THP-1 cells (human acute monocytic leukemia cell line) were maintained in RPMI supplemented with 10% FBS (EuroClone), 50 μM 2-mercaptoethanol, 20 mM Hepes, 2 mM glutamine, at 37 °C in 5% CO2. For Leishmania infections, THP-1 cells were plated at 5 × 105 cells/mL in 16-chamber Lab-Tek culture slides (Nunc) and treated with 0.1 μM phorbol myristate acetate (PMA, Sigma) for 48 h to achieve differentiation into macrophages. Cells were washed and infected with metacyclic L. infantum promastigotes at a macrophage/promastigote ratio of 1/10 for 24 h. Cell monolayers were then washed and incubated in the presence of test compounds for 72 h. Slides were fixed with methanol and stained with Giemsa. The percentage of infected macrophages in treated and non-treated cells was determined by light microscopy.
Lab tek culture slides
Manufactured by Thermo Fisher Scientific
Lab-Tek culture slides are a type of laboratory equipment designed for cell culture applications. They provide a controlled environment for the growth and observation of cells in vitro. The slides feature a chamber system that allows for the introduction and containment of cell cultures, media, and reagents. The product is intended to facilitate various cell-based experiments and analyses.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Euroclone (2 mentions)
Phorbol myristate acetate (pma), by Merck Group (2 mentions)
Rpmi 1640, by Euroclone (1 mentions)
Horse serum, by American Type Culture Collection (1 mentions)
Axio scope a1 microscope, by Zeiss (1 mentions)
Alexa fluor 488 goat anti mouse igg antibody, by Thermo Fisher Scientific (1 mentions)
J61899, by Thermo Fisher Scientific (1 mentions)
P7949, by Merck Group (1 mentions)
Dapi anti fade mounting medium, by Vector Laboratories (1 mentions)
H 1500, by Vector Laboratories (1 mentions)
Axiocam mrm camera, by Zeiss (1 mentions)
Phosphate buffered saline (pbs), by Thermo Fisher Scientific (1 mentions)
4 protocols using lab tek culture slides
1
Leishmania infantum Infection Assay in THP-1 Macrophages
2
Leishmania Infection of Differentiated THP-1 Macrophages
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
THP-1 cells (human acute monocytic leukemia cell line) were maintained in RPMI 1640 supplemented with 10% FBS (EuroClone), 50 μM 2-mercaptoethanol, 20 mM Hepes, 2 mM glutamine, at 37 °C in 5% CO2. For Leishmania infections, THP-1 cells were plated at 5 × 104 cells/well in 16-chamber Lab-Tek culture slides (Nunc) and treated with 0.1 μM phorbol myristate acetate (PMA, Sigma) for 48 h to achieve differentiation into macrophages. Cells were washed and infected with Leishmania spp. promastigotes at a macrophage:promastigote ratio of 1:10 for 24 h. The ability of the parasites to infect macrophages was examined after 4 h of promastigotes incubation in: (i) standard conditions (complete Schneider’s Drosophila Medium, 23 °C), (ii) cell culture conditions (cell medium, 37 °C, 5% CO2), (iii) co-incubation with HMEC-1 (1:10, cell:promastigote ratio) in cell culture conditions. After incubation, slides were fixed with methanol and stained with Giemsa. The percentage of infected macrophages was determined by light microscopy.
3
Immunofluorescence Imaging of Transfected Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
HEK-293 T transfected cells were cultured on 8 wells Lab-Tek culture slides (Nunc, C7182). 15 h or 48 h after transfection, cells were washed once with 200 µL of Phosphate-Buffered Saline 1× (prepared from PBS 10× pH 7.4 Gibco, 70,011–036) and fixed in 4% paraformaldehyde solution (Alfa Aesar, J61899) for 15 min. Cells were further washed three times with 200 µL of PBS 1× and permeabilized with 200 µL of PBS 1× supplemented with 0.2% Tween 20 (Sigma, P7949) for 15 min. After 30 min of incubation in 200 µL of blocking solution made with PBS 1×, 0.2% Tween 20 and 2.5% horse serum (ATCC, 30–2040), cells were incubated with 50 µg/mL of primary antibodies GN_mAb_Env01 diluted in blocking solution for 1 h. After 3 washes with 200 µL of PBS 1×, cells were incubated for 1 h with 4 µg/mL Alexa Fluor 488 goat anti-mouse IgG antibody (ThermoFisher, A11029) diluted in blocking solution. After 3 washes with 200 µL of PBS 1×, cells were counterstained with DAPI/anti-fade mounting medium (Vectashield, H-1500). Microscopy was performed with a Zeiss AXIO Scope A.1 microscope, equipped with Zeiss AxioCam MRm camera. Composite images were created using ImageJ software.
4
In Vitro Antileishmanial Assay Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For Leishmania spp. infections, 100 μL of THP-1 cells (5 × 105 cells/mL) or canine macrophages (2 × 106 cells/mL) were plated in 16-chamber Lab-Tek culture slides (Nunc) and further treated with 0.1 µM of PMA for 72 h to achieve differentiation into macrophages [21 (link)]. Cells were then washed with PBS and infected with stationary phase L. infantum promastigotes at a macrophage:promastigote ratio of 1:10. After 24 h, cell monolayers were extensively washed with PBS to remove non-internalized promastigotes and observed under an inverted light microscope to evaluate both the cellular size and the cellular morphology. Infected cells were then incubated in the presence of test compounds for 72 h. Infected and/or treated cells were then fixed with 100% methanol and stained with Giemsa at room temperature. The percentage of infected macrophages in treated and non-treated wells was determined by counting 200–300 macrophages/well under a light microscope. About 10 random microscopic fields/well were observed. The number of infected macrophages in the untreated control samples was considered 100% for calculating the percentage of infection in drug treated samples. IC50 values were then calculated. Data are the mean ± standard deviation of individual experiments performed in triplicate.
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!