Plat e cells

Manufactured by Promega

Sourced in United States

Plat-E cells are a laboratory-generated cell line designed for efficient retroviral vector production. They provide a stable platform for the high-titer production of retroviral particles, which can be used for a variety of applications, such as gene delivery and gene expression studies.

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Lab products found in correlation

Plat e cells, by Cell Biolabs (4 mentions) Fugene hd, by Promega (3 mentions) Giemsa stain, by Merck Group (1 mentions) Puromycin, by Merck Group (1 mentions) Fugene 6, by Promega (1 mentions) Polybrene, by Merck Group (1 mentions) Opti mem reduced serum media, by Thermo Fisher Scientific (1 mentions)

4 protocols using plat e cells

Lentiviral Transduction and Clonal Expansion

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Plat-E cells (Cell Biolabs) were maintained in complete media supplemented with 1 μg/ml puromycin and 10 μg/ml blasticidin and plated at ∼80% confluency the day before transfection. Plat-E cells were transfected with FuGENE HD (Promega) and transfection regent was incubated overnight before a media change. Viral supernatants were collected at ∼48, 56, 72, and 80 h post-transfection and incubated with MEFs in the presence of 8 mg/ml polybrene. ∼16 h after the last viral transduction, MEF cells were split and selection medium was added if needed (1 μg/ml puromycin or 200 μg/ml hygromycin).
Clonal populations were generated by plating cells at very low density and clones were collected onto sterile filter paper dots soaked in trypsin. After expansion, whole-cell extracts from clonal populations were screened by Western blot analysis for mitofusin against wild-type controls.

Samanas N.B., Engelhart E.A, & Hoppins S. (2020). Defective nucleotide-dependent assembly and membrane fusion in Mfn2 CMT2A variants improved by Bax. Life Science Alliance, 3(5), e201900527.

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Retrovirus-mediated Hras V12 Transformation Assay

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Plat-E cells (Cell Biolabs, San Diego, CA, USA) were used to produce retrovirus according to the manufacturer's instructions. Plat-E cells were transfected with pBABE-puro and pBABE-puro Hras V12 (a generous gift from Martine F. Roussel/Scott Lowe) by using FuGENE 6 (Promega, Fitchburg, WI, USA). Retroviral supernatant was filtered and mixed with 4 μg/ml polybrene (EMD Millipore). MEF cells were infected with retroviral supernatant twice and selected in the presence of 2 μg/ml of puromycin (Sigma-Aldrich, St. Louis, MO, USA) for 4 days.
Focus-formation assays were performed as previously described.^{35 (link)} Briefly, 10³ MEF cells infected with retrovirus were mixed with 3 × 10⁵ uninfected MEF cells, and the mixture was cultured in a 100-mm dish in triplicate. Medium was changed every 2 or 3 days. After 2 weeks incubation, foci were stained with giemsa stain (Sigma-Aldrich), and the number of foci was counted.

Ogi H., Sakuraba Y., Kitagawa R., Xiao L., Shen C., Cynthia M.A., Ohta S., Arnold M.A., Ramirez N., Houghton P.J, & Kitagawa K. (2015). The oncogenic role of the cochaperone Sgt1. Oncogenesis, 4(5), e149-.

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Generating Mitofusin-expressing Clonal MEFs

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Plat-E cells (Cell Biolabs) were maintained in complete media supplemented with 1 μg/ml puromycin and 10 μg/ml blasticidin and plated at ∼80% confluency the day before transfection. Plat-E cells were transfected with FuGENE HD (Promega) and transfection regent was incubated overnight before a media change. Viral supernatants were collected at ∼48, 56, 72, and 80 h posttransfection and incubated with MEFs in the presence of 8 mg/ml polybrene. Approximately 16 h after the last viral transduction, MEF cells were split and selection was added if needed (1 μg/ml puromycin or 200 μg/ml hygromycin).
Clonal populations were generated by plating cells at very low density and clones were collected onto sterile filter paper dots soaked in trypsin. Following expansion, whole cell extract from clonal populations was screened by Western blot analysis for mitofusin against wild-type controls.

Sloat S.R., Whitley B.N., Engelhart E.A, & Hoppins S. (2019). Identification of a mitofusin specificity region that confers unique activities to Mfn1 and Mfn2. Molecular Biology of the Cell, 30(17), 2309-2319.

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Overexpression of Mitofusin Proteins in MEFs

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Plat-E cells (Cell Biolabs) were maintained in complete media supplemented with 1 μg/ml puromycin and 10 μg/ml blasticidin and plated at ∼80% confluency the day before transfection. 350,000 Plat-E cells were plated in a six-well dish and the following day were transfected with pBABE plasmids (3 μg pBABE Mfn1 or an empty vector; 1.5–3 μg pBABE Mfn2; 3 μg mito-paGFP) using FuGENE HD (Promega) and Opti-MEM reduced serum media (Gibco). Transfection reagent was incubated overnight before a media change. Viral supernatants were collected at ∼48 and 72 h post-transfection and incubated with MEFs in the presence of 8 μg/ml polybrene. ∼24 h after the last viral transduction, MEF cells were split and selection was added (1 μg/ml puromycin or 200 μg/ml hygromycin B).

Sloat S.R, & Hoppins S. (2022). A dominant negative mitofusin causes mitochondrial perinuclear clusters because of aberrant tethering. Life Science Alliance, 6(1), e202101305.

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