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Four well culture slides

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Four-well culture slides provide a multi-well platform for cell culture and analysis. Each slide contains four individual wells to enable parallel experiments or observations.

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Lab products found in correlation

Axioscop 2 plus fluorescence microscope, by Zeiss (2 mentions) Bovine serum albumin (bsa), by Merck Group (2 mentions) Abcg2, by Abcam (1 mentions) Axiovert 200m, by Zeiss (1 mentions) C myc, by Abcam (1 mentions) Cd133, by Proteintech (1 mentions) A32727, by Thermo Fisher Scientific (1 mentions) Vimentin, by Abcam (1 mentions) Axioscop 2 plus, by Zeiss (1 mentions) Eclipse ti, by Nikon (1 mentions) Ab97050, by Abcam (1 mentions) Ab5694, by Abcam (1 mentions) E cadherin, by Sony (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions) Anti c jun antibody, by Santa Cruz Biotechnology (1 mentions) Vectashield mounting medium, by Vector Laboratories (1 mentions) 510 meta laser scanning confocal microscope, by Zeiss (1 mentions) Alexa fluor 647 goat anti rabbit, by Thermo Fisher Scientific (1 mentions) Dapi nuclear stain, by Thermo Fisher Scientific (1 mentions) Alexa fluor 568 goat anti mouse, by Thermo Fisher Scientific (1 mentions)

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Culture slides (24 citations) Four‐well glass culture slides (2 citations)

6 protocols using four well culture slides

1

Immunofluorescence Staining for Vimentin and Ki-67

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Cells (1 × 104) growing in four-well culture slides (BD Falcon, Bedford, MA) were fixed with ice cold methanol. For permeabilization, 0.1% Triton X-100 was added to cells for 10 min. To block nonspecific antibody binding, cells were incubated in 1% BSA (Sigma-Aldrich), 0.3 M glycine in PBST buffer (PBS with 0.1% Tween 20) for 30 min at RT. Next, cells were incubated with anti-vimentin and anti-Ki-67 antibodies for 40 min at RT. For visualization FITC-conjugated and Alexa Fluor® 594-conjugated secondary antibodies were used for 1 h at RT. Stained cells were visualized using an Axioscop 2 PLUS fluorescence microscope (Carl Zeiss, GmbH).
Nushtaeva A.A., Karpushina A.A., Ermakov M.S., Gulyaeva L.F., Gerasimov A.V., Sidorov S.V., Gayner T.A., Yunusova A.Y., Tkachenko A.V., Richter V.A, & Koval O.A. (2019). Establishment of primary human breast cancer cell lines using “pulsed hypoxia” method and development of metastatic tumor model in immunodeficient mice. Cancer Cell International, 19, 46.
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2

Visualizing Autophagy via LC3 Staining

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Cells (1×104) were cultured in four-well culture slides (BD Falcon, Bedford, MA) and treated with RL2 (200 μg/mL) for 8 hours. Cells were fixed with 4% paraformaldehyde and incubated with primary antibody against LC3 (1 μg/mL) for 20 min, followed by FITC-conjugated secondary antibody (1∶1000). Primary and secondary antibodies were diluted in blocking solution (PBS supplemented with 0.05% Tween 20 and 1% gelatin). Stained cells were visualized under an Axioscop 2 PLUS fluorescence microscope (Carl Zeiss, GmbH).
Koval O.A., Tkachenko A.V., Fomin A.S., Semenov D.V., Nushtaeva A.A., Kuligina E.V., Zavjalov E.L, & Richter V.A. (2014). Lactaptin Induces p53-Independent Cell Death Associated with Features of Apoptosis and Autophagy and Delays Growth of Breast Cancer Cells in Mouse Xenografts. PLoS ONE, 9(4), e93921.
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3

Immunofluorescent Analysis of Stem Cell Markers

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Cells growing in four-well culture slides (BD Falcon, Bedford, MA) were fixed in 4% paraformaldehyde for 10 min. For permeabilization, 0.1% Triton X-100 was added to the cells for 10 min, then, they were incubated in 5% goat serum in PBS for 30 min at RT to block nonspecific antibody binding. Next, the cells were incubated with Sox-2 (Abcam, Cambridge, MA), Oct-4 (Abcam, Cambridge, MA), c-Myc (Abcam, Cambridge, MA), CD133 (Proteintech, USA), and ABCG2 (Abcam, Cambridge, MA) primary antibodies overnight at 4°C. Secondary antibody staining was performed with either IgG/TRITC goat antirabbit or IgG/TRITC goat antimouse antibody at a 1 : 300 dilution for 2 h at room temperature. Images were captured under a fluorescence microscope (Zeiss Axiovert 200M).
Fan L., Wang X., Cheng C., Wang S., Li X., Cui J., Zhang B, & Shi L. (2023). Inhibitory Effect and Mechanism of Ursolic Acid on Cisplatin-Induced Resistance and Stemness in Human Lung Cancer A549 Cells. Evidence-based Complementary and Alternative Medicine : eCAM, 2023, 1307323.
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4

Immunofluorescence Staining of Cell Markers

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Cells (1  ×  104) growing in four-well culture slides (BD Falcon, Bedford, MA) were washed with PBS and 10% neutral-buffered formalin. To block nonspecific antibody binding, cells were incubated in 1% BSA (Sigma-Aldrich, Burlington, MA, USA) and 0.3 M glycine in PBST buffer (PBS with 0.1% Tween 20) for 30 min at RT. Next, cells were incubated with the VIMENTIN (#ab8069, Abcam, Cambridge, UK) and E-CADHERIN (#2220530, Sony Biotechnology Inc., San Jose, CA, USA), S100A4 (#MA5-31333, Invitrogen, Waltham, MA, USA) and αSMA (#ab5694, Abcam, Cambridge, UK) antibodies for 60 min at RT. For visualization, FITC-conjugated (#ab97050, Abcam, Cambridge, UK) or Alexa Fluor 555-conjugated (#A32727, Invitrogen, Waltham, MA, USA) secondary antibodies were used for 1 h at RT. Stained cells were visualized using an Axioscop 2 PLUS fluorescence microscope (Carl Zeiss, GmbH, Jena, Germany) or inverted microscope (Eclipse Ti, Nikon, Tokio, Japan).
Nushtaeva A., Ermakov M., Abdurakhmanova M., Troitskaya O., Belovezhets T., Varlamov M., Gayner T., Richter V, & Koval O. (2023). “Pulsed Hypoxia” Gradually Reprograms Breast Cancer Fibroblasts into Pro-Tumorigenic Cells via Mesenchymal–Epithelial Transition. International Journal of Molecular Sciences, 24(3), 2494.
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5

Immunofluorescence Staining of Cultured Cells

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Cells (5 × 104 each well) were seeded on four-well culture slides (Becton Dickinson Labware, Franklin Lakes, NJ, USA). After 24 h, slides were washed in PBS, fixed with 4% paraformaldehyde for 30 min and non-specific binding was blocked using 1% BSA/PBS. Slides were then incubated with an anti-c-Jun antibody (1:40; Santa Cruz Biotechnology, Heidelberg, Germany) or anti-TUB1 antibody (1:500; Sigma-Aldrich, Steinheim, Germany), washed three times with PBS, and incubated with the AlexaFlour-anti-rabbit or AlexaFlour-anti-mouse antibody (1:150; Invitrogen, Groningen, The Netherlands). Afterwards, they were washed again and finally sealed with VectaShield mounting medium (Vector Laboratories, Burlingame, CA, USA) including 1 mg/mL DAPI (Sigma-Aldrich, St. Louis, MO, USA). Images were collected by fluorescence microscopy or confocal microscopy.
Kappelmann-Fenzl M., Kuphal S., Krupar R., Schadendorf D., Umansky V., Vardimon L., Hellerbrand C, & Bosserhoff A.K. (2019). Complex Formation with Monomeric α-Tubulin and Importin 13 Fosters c-Jun Protein Stability and Is Required for c-Jun’s Nuclear Translocation and Activity. Cancers, 11(11), 1806.
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6

Subcellular Localization of Inflammasome Proteins

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Cells were grown to near confluency in four well culture slides (Becton Dickinson, Franklin Lakes, NJ) and exposed to either asbestos or CNT for 24 hours with or without pre-treatment of cyto D. Cells were fixed in 4% paraformaldehyde (PFA) for 10 minutes followed by permeabilization with 0.1% Triton-X for 10 minutes at room temperature (RT). Auto fluorescence was blocked with 0.3M glycine and non-specific antibody binding was blocked with 20% goat serum in 1x PBS for 1 hour. Cells were incubated with either anti-rabbit ASC or NLRP3 (Novus Biologicals, Littleton, CO) combined with either mitochondrial marker anti-mouse PRDX3 (AB Nova, Walnut, CA) or ER marker anti-mouse calnexin (Novus Biologicals) diluted 1:50 in 1% BSA/PBS overnight at 4°C in a humidified chamber. The following day, a mixture of secondary antibodies Alexa Fluor® 647 goat anti-rabbit and Alexa Fluor® 568 goat anti-mouse (Life Technologies, Grand Island, NY ) diluted 1:400 in 1xPBS were applied to cells for 1 hour at RT. The nuclear stain DAPI (Life Technologies) was diluted 1:200 and added to cells for 20 minutes at RT. Slides were then mounted and coverslipped with Aqua-Poly/Mount (Polysciences Inc., Warrington, PA) and stored at 4°C. Confocal images were acquired using a Zeiss 510 META laser scanning confocal microscope (Carl Zeiss, Thornwood, NY).
MacPherson M., Westbom C., Kogan H, & Shukla A. (2016). Actin polymerization plays a significant role in asbestos-induced inflammasome activation in mesothelial cells in vitro. Histochemistry and cell biology, 147(5), 595-604.
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