Collagenase type 1

Manufactured by Worthington

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Collagenase type I is an enzyme used to digest collagen, which is a key structural component of the extracellular matrix. It is commonly used in cell isolation and tissue dissociation procedures.

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Lab products found in correlation

Close spelling variants of this product

Collagenase type I and III Bacterial collagenase type I Collagenase type I and IV Collagenase type I solution Collagenase type 1 and 2 Collagenase Type I from Clostridium Histolyticum Collagenase I Type I Collagenase

694 protocols using collagenase type 1

Isolation of Keratinocytes from Mouse Skin

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To isolate keratinocytes from mouse back skin, shaved skin was floated
on 0.25% Trypsin without EDTA (Life Technologies) for 2 hours at 37°C.
Then the epidermis was scraped off the dermis, and cells were disaggregated by
gentle mincing with a scalpel and pipetting. For back skin in late anagen the
dermis was further minced and digested for 30 minutes at 37°C low-calcium
medium containing 1.25 mg/ml of Collagenase Type I, 0.5 mg/ml of Collagenase
Type II, 0.5 mg/ml of Collagenase Type IV (all from Worthington) and 0.1 mg/ml
of Hyaluronidase (Sigma-Aldrich).
To disaggregate cells from squamous tumours, the tumours were minced
with a scalpel and incubated for 1-2 hours at 37°C in low-calcium medium
containing 1.25 mg/ml of Collagenase Type I, 0.5 mg/ml of Collagenase Type II,
0.5 mg/ml of Collagenase Type IV (all from Worthington) and 0.1 mg/ml of
Hyaluronidase (Sigma-Aldrich). Then pieces were further incubated for another
hour with Trypsin without EDTA (Life Technologies) and cells disaggregated by
scraping with a scalpel blade. Trypsin was inactivated by washing the cell
suspension with low-calcium media containing 10% of FBS (Life Technologies).

Blanco S., Bandiera R., Popis M., Hussain S., Lombard P., Aleksic J., Sajini A., Tanna H., Cortés-Garrido R., Gkatza N., Dietmann S, & Frye M. (2016). Stem cell function and stress response are controlled by protein synthesis. Nature, 534(7607), 335-340.

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Isolation of Keratinocytes from Mouse Skin

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Glycocalyx Modification of Tumor Cells

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Tumor cells were detached from the different substrates after 7 days of culture. Cells cultured on tissue culture polystyrene were collected using 0.25% trypsin-EDTA. Cells cultured on collagen and mineralized collagen were collected using collagenase type I (1mg/mL in PBS, Worthington Biochemical) and passed through a 40 μm cell strainer. MCF10A-derived 1E7 cells cultured on tissue culture polystyrene were induced with 1 μg/mL doxycycline (204734, Santa Cruz Biotechnology) for 24 hours before being collected with 0.05% trypsin-EDTA.
For treatment with glycocalyx-editing enzymes, detached cells were incubated with 100 nM StcE mucinase or 100 nM Salmonella typhimurium sialidase or 2.5 U/mL Streptomyces hyalurolyticus hyaluronidase (H1136, Millipore Sigma) or 5,000 U/mL PNGase F (P0704S, New England BioLabs) in growth media for 1 hour at 37°C. Subsequently, cells were rinsed thoroughly twice with growth media and used further. For treatment with the sialylation inhibitor P-3F AX -Neu5Ac, tumor cells were cultured on collagen and mineralized collagen for 6 days before incubation with 100 μM P-3F AX -Neu5Ac in growth media for another 24 hours at 37°C in 5% CO 2 . Subsequently, cells were collected using collagenase type I (1mg/mL in PBS, Worthington Biochemical) and passed through a 40 μm cell strainer before use in subsequent assays.

Park S., Choi S., Shimpi A.A., Estroff L.A., Fischbach C, & Paszek M.J. (2024). Collagen Mineralization Decreases NK Cell-Mediated Cytotoxicity of Breast Cancer Cells via Increased Glycocalyx Thickness. Advanced materials (Deerfield Beach, Fla.).

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Glycocalyx Editing and Sialylation Inhibition

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Tumor cells were detached from the different substrates after 7 days of culture. Cells cultured on tissue culture polystyrene were collected using 0.25% trypsin-EDTA. Cells cultured on collagen and mineralized collagen were collected using collagenase type I (1mg/mL in PBS, Worthington Biochemical) and passed through a 40 μm cell strainer. MCF10A-derived 1E7 cells cultured on tissue culture polystyrene were induced with 1 μg/mL doxycycline (204734, Santa Cruz Biotechnology) for 24 hours before being collected with 0.05% trypsin-EDTA.
For treatment with glycocalyx-editing enzymes, detached cells were incubated with 100 nM StcE mucinase or 100 nM Salmonella typhimurium sialidase or 2.5 U/mL Streptomyces hyalurolyticus hyaluronidase (H1136, Millipore Sigma) or 5,000 U/mL PNGase F (P0704S, New England BioLabs) in growth media for 1 hour at 37°C. Subsequently, cells were rinsed thoroughly twice with growth media and used further. For treatment with the sialylation inhibitor P-3F_AX-Neu5Ac, tumor cells were cultured on collagen and mineralized collagen for 6 days before incubation with 100 μM P-3F_AX-Neu5Ac in growth media for another 24 hours at 37°C in 5% CO₂. Subsequently, cells were collected using collagenase type I (1mg/mL in PBS, Worthington Biochemical) and passed through a 40 μm cell strainer before use in subsequent assays.

Park S., Choi S., Shimpi A.A., Estroff L.A., Fischbach C, & Paszek M.J. (2024). COLLAGEN MINERALIZATION DECREASES NK CELL-MEDIATED CYTOTOXICITY OF BREAST CANCER CELLS VIA INCREASED GLYCOCALYX THICKNESS. bioRxiv.

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Isolation and Culture of Adipose-Derived and Synovial Mesenchymal Stem Cells

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Adipose-derived MSCs (ASCs) were obtained as previously described [24 ]. Briefly, the adipose tissue was enzymatically digested (37 °C, 30 min) by 0.075% w/v type I collagenase (Worthington Biochemical Co., Lakewood, NJ, USA). After digestion, samples were filtered through a cell strainer (100 μm) and centrifuged (1000×g, 5 min). Released cells were seeded at 5 × 10³ cells/cm² in DMEM supplemented with 10% FBS (GE Healthcare, Piscataway, NJ, USA) and pen/strepto (Life Technology, Carlsbad, CA, USA). Synovial membranes were minced in small pieces and enzymatically digested (37 °C, 3 h) by 0.25% w/v type I collagenase (Worthington Biochemical Co., Freehold, NJ, USA). After digestion, samples were filtered through a cell strainer (100 μm) and centrifuged (376×g, 5 min). Cells were seeded at 5000 cell/cm² density and fibroblast-like synoviocytes (FLSs) selected for plastic adherence [25 ]. ASCs and FLSs were cultured in DMEM supplemented with 10% FBS and pen/strepto. All cell types were maintained in an incubator at 37 °C in a humidified atmosphere with 5% CO₂ and used for the following experiments between passages 3 and 5.

Ragni E., Perucca Orfei C., De Luca P., Lugano G., Viganò M., Colombini A., Valli F., Zacchetti D., Bollati V, & de Girolamo L. (2019). Interaction with hyaluronan matrix and miRNA cargo as contributors for in vitro potential of mesenchymal stem cell-derived extracellular vesicles in a model of human osteoarthritic synoviocytes. Stem Cell Research & Therapy, 10, 109.

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Adipose Tissue Emigration and Enzymatic Digestion

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To obtain emigrants, subcutaneous fat was removed and put in the culture medium for 2 days at 37 °C and then analyzed on day 2. For enzymatic digestion, subcutaneous fat was placed in PBS (Gibco) containing 2.5 mg/ml dispase II (Roche Diagnostics, Indianapolis, IN) overnight at 4 °C and/or 0.15% collagenase type Ⅰ (Worthington Biochemical, Lakewood, NJ) for 60 minutes at 37 °C.

Sato T., Ogawa Y., Ishikawa A., Nagasaka Y., Kinoshita M., Shiokawa I., Shimada S., Momosawa A, & Kawamura T. (2022). Revisiting the Experimental Methods for Human Skin T-Cell Analysis. JID Innovations, 2(4), 100125.

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Skin Cell Isolation and Emigration

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To obtain epidermal and dermal sheets, the subcutaneous fat and deep dermis were thoroughly removed using a pair of scissors. The parts of the skin composed of the epidermis and upper dermis were cut into approximately 20 × 10 mm square pieces and then incubated with dispase II (2.5 mg/ml; Roche Diagnostics) dissolved with PBS (Gibco) overnight at 4 °C. On day 1, to make a single-cell suspension from the epidermis and dermis, the epidermal sheets were incubated with collagenase type IV (200 U/ml; Worthington Biochemical) for 30 minutes at 37 °C, whereas the dermal sheets were incubated with PBS solution containing 0.2% collagenase type Ⅰ (Worthington Biochemical) and 0.05% DNase I (Sigma-Aldrich, St. Louis, MO) for 120 minutes at 37 °C. After incubation, the epidermal and dermal sheets were divided into small pieces using a pair of forceps. To generate single-cell suspensions, these pieces were aspirated with a 50-cc syringe up and down 5 times for epidermal suspension and 10 times for the dermal and subcutaneous fat suspension. This was then filtered thrice through a sterile mesh, with subsequent analysis on the same day (day 1). To obtain emigrants, the epidermal or dermal sheets were floated separately in the culture medium for 2 days at 37 °C. On day 3, these emigrants were analyzed.

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Adipose Tissue Isolation and RNA Extraction

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Minced adipose tissue samples were treated with 1 mg/mL Collagenase Type I (Worthington Biochemical Corporation) in Krebs-Ringer Buffer + 10% FCS + 1% Penicillin/Streptomycin + 2% BSA, and incubated at 37°C for 60 min. Dispersed cells w ere centrifuged at 500 g for 5 min. The precipitated cells from stromal vascular fractions were centrifuged at 500 g for 5 min and resuspended in DMEM supplemented with 10% FCS and 1% Penicillin/Streptomycin twice. Total mRNA was extracted from pelleted fractions as described above.

Martinez-Lopez N., Tarabra E., Toledo M., Garcia-Macia M., Sahu S., Coletto L., Batista-Gonzalez A., Barzilai N., Pessin J.E., Schwartz G.J., Kersten S, & Singh R. (2017). System-wide benefits of intermeal fasting by autophagy. Cell metabolism, 26(6), 856-871.e5.

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Isolation and Culture of Stem Cells from Apical Papilla

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Tooth tissue acquisition complied with the approved guidelines established by Beijing Stomatological Hospital, Capital Medical University, with informed patient consent. The third molar was disinfected with 75% ethanol and then washed with phosphate‐buffered saline (PBS). We isolated and cultured the SCAPs, as described previously,23 and then identified the cell type. Briefly, SCAPs were separated from the apical papilla tissues. The tissues were separately digested in a solution containing 3 mg/mL collagenase type I (Worthington Biochem, Lakewood, NJ, USA) and 4 mg/mL dispase (Roche, Basel, Switzerland) for 1 hour at 37°C. Single‐cell suspensions were obtained by passing the cells through a 70‐μm strainer (Falcon; BD Labware, San Jose, CA, USA). WJCMSCs and BMSCs were purchased from ScienCell Research Laboratories (San Diego, CA, USA). MSCs were cultured in complete medium containing MEM alpha‐modified Eagle's medium (Invitrogen, Carlsbad, CA, USA), 15% foetal bovine serum (FBS; Invitrogen), 2 mmol/L glutamine, 100 U/mL penicillin and 100 µg/mL streptomycin (Invitrogen). The medium was replaced every 3 days. The cultured MSCs were placed in a humidified 5% CO₂ incubator at 37°C. Human embryonic kidney 293 T cells were maintained in complete DMEM medium with 10% foetal bovine serum (FBS; Invitrogen).

Lin X., Yang H., Wang L., Li W., Diao S., Du J., Wang S., Dong R., Li J, & Fan Z. (2018). AP2a enhanced the osteogenic differentiation of mesenchymal stem cells by inhibiting the formation of YAP/RUNX2 complex and BARX1 transcription. Cell Proliferation, 52(1), e12522.

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Isolation of Canine Adipose-Derived Cells

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Adipose tissue was aseptically collected under anesthesia from subcutaneous fat of a two-year-old dog during castration, with permission from the owners. Tissue was minced into small pieces for enzymatic digestion with 0.2% collagenase type I (Worthington Biochemical Corporation, Lakewood, NJ, USA) for 50 min at 37 °C. Cells were plated and expanded in Dulbecco's Modified Eagle Medium (DMEM) low glucose supplemented with 20% fetal bovine serum (FBS, Thermo Fisher Scientific, Waltham, MA, USA), 2 mM L-glutamine, and 1% penicillin-streptomycin. Cultures were expanded, and experiments were performed at passage 3.

Escalhão C.C., Ramos I.P., Hochman-Mendez C., Brunswick T.H., Souza S.A., Gutfilen B., dos Santos Goldenberg R.C, & Coelho-Sampaio T. (2017). Safety of Allogeneic Canine Adipose Tissue-Derived Mesenchymal Stem Cell Intraspinal Transplantation in Dogs with Chronic Spinal Cord Injury. Stem Cells International, 2017, 3053759.

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