Trypsin like enzyme

Manufactured by Thermo Fisher Scientific

Sourced in United States

Trypsin-like Enzyme is a proteolytic enzyme that catalyzes the hydrolysis of peptide bonds, typically after lysine or arginine residues. It is commonly used in cell culture and protein research applications to dissociate adherent cells and digest proteins.

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Lab products found in correlation

L glutamine, by Thermo Fisher Scientific (3 mentions) Countess hemocytometer, by Thermo Fisher Scientific (2 mentions) Growth supplement, by ScienCell (1 mentions) Gentamicin, by Harvard Bioscience (1 mentions) Foetal bovine serum (fbs), by Harvard Bioscience (1 mentions) Amphotericin b, by Harvard Bioscience (1 mentions) Amphotericin b, by Merck Group (1 mentions) Trypan blue dye exclusion, by Merck Group (1 mentions) Gentamicin, by Merck Group (1 mentions) Collagenase type h, by Roche (1 mentions) 70 μm cell strainer, by BD (1 mentions) Gibco cell culture freezing medium, by Thermo Fisher Scientific (1 mentions) Advanced minimum essential medium, by Thermo Fisher Scientific (1 mentions) Collagenase type 1, by Thermo Fisher Scientific (1 mentions) Celltiter 96 non radioactive cell proliferation assay, by Promega (1 mentions) 100 μm cell strainer, by BD (1 mentions)

4 protocols using trypsin like enzyme

Mesenchymal Stem Cell Culture and Secretome Analysis

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Additional Mesenchymal stem cells from the subjects were grown in T-75 cm² flasks at a density of (1.0–2.5 × 10³) cells/cm² in Advanced MEM* with 5 % PLTmax (Mill Creek Life Sciences, Rochester, MN, USA) and 2 mM L-glutamine (Invitrogen, Carlsbad, CA, USA) in a 37 °C and 5 % CO2 incubator for 3–4 days. When cells were 60–80 % confluent, they were passaged using TrypLE (Trypsin-like Enzyme, Invitrogen) ^{37 (link)}. Cell count was quantified with a Countess hemocytometer (Thermo Fisher Scientific, Inc., NY, USA). Aliquots of the conditioned media were collected before harvesting the cells and stored at −80 °C until they were assayed.
Hepatocyte growth factor (HGF), vascular endothelial growth factor (VEGF), epidermal growth factor (EGF) and Inter-Cellular Adhesion Molecule-1 (ICAM-1) were measured by enzyme-linked immunosorbent assays (ELISA) according to manufacturer protocols as previously described ^{28 (link)}.

Abumoawad A., Saad A., Ferguson C.M., Eirin A., Herrmann S.M., Hickson L.J., Goksu B.B., Bendel E., Misra S., Glockner J., Dietz A.B., Lerman L.O, & Textor S.C. (2019). In a Phase 1a escalating clinical trial, autologous mesenchymal stem cell infusion for renovascular disease increases blood flow and the glomerular filtration rate while reducing inflammatory biomarkers and blood pressure.. Kidney international, 97(4), 793-804.

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Isolation of Primary Human Alveolar Osteoblasts

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Primary human alveolar osteoblasts were obtained from drilling bone particles that were generated during dental implant surgery of one patient, after the informed consent form was received and signed. Briefly, harvested alveolar bone was gathered in phosphate-buffered saline with 50 mg/mL gentamicin and 2.5 mg/mL amphotericin B (both from Sigma-Aldrich Inc., St. Louis, MO, USA) by using dental drills. The bone tissue was cultured as explants in osteoblast culture medium (ObM) enriched with growth supplements (ScienCell Research Laboratories Inc., Carlsbad, CA, USA), 5% foetal bovine serum (FBS) (v/v) (Biochrom AG, Leonorenstr, Berlin, Germany), 50 mg/mL gentamicin, and 2.5 mg/mL amphotericin B. Culture medium was changed 2 times a week. Cells leaving the explants were detached with animal origin-free trypsin-like enzyme (Gibco-Invitrogen, Grand Island, NY, USA) when they covered the culture surface entirely (Figure 2A). Trypan blue dye exclusion (Sigma-Aldrich) was used for cell viability assessment. From the first subculture onwards, primary osteoblasts were maintained in ObM supplemented with 15% FBS (v/v) and 50 mg/mL gentamicin (hereafter, ObGM, osteoblast growth culture medium). Cells between the 4th and 6th passages were used in the experiments.

Anitua E., Zalduendo M., Tierno R, & Alkhraisat M.H. (2024). Plasma Rich in Growth Factors in Bone Regeneration: The Proximity to the Clot as a Differential Factor in Osteoblast Cell Behaviour. Dentistry Journal, 12(5), 122.

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Isolation and Expansion of Subcutaneous Adipose-Derived Cells

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A subcutaneous fat biopsy (approximately 2–4 g) was obtained under sterile conditions from RVD patients in an outpatient surgical suite or during the surgical nephrectomy from kidney donors. Cells were expanded ex vivo as follows. The fat tissues were minced with surgical scalpels and incubated in 2 % collagenase type H (Roche, Mannheim, Germany) for 90 min at 37 °C. Digested tissue was centrifuged at 400 g for 5 min with the pellet washed in PBS, passed through a 70-μm cell strainer (BD Biosciences, San Jose, CA, USA), and incubated in red blood cell lysis buffer (154 mM NH4Cl, 10 mM KHCO3, 0.1 mM EDTA). Cells were grown in T-75 cm² flasks at a concentration of 1.0–2.5 × 10³ cells/cm² in Advanced MEM with 5 % PLTmax (Mill Creek Life Sciences, Rochester, MN, USA) and 2 mM L-glutamine (Invitrogen, Carlsbad, CA, USA) in a 37 °C 5 % CO² incubator for 3–4 days. When cells were 60–80 % confluent, they were passaged using TrypLE (Trypsin-like Enzyme, Invitrogen) [15 (link)]. Cell yield was quantified with a Countess hemocytometer (Thermo Fisher Scientific, Inc., NY, USA).

Saad A., Zhu X.Y., Herrmann S., Hickson L., Tang H., Dietz A.B., van Wijnen A.J., Lerman L, & Textor S. (2016). Adipose-derived mesenchymal stem cells from patients with atherosclerotic renovascular disease have increased DNA damage and reduced angiogenesis that can be modified by hypoxia. Stem Cell Research & Therapy, 7(1), 128.

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Isolation and Characterization of Adipocyte Progenitors

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Adipocyte progenitors, also termed adipose tissue-derived mesenchymal stem cells or preadipocytes, were harvested from abdominal subcutaneous fat samples as previously described [48 (link),49 (link)]. Adipose biopsies were minced and digested in 2% collagenase type I at 37 °C for 45 min (Gibco, Waltham, MA), filtered through a 100 μm cell strainer (BD Biosciences, San José, CA) to remove remaining tissue pieces, and centrifuged to pellet cells. Cells were cultured in advanced minimum-essential-medium (Thermo Fisher Scientific, Waltham, MA) supplemented with 5% platelet lysate (PLTMax, Mill Creek Life Sciences, Rochester, MN) and 2 mM l-glutamine (Invitrogen, Carlsbad, CA) in a 37 °C/5% CO₂ incubator for 3–4 days. When 60–80% confluent, cells were passaged using TrypLE (Trypsin-like Enzyme, Invitrogen, Invitrogen, Waltham, MA). The third passage cells were collected and kept in Gibco Cell Culture Freezing Medium (Life Technologies, Carlsbad, CA) at −80 °C. Change in adipocyte progenitor numbers over time was measured in passage 3–4 cultures using a tetrazolium compound (3-[4,5-dimethylthiazol-2-yl]-5-[3-carboxymethoxyphenyl]-2-[4-sulfophenyl]-2H-tetrazolium, inner salt; MTS) assay (CellTiter 96 Non-Radioactive Cell Proliferation Assay; Promega, Madison, WI) according to the manufacturer's instructions.

Hickson L.J., Langhi Prata L.G., Bobart S.A., Evans T.K., Giorgadze N., Hashmi S.K., Herrmann S.M., Jensen M.D., Jia Q., Jordan K.L., Kellogg T.A., Khosla S., Koerber D.M., Lagnado A.B., Lawson D.K., LeBrasseur N.K., Lerman L.O., McDonald K.M., McKenzie T.J., Passos J.F., Pignolo R.J., Pirtskhalava T., Saadiq I.M., Schaefer K.K., Textor S.C., Victorelli S.G., Volkman T.L., Xue A., Wentworth M.A., Wissler Gerdes E.O., Zhu Y., Tchkonia T, & Kirkland J.L. (2019). Senolytics decrease senescent cells in humans: Preliminary report from a clinical trial of Dasatinib plus Quercetin in individuals with diabetic kidney disease. EBioMedicine, 47, 446-456.

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