Collagenase p

Manufactured by Roche

Sourced in Germany, Switzerland, United States, Japan, Sweden, France, China, United Kingdom, Spain

Collagenase P is a laboratory reagent used for the enzymatic digestion of collagen, a structural protein found in the extracellular matrix of various tissues. It is commonly used in cell isolation and tissue dissociation protocols.

Automatically generated - may contain errors

Lab products found in correlation

567 protocols using collagenase p

Islet Isolation and Purification from Rat Pancreas

Check if the same lab product or an alternative is used in the 5 most similar protocols

Pancreatic islets were isolated from rats by collagenase P (Roche Diagnostics, Milano, Italy) perfusion [28] (link) and purified by continuous-density Ficoll gradient. Briefly, the pancreas was distended by bile duct injection of 15 mL 4°C-cold 1 mg/mL collagenase P (Roche Diagnostics) diluted in HEPES-buffered Hank's balanced salt solution (HBSS) (Sigma-Aldrich), and then it was excised and minced. Islets were digested at 37°C for 20 min under constant agitation. Islets were separated from exocrin tissue by centrifugation on a Histopaque (Sigma-Aldrich) discontinuous gradient, were removed from the interface of the layers, were washed in HBSS, and finally resuspended into 10 mL of RPMI (Eurobio, Milano, Italy) supplemented with 10% fetal calf serum (FCS) (Eurobio), 1% L-glutamine (Eurobio), 10 mM glucose (Sigma-Aldrich), penicillin (50 U/ml, Eurobio), streptomycin (50 µg/ml, Eurobio), amphoterycin B (0,2 µg/ml, Eurobio) and 1% HEPES buffer (Sigma-Aldrich) in free floating culture flask. Islets were handpicked under an inverted microscope under sterile conditions and purity was assessed by Dithizone staining (Sigma-Aldrich). For each graft, the total islet mass, expressed as the 150 µm diameter islet equivalent (IE) which was calculated based on volumetric assumptions.
Pancreatic islets were incubated at 37°C (95% air and 5% CO₂) for 1–2 days before transplantation.

Quaranta P., Antonini S., Spiga S., Mazzanti B., Curcio M., Mulas G., Diana M., Marzola P., Mosca F, & Longoni B. (2014). Co-Transplantation of Endothelial Progenitor Cells and Pancreatic Islets to Induce Long-Lasting Normoglycemia in Streptozotocin-Treated Diabetic Rats. PLoS ONE, 9(4), e94783.

+ Open protocol

+ Expand

Isolation and Stimulation of Skin T Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

For freshly isolated (FI) T-cell stimulation experiments, Collagenase P (Roche, Basel, Switzerland) digestion of fresh human skin was used essentially as previously described (31 (link)). In brief, dermatome-cut skin specimens were digested in complete Roswell Park Memorial Institute (RPMI) 1640 medium (Thermo Fisher Scientific, Waltham, MA, USA) supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin (complete RPMI) containing 1 mg/ml Collagenase P (Roche) at 37°C on a rotator for 3 h. Next, 200 U/ml DNase I (Roche) were added and incubated for 15 min at 37°C. The cell suspension was diluted with 10 mM EDTA/PBS and filtered through 100- and 40-µm cell strainers. Dead cells were eliminated using a Dead Cell Removal Kit (Miltenyi Biotec, Bergisch Gladbach, Germany), and T cells were enriched (purity range: 91–98%) with CD2 microbeads (Miltenyi Biotec) according to manufacturer’s instructions. Next, either IL-9 staining was performed or cells were further stimulated. For flow-cytometric analysis of surface proteins on FI skin T cells, the Whole Skin Dissociation Kit (Miltenyi Biotec) was used according to manufacturer’s instructions to digest human skin.

Kienzl P., Polacek R., Reithofer M., Reitermaier R., Hagenbach P., Tajpara P., Vierhapper M., Gschwandtner M., Mildner M., Jahn-Schmid B, & Elbe-Bürger A. (2019). The cytokine environment influence on human skin–derived T cells. The FASEB Journal, 33(5), 6514-6525.

+ Open protocol

+ Expand

Isolation of Mouse Liver Non-Parenchymal Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

Mice were anesthetized using isoflurane, and liver NPCs were isolated following a two‐step perfusion protocol as previously described.¹⁴ Briefly, the liver was perfused in situ with calcium‐free Hank's Balanced Salt Solution (HBSS) containing 0.2 mg/mL EDTA, then perfused with 0.5 mg/mL pronase E (1074330005; Merck) and 0.75 U/mL collagenase P (11213857001; Roche) for 10 mL each. The liver was minced and further digested with HBSS containing 0.2% collagenase P, 0.4 mg/mL pronase, and 0.1 mg/mL DNase I (R104159001; Roche) in 37°C with shaking for 20 min. The digestion was passed through 40 μm cell strainer and centrifuged at 50×g for 3 min to remove the majority of hepatocytes. The NPCs supernatant suspension was collected and treated with a Dead Cell Removal Kit (130‐090‐101; Miltenyi Biotec) according to the manufacturer's instructions to remove dead cells. The resulting NPCs were subjected to scRNA‐seq analysis using 10× Genomics Chromium Single‐Cell processing. Additional analysis was then performed by using the “Seurat” (v2.3.2) package for R (v3.5) (https://www.r‐project.org/).

Cheng S., Zou Y., Zhang M., Bai S., Tao K., Wu J., Shi Y., Wu Y., Lu Y., He K., Sun P., Su X., Hou S, & Han B. (2023). Single‐cell RNA sequencing reveals the heterogeneity and intercellular communication of hepatic stellate cells and macrophages during liver fibrosis. MedComm, 4(5), e378.

+ Open protocol

+ Expand

Isolation of Mouse Pancreatic Islets

Check if the same lab product or an alternative is used in the 5 most similar protocols

Mouse islets were isolated following an injection of 0.6 mg/mL Collagenase P (Roche) into the pancreatic bile duct. Partially dissociated tissue was fractionated using a Histopaque-1077 (Sigma) gradient followed by hand-picking of islets. Islets were isolated by injection of 0.6 mg/mL Collagenase P (Roche) into the pancreatic bile duct, and partially dissociated tissue was fractionated using a Histopaque-1077 (Sigma) gradient followed by hand-picking of islets. Islet isolations were performed by the Vanderbilt Islet Procurement and Analysis Core.

Osipovich A.B., Dudek K.D., Trinh L.T., Kim L.H., Shrestha S., Cartailler J.P, & Magnuson M.A. (2023). ZFP92, a KRAB domain zinc finger protein enriched in pancreatic islets, binds to B1/Alu SINE transposable elements and regulates retroelements and genes. PLOS Genetics, 19(5), e1010729.

+ Open protocol

+ Expand

Isolation and Compression of Rat Intervertebral Disc Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

A total of 32 male Sprague-Dawley (SD) rats (8-weeks old, skeletally mature) were used for the isolation of primary rat NP cells in this study. Rats were euthanized by inhaling an excessive amount of isoflurane. Rat tails were removed under aseptic conditions, and coccygeal IVDs were separated. The gelatinous NP tissue was separated from the AF under microscope. For rat NP cells isolation, the NP tissue obtained was digested for 30 min in a mixture of 0.4% Pronase (Roche Diagnostics, 10165921001) and 0.0125% collagenase P (Roche Diagnostics, 11213865001) [ 71 ]; for human NP cells isolation, the NP tissue obtained was digested for 30 min in 0.0125% collagenase P (Roche Diagnostics, 11213865001) [ 72 ]. The digested tissue was then passed through a cell strainer (BD Falcon, 352360) with a pore size of 100 μm and was washed 3 times with phosphate-buffered saline (PBS; Gibco, 20012027). The isolated cells were maintained in Dulbecco's Modified Eagle Medium/Nutrient Mixture F-12 (DMEM/F-12; Gibco, 11320033) containing 10% fetal bovine serum (FBS; Gibco 10099141C), supplemented with 1% penicillin-streptomycin combination (Gibco, 15070063) at 37°C in a humidified atmosphere of 5% CO 2 . Passage 1 and 2 was used in this study. For in vitro compression model, the cells were placed in a compression apparatus as previously described [ 27,42,73 ].

Wang D., Shang Q., Mao J., Gao C., Wang J., Wang D., Wang H., Jia H., Peng P., Du M., Luo Z, & Yang L. (2023). Phosphorylation of KRT8 (keratin 8) by excessive mechanical load-activated PKN (protein kinase N) impairs autophagosome initiation and contributes to disc degeneration. Autophagy, 19(9).

+ Open protocol

+ Expand

Tissue Digestion and Single-Cell Suspension Preparation

Check if the same lab product or an alternative is used in the 5 most similar protocols

Ears were dissected into small pieces and digested in 500 μl RPMI containing 10% FCS and 1 mg/ml Collagenase P (Roche Diagnostic) for an initial 1 hr at 37 °C. The digested tissues were briefly spin at 2,000 rpm for 5 min and the 200 μl clear supernatant was collected and stored at -20 °C for LEGENDplex analysis. The remaining tissue pellets were resuspended thoroughly with another 1 ml of RPMI + Collagenase P with another 1 hr incubation at 37 °C, and 100 µg/ml DNase I (Roche Diagnostic) was added for the final 30 min. The digested samples were passed through 70-μm cell strainers (BD Biosciences) and the digestion was stopped with 500 µl cold 10 mM EDTA in PBS, and a single cell suspension was obtained in FACS cell staining buffer (BioLegend). Auricular lymph nodes were harvested and meshed through 70-μm cell strainers (BD Biosciences) to obtain single cell suspensions in FACS cell staining buffer (BioLegend).

Chen Y.L., Ng J.S., Babu R.O., Woo J., Nahler J., Hardman C.S., Kurupati P., Nussbaum L., Gao F., Dong T., Ladell K., Price D.A., Duncan D.A., Johnson D., Gileadi U., Koohy H, & Ogg G.S. (2023). Group A streptococcus induces CD1a-autoreactive T cells and promotes psoriatic inflammation. Science immunology, 8(84), eadd9232.

+ Open protocol

+ Expand

Isolation of Mouse Pancreatic Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

Whole adult mouse pancreas was harvested and digested in 1 mg/mL collagenase-P (Boehringer Mannheim) at 37 °C for 30 min. Following multiple washes with HBSS supplemented with 5% FBS, collagenase-digested pancreatic tissue was filtered through a 600 µm polypropylene mesh (Spectrum Laboratories) and spun down. The pellet was then diluted in trypsin (0.05%) (Mediatech) and incubated at 37 °C for 5 min. After multiple washes, cells were finally filtered through a 100 µm cell strainer and directly re-suspended in HBSS for flow cytometry.

McAllister F., Bailey J.M., Alsina J., Nirschl C.J., Sharma R., Fan H., Rattigan Y., Roeser J.C., Lankapalli R.H., Zhang H., Jaffee E.M., Drake C.G., Housseau F., Maitra A., Kolls J.K., Sears C.L., Pardoll D.M, & Leach S.D. (2014). Oncogenic Kras activates a hematopoietic-to-epithelial IL-17 signaling axis in preinvasive pancreatic neoplasia. Cancer cell, 25(5), 621-637.

+ Open protocol

+ Expand

Enzyme-based Tissue Dissociation Protocol

Check if the same lab product or an alternative is used in the 5 most similar protocols

Human NPCs were separated from the nuclear tissue by sequential enzymatic digestion, first with 0.4% pronase (Sigma, St. Louis, MO, USA) for 1 h and subsequently with 0.025% collagenase P (Boehringer Mannheim, Germany) and 0.004% DNase II (Sigma) at 37 °C overnight. After digestion, the cells were washed extensively with DMEM/F-12, then seeded in 3 fresh flasks at a density of 5000 cells/cm² and incubated in a humidified atmosphere of 5% CO₂ and 95% air until the cells attained confluence.

Lin S.S., Ueng S.W., Chong K.Y., Chan Y.S., Tsai T.T., Yuan L.J., Liu S.J., Yang C.Y., Hsiao H.Y., Hsueh Y.J., Chen C.A, & Niu C.C. (2023). Effects of Hyperbaric Oxygen Intervention on the Degenerated Intervertebral Disc: From Molecular Mechanisms to Animal Models. Cells, 12(16), 2111.

+ Open protocol

+ Expand

Isolation of Pancreatic Islets

Check if the same lab product or an alternative is used in the 5 most similar protocols

Rats and mice were killed by high CO₂ and cervical dislocation, respectively. Collagenase solution (Boehringer Mannheim Collagenase P, 1 mg/ml, pH 7.4) was injected into the main pancreatic duct. The pancreas was resected and incubated for 17 min at 37 °C. The islets were handpicked under a stereomicroscope. Whole islets were cultured in petri dishes (Sarstedt) containing RPMI 1640 (HyClone) as above but substituted with 5 mM d-glucose and 10% fetal bovine serum (for rat islets)/10 mM d-glucose and 5% fetal bovine serum (for mouse islets) and lacking 2-mercaptoethanol.

Ye Y., Barghouth M., Dou H., Luan C., Wang Y., Karagiannopoulos A., Jiang X., Krus U., Fex M., Zhang Q., Eliasson L., Rorsman P., Zhang E, & Renström E. (2022). A critical role of the mechanosensor PIEZO1 in glucose-induced insulin secretion in pancreatic β-cells. Nature Communications, 13, 4237.

+ Open protocol

+ Expand

Isolation and Culture of Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

Unless stated otherwise, all chemical reagents were obtained from Sigma (St. Louis, MO). Collagenase P was purchased from Boehringer Mannheim (Indianapolis, IN). Cell culture reagents, including Hanks’ balanced salt solution, Medium 199, fetal bovine serum (FBS) and 0.25% trypsin-EDTA, were purchased from Gibco (Gaithersburg, MD)

Benson J.D., Benson C.T, & Critser J.K. (2014). Mathematical Model Formulation And Validation Of Water And Solute Transport In Whole Hamster Pancreatic Islets. Mathematical biosciences, 254, 64-75.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!