LABORATORY FOR CELL AND TISSUE CULTURES

Laboratory for Cell and Tissue Cultures (LCTC) is dedicated for basic and applicative research and higher education in cell culturing and tissue engineering. We develop various 2D and 3D in vitro models with emphasis on the development of biomimetic in vitro models. We use the in vitro models developed from normal or pathological samples that mimic healthy tissue or diseases to test potential new therapeutic agents. We perform research with good laboratory practice in clean rooms. The LCTC is also accredited for the GSO2 use.

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The main objectives of the LCTC are (1) to provide high-quality services and technical expertise in cell and tissue culturing, (2) to run basic and practical courses, (3) to support interdisciplinary research in the development and application of advanced methods in tissue engineering and preclinical studies, and (4) to collaborate with industry in the development and testing of new therapeutics for medical purposes.

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The LCTC is part of the Centre for Electron Microscopy and therefore also offers comprehensive analysis of samples with various electron and light microscopic analyses.

 

Expertise of Laboratory for Cell And Tissue Cultures - morphological, ultrastructural, molecular, functional tests (in alphabetical order):

 

• Analysis of inflammatory factors

• Analysis of the nanoparticle effects on cellular level

• Cell transfections with different vectors (cell biological analyses)

• Cell viability tests

• Ciliary beat frequency analysis

• Correlative light and electron microscopy

• Culturing and optimization of various in vitro models of normal, benign and malignant cells (e.g. nasal cell line RPMI1640, bronchiolar cell line Calu-3, primary human nasal cells, MucilAir, EpiCorneal, oesophageal human cell line FLO-1, colorectal cell line Caco-2, urinary bladder urothelial cells, breast cells, keratinocytes, fibroblasts, macrophages, human amniotic membrane stem/stromal cells)

• ELISA

• Extracellular vesicles analysis

• Freeze-fracture immunolabeling

• Histological and histochemical analysis

• Immunolabelling of different proteins on parafin sections, cryo sections and in situ

• Lectin immunolabeling

• Liposome, multilamellar vesicle analysis

• Live-cell imaging with confocal Airy-disk microscopy

• Ex vivo models – related to urological diseases

• Microbial and antimicrobial tests

• Migration and invasion tests

• Preparation of human amniotic membrane preparations with antimicrobial and anticancer activities

• RT-qPCR of drug transporters and other genes

• Scanning electron microscopy

• Single cell sequencing

• Spectroscopy analysis

• Tissue engineering (different scaffold and cells)

• Transepithelial electrical resistance measurements

• Transmission electron microscopy

• Western blot

• Wound healing model

 

Main equipment:

• Centrifuge 5804 R (Eppendorf)

• CO2 incubator CB170 Binder with O2 regulation

• CO2 incubator Heracell (Heraeus) (2×)

• CO2 incubator Heracell Vios 160 with O2 control (Thermo Scientific)

• FemtoJet, InjecMan (Eppendorf)

• Fluorescence microscope Eclipse TE300 (Nikon)

• High resolution confocal microscope, confirmed purchase of equipment to be installed at ICB in June 2020 in the context of ELIXIR-SI ESFRI call

• Inverted microscope DM IL (Leica)

• Laminar vertical cabinet, LFV 9 (Iskra Pio)

• Live cell imaging

• Microbiological safety cabinet M12, Class II (Iskra Pio)

• Microbiological safety cabinet M9-2, M12-2, M18, Class II (Iskra Pio)

 

Notable publications with industry:

• In vitro Ciliotoxicity and Cytotoxicity Testing of Repeated Chronic Exposure to Topical Nasal Formulations for Safety Studies. Tratnjek L, Sibinovska N, Kristan K, Kreft ME. Pharmaceutics. 2021 Oct 20;13(11):1750. doi: 10.3390/pharmaceutics13111750.

• Standardization of esophageal adenocarcinoma in vitro model and its applicability for model drug testing. Tratnjek L, Sibinovska N, Kralj S, Makovec D, Kristan K, Kreft ME. Sci Rep. 2021 Mar 23;11(1):6664. doi: 10.1038/s41598-021-85530-w.

• Different Culture Conditions Affect Drug Transporter Gene Expression, Ultrastructure, and Permeability of Primary Human Nasal Epithelial Cells. Kreft ME, Tratnjek L, Lasič E, Hevir N, Rižner TL, Kristan K. Pharm Res. 2020 Aug 20;37(9):170. doi: 10.1007/s11095-020-02905-w.

• Ciliary beat frequency of in vitro human nasal epithelium measured with the simple high-speed microscopy is applicable for safety studies of nasal drug formulations. Tratnjek L, Kreft M, Kristan K, Kreft ME. Toxicol In vitro. 2020 Aug;66:104865. doi: 10.1016/j.tiv.2020.104865.

• Demonstrating suitability of the Caco-2 cell model for BCS-based biowaiver according to the recent FDA and ICH harmonised guidelines. Jarc T, Novak M, Hevir N, Rižner TL, Kreft ME, Kristan K. J Pharm Pharmacol. 2019 Aug;71(8):1231-1242. doi: 10.1111/jphp.13111.

• The characterization of the human cell line Calu-3 under different culture conditions and its use as an optimized in vitro model to investigate bronchial epithelial function. Kreft ME, Jerman UD, Lasič E, Hevir-Kene N, Rižner TL, Peternel L, Kristan K. Eur J Pharm Sci. 2015 Mar 10;69:1-9. doi:  10.1016/j.ejps.2014.12.017.

• The characterization of the human nasal epithelial cell line RPMI 2650 under different culture conditions and their optimization for an appropriate in vitro nasal model. Kreft ME, Jerman UD, Lasič E, Lanišnik Rižner T, Hevir-Kene N, Peternel L, Kristan K. Pharm Res. 2015 Feb;32(2):665-79. doi: 10.1007/s11095-014-1494-0.