Samples of 3 × 105 hCMEC/D3 cells were seeded in 6-well plates containing EBM-2 medium complemented with 1.25% FBS (other supplements as above). sEVs at various concentrations (0–250 µg/mL) obtained from (a) MSC culture medium (DMEM low, Lonza) supplemented with 10% platelet lysate (sEVplatelet), (b) supernatants of MSCs cultured under regular ‘normoxic’ conditions (21% O2; sEVnormoxic), or (c) supernatants of MSCs cultured under hypoxic conditions (1% O2; sEVhypoxic) were added. Forty-eight hours later, cells were stained with 0.4% tryptan blue (Sigma-Aldrich). The number of viable cells was determined using an automatic cell counter (EVE Automatic Cell Counter; NanoEnTek, Waltham, MA, USA).
Eve automatic cell counter
Manufactured by NanoEnTek
Sourced in United States
The EVE automatic cell counter is a compact and efficient device designed for cell counting purposes. It utilizes advanced optical technology to accurately determine the number of cells in a sample. The core function of the EVE cell counter is to provide reliable and automated cell counting capabilities.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (6 mentions)
Non enzymatic cell dissociation solution, by Merck Group (2 mentions)
Glutamine, by Merck Group (2 mentions)
Antibiotic antimycotic solution, by Merck Group (2 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
Trypsin edta, by Thermo Fisher Scientific (2 mentions)
Tryptan blue, by Merck Group (1 mentions)
Lsm 710 confocal microscope, by Zeiss (1 mentions)
Apoptag fluorescein direct in situ apoptosis detection kit, by Merck Group (1 mentions)
Streptomycin, by Merck Group (1 mentions)
Phorbol myristate acetate (pma), by Merck Group (1 mentions)
Penicillin, by Merck Group (1 mentions)
Axiovert 40c microscope, by Zeiss (1 mentions)
D5796, by Merck Group (1 mentions)
Dulbecco modified eagle medium (dmem), by Cayman Chemical (1 mentions)
Cx 5461, by Cayman Chemical (1 mentions)
Countess cell counting chamber slide, by Thermo Fisher Scientific (1 mentions)
Trypan blue, by Thermo Fisher Scientific (1 mentions)
Tryple reagent, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Biowest (1 mentions)
36 protocols using eve automatic cell counter
1
Evaluating hCMEC/D3 Cell Viability
2
Serum Deprivation and Apoptosis Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
A10 cells seeded into 12-well cluster dishes (2.0 × 105 cells/well) were deprived of serum for 24 h and further incubated in the presence or absence of SBSN_HUMAN[225–237] or SBSN_HUMAN[243–259] for the indicated times. Cells were then trypsinized, and cell numbers measured using an EVE automatic cell counter (NanoEnTek Inc, Seoul, Korea)50 (link).
HAoSMCs were cultured to subconfluency and serum deprived for 9 h in the presence or absence of SBSN_HUMAN[225–237] (10–6 M) or SBSN_HUMAN[243–259] (10–6 M). The terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) assay was performed using an ApopTag Fluorescein Direct In Situ Apoptosis Detection Kit (EMD Millipore, USA)51 (link). DNA fragmentation was detected by laser scanning confocal microscopy using an LSM710 confocal microscope (Carl Zeiss, Jena, Germany).
HAoSMCs were cultured to subconfluency and serum deprived for 9 h in the presence or absence of SBSN_HUMAN[225–237] (10–6 M) or SBSN_HUMAN[243–259] (10–6 M). The terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) assay was performed using an ApopTag Fluorescein Direct In Situ Apoptosis Detection Kit (EMD Millipore, USA)51 (link). DNA fragmentation was detected by laser scanning confocal microscopy using an LSM710 confocal microscope (Carl Zeiss, Jena, Germany).
3
Differentiation and Proliferation of MO3.13 Oligodendrocytes
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The human oligodendrocytic cell line MO3.13 was purchased from Cedarlane (Burlington, ON, Canada). Cells were cultured according to the manufacturer’s protocol in DMEM (D5796, Sigma-Aldrich, St. Louis, MO, USA) supplemented with 10% FBS (Thermo Fisher Scientific, Waltham, MA, USA), 100 μg/ml streptomycin, and 100 U/mL penicillin (both from Sigma-Aldrich, St. Louis, MO, USA) in 5% CO2 at 37 °C. The medium was changed every 2–3 days, and cells were passaged when confluent. To induce differentiation, MO3.13 cells were incubated in the presence of 100 nM PMA (Sigma-Aldrich, St. Louis, MO, USA) or in the presence of 200 nM Pol I inhibitor, CX-5461 (Cayman Chemical, Ann Arbor, MI, USA) in a medium containing DMEM supplemented with 1% FBS. The medium was exchanged every second day, and cell morphology was analyzed daily. Cell morphology was examined using an Axiovert 40C microscope (Carl Zeiss, Jena, Germany) equipped with an A-Plan 10×/0.25 Ph1-objective.
Proliferation of MO3.13 cells was assessed by cell counting, using an EVE automatic cell counter (NanoEnTek, Seoul, Korea). Cells were plated onto a 12-well plate at the density of 150,000 cells per well. After 24 h, cells were treated with PMA (Sigma-Aldrich, St. Louis, MO, USA) at a final concentration of 100 nM or left untreated (control). Cells were cultured for additional 72 h, then trypsinized and counted.
Proliferation of MO3.13 cells was assessed by cell counting, using an EVE automatic cell counter (NanoEnTek, Seoul, Korea). Cells were plated onto a 12-well plate at the density of 150,000 cells per well. After 24 h, cells were treated with PMA (Sigma-Aldrich, St. Louis, MO, USA) at a final concentration of 100 nM or left untreated (control). Cells were cultured for additional 72 h, then trypsinized and counted.
4
Culturing and Characterizing MDA-MB-231 Breast Cancer Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The study was conducted using the MDA-MB-231 cell line. This is an epithelial human breast cancer cell line, established from a pleural effusion of a 51-year-old Caucasian woman with metastatic breast adenocarcinoma. The MDA-MB-231 breast cancer cell line was purchased from the American Type Culture Collection (ATCC) and authenticated by DNA profiling using short tandem repeat (STR) (GenePrint® 10 System, Promega) at Genomics Core Facility, Instituto de Investigaciones Biomédicas “Alberto Sols” CSIC-UAM. Viral, bacterial and parasitic pathogen analysis by RT-PCR/PCR was performed at Dynamimed Research Company: no genetic material was detected. Cells were maintained at 37 °C in a 5% CO2 humidified incubator (AutoFlow UN-5510, Nuaire), in Dulbecco’s modified Eagle’s medium (DMEM) (Life Technologies) supplemented with 10% heat-inactivated fetal bovine serum (FBS) (Biowest) and antibiotics (penicillin, streptomycin) (Gibco®-Invitrogen). Cells were trypsinized/passaged every 2–3 days using TrypLE reagent (ThermoFisher Scientific). Cells were automatically counted using Countess cell counting chamber slides (Invitrogen) and an Eve Automatic cell counter (NanoEntek), excluding dead cells by trypan blue (Invitrogen) staining.
5
VEGF-A Secretion Quantification
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Secreted VEGF-A concentration was collected 24 hours and 48 hours after IR and detected in filtered conditioned media using a VEGF-A DuoSet ELISA kit according to the manufacturer's guidelines (R&D Systems; DY293B-05). The absorbance was determined with a plate reader TECAN Infinite 200 PRO at 450 nm excitation and 540 nm emission and normalized to live cell count using EVE Automatic cell counter (NanoEnTek).
6
Cell Viability and Proliferation Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
A549 and Calu-1 cells were seeded into T-25-cm2 flasks and treated with 5-dAzaC or TSA as described above. After each incubation period, cells were detached with Trypsin–EDTA solution, Sigma-Aldrich Co. (St. Louis, MO), centrifuged and resuspended in 8 ml of phosphate-buffered saline (PBS). Next, cell viability and proliferation were determined by a trypan blue staining. Fifty μl of cell suspension was taken and mixed with the same volume of trypan blue. Cells were counted with an EVE™ automatic cell counter, NanoEnTek Inc. (Seoul, Korea). For cell viability, results are expressed as the percentage of viable cells relative to respective controls (100 %), and for cell proliferation, the number of counted cells per 1 ml of solution is presented. All experiments were performed in triplicate, and data represent mean ± standard deviation (SD).
7
Evaluating Cell Viability with Trypan Blue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cell viability was tested using the EVE Automatic Cell Counter (NanoEnTek Inc., Seoul, Republic of Korea) (Figure 10 ) based on the different staining of live and dead cells when using Trypan blue dye.
This test method is an efficient and economical method of counting and testing cell viability.
To express cell viability as % viability, the calculation formula was also used:
NHDF cells were seeded in 24-well plates at a cell density of 1 × 104 cells/well and maintained at 37 °C with 5% CO2 for 24 h, and then the cells were treated with EPF extract at different concentrations: E200 µg/mL, E100 µg/mL, and E50 µg/mL for 24 h. Cells were trypsinized, neutralized, and centrifuged (1000 rpm/5 min).
The obtained cell pellet was suspended in cell medium, and cell viability was determined.
This test method is an efficient and economical method of counting and testing cell viability.
To express cell viability as % viability, the calculation formula was also used:
NHDF cells were seeded in 24-well plates at a cell density of 1 × 104 cells/well and maintained at 37 °C with 5% CO2 for 24 h, and then the cells were treated with EPF extract at different concentrations: E200 µg/mL, E100 µg/mL, and E50 µg/mL for 24 h. Cells were trypsinized, neutralized, and centrifuged (1000 rpm/5 min).
The obtained cell pellet was suspended in cell medium, and cell viability was determined.
8
Keratinocyte Proliferation and Viability Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
At passage three (P3) subconfluence, six keratinocyte cultures in 75 cm2 cell culture flasks were starved (DMEM only) for 12 h and then randomised into three groups. Two cultures were subsequently given only DMEM, two were kept in DMEM with 50% AF and two received DMEM with 50% FCS. All cultures were trypsinised and pelleted 24 h later.
Cell counting was performed by staining an aliquot of the trypsinised cell suspension in Trypan Blue and quantifying total, live and dead cells in the EVE Automatic cell counter (NanoEnTek). Cell counts are given as live cells per ml and viability as % of total cells counted. Keratinocyte cultures in KSFM, DMEM and DMEM with 50% FCS (n = 6) were quantified for proliferation and viability at 24 and 48 h and presented as mean with SD (FigureS1 ).
Cell counting was performed by staining an aliquot of the trypsinised cell suspension in Trypan Blue and quantifying total, live and dead cells in the EVE Automatic cell counter (NanoEnTek). Cell counts are given as live cells per ml and viability as % of total cells counted. Keratinocyte cultures in KSFM, DMEM and DMEM with 50% FCS (n = 6) were quantified for proliferation and viability at 24 and 48 h and presented as mean with SD (Figure
9
PBMC Isolation from Healthy Human Blood
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Human blood was obtained from healthy volunteers following informed consent. The Institutional Review Board of North Dakota State University approved the study (#3735). The age of the donors is 39.4 ± 3.0 for females and 38.5 ± 3.6 for males. Blood was collected in 3.2% sodium citrate vacutainers. Blood was diluted with RPMI1640, added to 10 ml of Ficoll-Paque, and centrifuged at 300 g for 30 minutes at RT. Peripheral blood mononuclear cells (PBMC) were collected from the interphase and washed twice in PBS. Viable cell numbers were determined using Eve Automatic Cell counter (NanoEntek, Walthan, MA). Cells with viability higher than 85% were selected and viability was taken into account when the cell number was calculated. Cells (0.5x106 cells/mL) were then cultured at 37°C and 5% CO2 in RPMI 1640 media, fully supplemented with penicillin-streptomycin (each at 0.8 mM) and L-glutamine (2 mM).
10
Colony Formation Assay for PDPN Silencing
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After PDPN silencing or inhibitor treatment experiments, the TPC1 and BcPAP cells were counted using the EVE automatic cell counter (NanoEnTek, Korea). Then, 500 cells were seeded in one 100 mm Petri dish (three replicates for each experiment) and incubated in a 5% CO2 humidified environment at 37 °C for 7–12 days, depending on the cell line. Next, the cells were fixed with 10% PFA for 25 min, washed twice with water, and stained with crystal violet (0.5% w/v) for 15 min. Digital images of the dried colonies were obtained using a scanning device and visible clones were counted using ImageJ.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!