Venorgem advance kit

Manufactured by Minerva Biolabs

Sourced in Germany

The VenorGeM Advance kit is a laboratory product designed for the detection of mycoplasma contamination in cell cultures. The kit utilizes a sensitive PCR-based method to identify the presence of mycoplasma species.

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6 protocols using venorgem advance kit

Cell Line Maintenance and Authentication

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All cell lines were maintained at 37°C in 5% CO₂ and split every 2–4 days using 0.05% trypsin/EDTA (Gibco) for detachment. MG-63, U2OS, Saos2, CAL72 and HeLa wt cell lines were cultured in DMEM medium containing 1 g l⁻¹ glucose, supplemented with 10% doxycycline-free fetal bovine serum, 1× penicillin/streptomycin and 2 mM stable glutamine. HeLa ST, HeLa LT, MGBM1, SF188 cells were cultured in DMEM containing 4.5 g l⁻¹ glucose and HOS cells in EMEM, both supplemented as described above. Human umbilical vein endothelial cells (HUVECs) (Lonza) were cultured in EGM-2 medium. NEM165, NEM157, NEM168 parental lines and ATRX knockout clones were maintained in Amniopan complete medium (PAN Biotech). U2OS, HeLa wt, CAL72 and Saos2 cells were obtained from the German Collection of Microorganisms and Cell Culture (DSMZ). HOS and MG-63 were purchased from CLS Cell Lines Services. NEM, MGBM1 and SF188 cell lines are described in (27 (link)) and HeLa ST and LT cell lines were kindly provided by Jan Karlseder (Salk Institute, USA) and are described in (28 (link)). All cell lines were tested for the absence of mycoplasma with the VenorGeM Advance kit (Minerva Biolabs) and were authenticated using single nucleotide polymorphism-profiling (Multiplexion).

Frank L., Rademacher A., Mücke N., Tirier S.M., Koeleman E., Knotz C., Schumacher S., Stainczyk S.A., Westermann F., Fröhling S., Chudasama P, & Rippe K. (2022). ALT-FISH quantifies alternative lengthening of telomeres activity by imaging of single-stranded repeats. Nucleic Acids Research, 50(11), e61.

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Cell culture conditions and validation

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Cells were grown in GIBCO DMEM (Thermo Fisher Scientific) supplemented with 10% fetal calf serum (PAA), 2 mM L-glutamine, 1% penicillin/streptomycin (PAA) and 1 g/l glucose for U2OS or 4.5 g/l glucose for iMEF and 3T3 cells. Cells were cultured at 37°C and 5% CO₂. References to the descriptions and the sources of the cell lines are given in the Key Resources Table above. Cell lines were generated and initially characterized in the respective laboratories. We tested them for the absence of mycoplasma with the VenorGeM Advance kit (Minerva Biolabs) and assessed their authenticity by analyzing RNA-seq data generated with them as compared to published datasets.

Erdel F., Rademacher A., Vlijm R., Tünnermann J., Frank L., Weinmann R., Schweigert E., Yserentant K., Hummert J., Bauer C., Schumacher S., Al Alwash A., Normand C., Herten D.P., Engelhardt J, & Rippe K. (2020). Mouse Heterochromatin Adopts Digital Compaction States without Showing Hallmarks of HP1-Driven Liquid-Liquid Phase Separation. Molecular Cell, 78(2), 236-249.e7.

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Culturing Mycoplasma-Free HEK-293 Cells

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The HEK-293 cell line was purchased from ATCC and cultured in DMEM high glucose (HG), without pyruvate (Thermo Fisher Scientific) with 10% FBS (Thermo Fisher Scientific), 1% pyruvate (Thermo Fisher Scientific), 1% NEAA (Thermo Fisher Scientific), and 100 U/ml penicillin–streptomycin (Thermo Fisher Scientific) in a 37 °C and 5% CO₂ incubator. The cell line was tested to be mycoplasma-free according to Venor®GeM Advance kit (Minerva Biolabs, Berlin, Germany) at multiple times throughout the study.

Disciglio V., Forte G., Fasano C., Sanese P., Lepore Signorile M., De Marco K., Grossi V., Cariola F, & Simone C. (2021). APC Splicing Mutations Leading to In-Frame Exon 12 or Exon 13 Skipping Are Rare Events in FAP Pathogenesis and Define the Clinical Outcome. Genes, 12(3), 353.

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Cell Line Maintenance and Mycoplasma Testing

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The murine colon carcinoma CT26 cells and the fibrosarcoma cell line MCA 205 were kindly provided by Prof. Zhihai Qin. The myeloid suppressor cell line MSC 2 cell line was a kind gift from Professor Ghiringhelli (Institut National de la Santé et de la Recherche Médicale [INSERM] U866, Dijon, France). All cell lines were tested for mycoplasma contamination on a monthly basis using Venor^®GeM Advance Kit (Minerva Biolabs GmbH, Berlin, Germany) according to the manufacturer's protocols.

Wu H., Han Y., Rodriguez Sillke Y., Deng H., Siddiqui S., Treese C., Schmidt F., Friedrich M., Keye J., Wan J., Qin Y., Kühl A.A., Qin Z., Siegmund B, & Glauben R. (2019). Lipid droplet‐dependent fatty acid metabolism controls the immune suppressive phenotype of tumor‐associated macrophages. EMBO Molecular Medicine, 11(11), e10698.

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Cell Culture Maintenance and Mycoplasma Testing

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HeLa 229, HEK293T and Vero cells (all from the European Collection of Cell Culture; ECACC) were maintained in high-glucose Dulbecco's modified Eagle Medium (DMEM; Thermo Fisher Scientific) supplemented with heat-inactivated 10% (v/v) fetal bovine serum (FBS; Thermo Fisher Scientific) at 37°C in a humidified atmosphere of 5% (v/v) CO₂. Cells were checked for Mycoplasma by conventional PCR either using the Venor® GeM Advance kit (Minerva Biolabs) or as described (Uphoff and Drexler, 2011 (link)).

Almeida F., Luís M.P., Pereira I.S., Pais S.V, & Mota L.J. (2018). The Human Centrosomal Protein CCDC146 Binds Chlamydia trachomatis Inclusion Membrane Protein CT288 and Is Recruited to the Periphery of the Chlamydia-Containing Vacuole. Frontiers in Cellular and Infection Microbiology, 8, 254.

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Culturing and Transfecting HeLa and Vero Cells

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HeLa 229 and Vero cells (from the European Collection of Cell Culture; ECACC) were maintained in high-glucose Dulbecco’s modified Eagle Medium (DMEM; Thermo Fisher Scientific) supplemented with heat-inactivated 10% (v/v) fetal bovine serum (FBS; Thermo Fisher Scientific) at 37 °C in a humidified atmosphere of 5% (v/v) CO₂. Cells were checked for Mycoplasma by conventional PCR either using the Venor GeM Advance kit (Minerva Biolabs) or as described^{63 (link)}. HeLa cells were transfected by using the jetPEI reagent (Polyplus-Transfection) according to the instructions of the manufacturer.

Pais S.V., Key C.E., Borges V., Pereira I.S., Gomes J.P., Fisher D.J, & Mota L.J. (2019). CteG is a Chlamydia trachomatis effector protein that associates with the Golgi complex of infected host cells. Scientific Reports, 9, 6133.

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