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Trypsin

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Trypsin is a proteolytic enzyme used in cell culture applications to detach adherent cells from surfaces. It acts by cleaving peptide bonds at the carboxyl side of lysine and arginine residues, leading to the dissociation of cell-cell and cell-substrate interactions.

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Lab products found in correlation

Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions) Glutamax, by Thermo Fisher Scientific (1 mentions) Non essential amino acids (neaa), by Thermo Fisher Scientific (1 mentions) Dexamethasone (dex), by Merck Group (1 mentions) 3t3 l1 fibroblasts, by ZenBio (1 mentions) Rosiglitazone, by Adipogen (1 mentions) Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)

Spelling variants of this product

Trypsin/EDTA (4 citations) Trypsin neutralization solution (3 citations) Trypsin neutralizing solution (2 citations) Trypsin/EDTA 0.05%, (2 citations) Trypsin/EDTA solution (2 citations)

5 protocols using trypsin

1

VZV-infected PBMC Transmission Assay

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After FACS sorting of PBMCs, individual VZV-infected immune cell subsets (15,000–20,000 per subset) were resuspended in complete RPMI media and added to semi-confluent monolayers of HFLs in 96-well plates. After 3 days of co-culturing, HFLs were washed with PBS and harvested using trypsin (Sciencell) then plated on ibidi 24-well μ-Plate (ibidi, Martinsried, Germany) for an additional 48 h to allow viral infection to spread. In some experiments, sorted PBMCs were washed in citrate buffer (40 mM C6H507Na3, 135 mM KCl, pH = 3.5) for 3 min, then washed with FACS buffer before co-culturing with HFLs to confirm no potential virus was “stuck” to PBMCs, ensuring that productive viral transmission occurred from VZV-infected PBMCs as previously described [29 (link), 32 (link)]. For flow cytometry analyses of productively infected HFLs, HFLs were harvested 5 days after co-culturing as described above, followed by cell surface staining with VZV gE (R-PE) for 30 min on ice. HFLs were then fixed with 1% paraformaldehyde and analyzed using flow cytometry as described above.
Jones D., Como C.N., Jing L., Blackmon A., Neff C.P., Krueger O., Bubak A.N., Palmer B.E., Koelle D.M, & Nagel M.A. (2019). Varicella zoster virus productively infects human peripheral blood mononuclear cells to modulate expression of immunoinhibitory proteins and blocking PD-L1 enhances virus-specific CD8+ T cell effector function. PLoS Pathogens, 15(3), e1007650.
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2

Synthetic PEG Hydrogel Encapsulation of Endothelial Cells and Pericytes

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The synthetic PEG hydrogel formulation used for the encapsulation of ECs and pericytes was adapted from previous publications in our laboratory.[10 (link),16 ]At day 6 of culture, ECs were detached using Accutase treatment for 3–4 min at 37 °C, followed by 3–5 min centrifugation at 300 ×g. Pericytes were detached using 0.25% trypsin (ScienCell Research Laboratories) for 7–8 min at 37 °C, followed by 5 min centrifugation at 188 ×g. ECs (3400 cells μL−1) and pericytes (1700 cells μL−1) were resuspended in photoinitiator solution (0.1% I2959) in PBS and added 1:1 to a 2× PEG/peptide monomer solution. The final solution was pipetted into 24-well transwell inserts (Corning Falcon, 1 μm pores, 40 μL per insert) and polymerized for 4 min under a UV lamp (365 nm, 5–10 mW cm−2, UVP XX-15L, Fisher). The polymerized gels were then immersed in E7V medium supplemented with 1× nonessential amino acids (Thermo Fisher Scientific) and 1× Glutamax (Thermo Fisher Scientific) for the duration of the experiment (1 mL under transwell, 200 μL in transwell).
Kaushik G., Gil D.A., Torr E., Berge E.S., Soref C., Uhl P., Fontana G., Antosiewicz-Bourget J., Edington C., Schwartz M.P., Griffith L.G., Thomson J.A., Skala M.C., Daly W.T, & Murphy W.L. (2018). Quantitative Label-Free Imaging of 3D Vascular Networks Self-Assembled in Synthetic Hydrogels. Advanced healthcare materials, 8(2), e1801186.
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3

HUVEC Culture and PEDF Signaling

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Human umbilical vein endothelial cells (HUVECs) were cultured at 37°C in 5% CO2 in endothelial cell medium (ECM, HyClone) containing 10% fetal bovine serum (FBS, Gibco). The culture medium was replaced every 2 to 3 days. After reaching 70% to 80% confluence, the cells were harvested with 0.25% trypsin (ScienCell) and passaged at a ratio of 1 : 3.
Recombinant human SERPINF1/PEDF protein (Sino Biological Inc.) was added and incubated at 37°C at indicated time points. The p38/MAPK inhibitor SB203580 (Calbiochem) was added and incubated at 37°C for 1 h before the PEDF treatment under serum-free conditions.
He T., Zhao L., Zhang D., Zhang Q., Jia J., Hu J, & Huang Y. (2015). Pigment Epithelium-Derived Factor Induces Endothelial Barrier Dysfunction via p38/MAPK Phosphorylation. BioMed Research International, 2015, 791825.
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4

3T3-L1 Adipocyte Differentiation Protocol

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3T3-L1 fibroblasts (Zenbio, Research Triangle Park, NC, USA; ATCC, Manassas, VA, USA) were seeded (2 × 104 cells/cm2) in a plastic T25 flask (Corning Inc. Corning, NY USA) and cultured in preadipocyte media [DMEM, 44.05 mM bicarbonate, 100 µM ascorbic acid, 33 µM biotin, 17 µM pantothenate, 10% calf serum, 1% antibiotic, 4 mM L-glutamine, 1 mM sodium pyruvate, 20 mM HEPES]. At 85–90% confluency, cells were passed using trypsin (0.25%, Sciencell, Carlsbad CA) and seeded in Falcon eight-well chamber slides (1.5 × 104 cells/cm2)(Corning Inc., Corning, USA), six-well (5 × 103 cells/cm2) and 96-well plates (6 × 103 cells/cm2) for uptake experiments, Oil Red O staining, and AdipoRed− assays, respectively. Cultures were maintained in a humidified incubator at 5% CO2 and 37°C. After two days at 100% confluency, 3T3-L1 fibroblasts were induced into adipocytes by adding 0.5 mM IBMX (Acros Organics, Fisher Scientific, Hampton, NH, USA) and 0.25 µM dexamethasone (Sigma-Aldrich, St. Louis, MO, USA) in addition to the maintenance media [which is preadipocyte media and 1 µg/mL insulin, 2 µM rosiglitazone (Adipogen, San Diego, CA, USA)]. After two days on differentiation media, cells were kept in maintenance media.
Ismail A., Ayala-Lopez N., Ahmad M, & Watts S.W. (2016). 3T3-L1 cells and perivascular adipocytes are not equivalent in amine transporter expression. FEBS letters, 591(1), 137-144.
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5

Culturing HeLa and GH354 Cell Lines

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Two human cervical cancer cell lines HeLa (ATCC CCL-2) and GH354 (ATCC CRL-13003) were purchased from ATCC (Manassas, VA) and cultured in DMEM (Dulbecco's modified Eagle's medium) containing 10% fetal bovine serum (Gibco, New Zealand) and 500 μL penicillin-streptomycin (Gibco, USA) following the manufacturer's instructions. All cells were cultured in a 37°C incubator with 5% CO2 under aseptic conditions. The growth of the cells was observed daily and the medium was changed according to the growth of cells. The cells were digested with 0.05% trypsin (ScienCell, USA) for inoculation when fused to 80%.
Chen A., Xu M., Chen J., Chen T., Wang Q., Zhang R, & Qiu J. (2022). Plasma-Based Metabolomics Profiling of High-Risk Human Papillomavirus and their Emerging Roles in the Progression of Cervical Cancer. BioMed Research International, 2022, 6207701.
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