptors
were seeded in growth media at 3 × 104 cells per well onto
PDL coated 96-well ImageLock plates (Sartorius) and incubated
overnight at 37°C. The following day media was replaced with warm
imaging media (Fluobrite DMEM with glutamax) and serially diluted
antibodies labelled with pH sensitive Zenon™ pHrodo™ iFL red human IgG
labeling reagent (Thermo) were tested in duplicate according to
manufacturer’s instructions. Plates were immediately placed in the
Incucyte (Sartorius) at 37°C, 5% CO2 and fluorescence
images were captured for three fields of view per well every 15 min
for 11 h.
were seeded in growth media at 3 × 104 cells per well onto
PDL coated 96-well ImageLock plates (Sartorius) and incubated
overnight at 37°C. The following day media was replaced with warm
imaging media (Fluobrite DMEM with glutamax) and serially diluted
antibodies labelled with pH sensitive Zenon™ pHrodo™ iFL red human IgG
labeling reagent (Thermo) were tested in duplicate according to
manufacturer’s instructions. Plates were immediately placed in the
Incucyte (Sartorius) at 37°C, 5% CO2 and fluorescence
images were captured for three fields of view per well every 15 min
for 11 h.
3
Scratch Wound Migration Assay
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The effect of treatment on cell migration was assessed in a scratch−wound assay using the IncuCyte® Zoom Kinetic Imaging System (Essen BioScience, USA). UW-CSCC1 and UW-CSCC2 were seeded (n = 10) onto collagen 1−coated 96−well ImageLock plates (Essen BioScience, USA) and grown to near confluency in their respective growth media. To prevent proliferation and isolate migratory effects, cells were serum−starved prior to investigation of their motility. This was achieved by replacing media in the wells with a 1% FCS analogue of the relevant growth factor−free culture media. After 24 h incubation in this low serum containing media, the cells were scratched according to manufacturer’s instructions using the 96−pin Woundmaker™ (Essen BioScience, USA). The cells were subsequently washed with serum−free media, then incubated in low serum media containing at 37 °C, 5% CO2, and imaged over 48 h at ×10 objective to track cell motility and wound width. IncuCyte™ ZOOM software (Essent BioScience, USA) was used to interpret wound width reduction over time. Output was analysed using GraphPad Prism (GraphPad Software, USA), applying a one−way ANOVA with Tukey’s multiple−comparison post−test.
4
Assessing Cell Migration and Invasion
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Migration and invasion of MM cells was assessed as described before [24 (link)]. Briefly, MSTO-wt and MSTO-shCALB2-IPTG (induced) cells were grown to confluence in 96-well ImageLock plates (Essen Bioscience) pre-coated with a thin layer of 0.1 mg/mL Matrigel Basement Membrane Matrix (Corning, Oneonta, NY, USA, Cat. No.354234). A scratch of about 1 mm was created using the 96 well-plate woundmaker tool (Essen Bioscience) as described by the manufacturer. In the case of the invasion assay, 50 µL of Matrigel (0.6 mg/mL) was added to each well after the scratch. After 30 min incubation, 100 µL of medium was added on top and plates were scanned at a 2 h-frequency using the Incucyte™ system (Essen Bioscience). Images were evaluated with the IncuCyte™ software system (version 2011A, Essen Bioscience).
5
Scratch Assay for GB Cell Migration
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U-87MG and T98G human GB cells (7.5 × 104 cells per well) were plated onto a 96-well plate (ImageLock, Essen BioScience, Hertfordshire, UK) in DMEM medium. A scratch was performed using a WoundMaker, followed by two washes with PBS. Cells were then treated with either 0.5 μM SELPi or with DMSO vehicle at corresponding percent (v/v). Wound density percent was monitored and quantified by Incucyte ZOOM (Essen BioScience). Each treatment was assayed in triplicates, and the experiment was repeated in three independent times.
6
Proliferation, Invasion, and Migration Assays
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For proliferation analysis, cells were seeded at low densities (2,500 to 5,000 cells per well). Cells were incubated using an IncuCyte ZOOM® (Essen Bioscience). Cell growth based on percent confluence was determined from phase contrast images. For proliferation assays based on automated counting of fluorescent nuclei, cells were first infected with the NucLight Red reagent (4476, Essen Bioscience) and selected using 1 μg per ml of puromycin to obtain a stable nuclear red fluorescent label. Invasion assays were performed by seeding cells at a density of 30,000 cells/well on a thin coating of Matrigel (0.1 mg/ml, standard formulation, Corning) in 96 well ImageLock microplates (4379, Essen BioScience). Cells were then allowed to grow confluent. Prior to scratching, cell proliferation was inhibited by 30 minutes exposure to 10 μg/ml mitomycin C (M4287, Sigma-Aldrich). Monolayers were scratched using the WoundMaker™ pin tool (4493, Essen BioScience). Medium was changed and cells were covered in 2 mg/ml Matrigel (standard formulation, Corning) diluted in complete medium. Relative wound density was calculated using the IncuCyte™ Scratch Wound Cell Migration Software Module (9600–0012).
7
Monitoring Macrophage Migration Dynamics
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Bone marrow-derived macrophages (BMDM; 106) were seeded onto wells of 96-well plates (ImageLock, Essen Bioscience Inc., Ann Arbor, MI, USA) and cultured in 30% L cell conditioned medium/IMDM for 7 days. Cells were then washed and cultured in fresh IMDM containing 2 μM KD025 or DMSO and analysed at the indicated times via the IncuCyte ZOOM System (Essen Bioscience) and the 2016A Live Cell Imaging sofware after application of a scratch.
8
Scratch Assay for Cell Migration
9
Automated Cell Proliferation and Migration Assay
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Cell proliferation assays were performed by seeding cells at a density of 2.25 x 104 cells per well of a 12-well plate in full growth media. All cultures were performed in quadruplicates (n=3). Scratch wound healing migration assay was performed by growing cells to confluency in 96-well plates (ImageLock; Essen Bioscience, Ann Arbor, MI) in a standard cell culture incubator. The 96-pin wound-making tool (WoundMaker; Essen Bioscience) was used to create a precise and reproducible wound in each well simultaneously. Cell proliferation and migration were monitored by Incucyte Zoom (Incucyte, Essen Bioscience). This allows an automated and non-invasive method of monitoring live cells in culture.
10
In Vitro Wound Closure Assay
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Confluent cells were serum starved for 4 hr prior to wound generation. Wounds were created in confluent monolayers of cells in a 24-well ImageLock (Essen Bioscience) plate using a WoundMaker™ (Essen Bioscience). Cells were washed twice to remove debris and finally placed in DMEM supplemented with 0.5% FBS. An IncuCyteTM was used to take 3 images per well, every 2 hr for a 12 hr period. Closure of the wound area was assessed by the relative wound density (%) automatic analysis metric of the IncuCyteTM, which calculates cell density in the wound area expressed relative to the cell density outside of the wound area over time.
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