Papain buffer

Manufactured by Merck Group

Sourced in United States

Papain buffer is a solution used to maintain the optimal pH and ionic environment for the activity of the enzyme papain. Papain is a cysteine protease often used in biological research and applications. The buffer helps to preserve the structural integrity and catalytic function of papain during experiments or processes.

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Lab products found in correlation

L cysteine, by Merck Group (2 mentions) Arabinosylcytosine, by Merck Group (2 mentions) Basal medium eagle, by Merck Group (2 mentions) Blyscan assay, by Biocolor (1 mentions) Lrrk2 in 1, by Bio-Techne (1 mentions) Chondroitin sulfate c, by Merck Group (1 mentions) Tetrodotoxin ttx, by Merck Group (1 mentions) Mli 2, by Bio-Techne (1 mentions) Bicuculline, by Merck Group (1 mentions) Sodium phosphate, by Merck Group (1 mentions) Blyscan gag assay kit, by Biocolor (1 mentions)

Close spelling variants of this product

Papain lysis buffer Papain

6 protocols using papain buffer

Quantifying Sulfated Glycosaminoglycans in Cartilage

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After 0, 7, 14, and 21 days of culture, each pellet was digested for 18 h at 65°C in 200 mg/ml papain buffer (Sigma–Aldrich) and dissolved in 200 mM phosphate buffer containing 8 mg/ml sodium acetate, 0.8 mg/ml cysteine‐HCl, and 4 mg/ml EDTA. The sulfated glycosaminoglycan (sGAG) concentration of the supernatant was determined by the Blyscan assay (Biocolor Ltd, Newtonabbay, Ireland), according to the manufacturer's instructions.

Naritomi M., Mizuno M., Katano H., Ozeki N., Otabe K., Komori K., Fujii S., Ichinose S., Tsuji K., Koga H., Muneta T, & Sekiya I. (2018). Petaloid recombinant peptide enhances in vitro cartilage formation by synovial mesenchymal stem cells. Journal of Orthopaedic Research, 37(6), 1350-1357.

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Chondrogenesis Glycosaminoglycan Quantification

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At day 21 of chondrogenesis, cultures were washed in PBS and frozen at -80 °C before their digestion in 400μg/ml Papain buffer (Sigma Aldrich) supplemented with 0.2M Sodium Phosphate + 5mM Ethylenediaminetetraacetic acid + 5mM L-Cysteine (all Sigma Aldrich) at 65 °C for 18 h. Glycosaminoglycans were quantified from Papain-digested lysates using the GAG assay kit by Blyscan™ in which GAGs were dyed with 1,9-dimethyl-methylene blue and subsequently dissociated with Propan-1-ol solution before quantification on an absorbance spectrophotometre at 640nm. Values were normalised to levels of dsDNA, which were quantified using the PicoGreen assay.

Taheem D.K., Foyt D.A., Loaiza S., Ferreira S.A., Ilic D., Auner H.W., Grigoriadis A.E., Jell G, & Gentleman E. (2018). Differential Regulation of Human Bone Marrow Mesenchymal Stromal Cell Chondrogenesis by Hypoxia Inducible Factor‐1α Hydroxylase Inhibitors. Stem Cells (Dayton, Ohio), 36(9), 1380-1392.

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Preparation of Primary Neuronal Cultures

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Primary neuronal cultures from cortex, striatum and hippocampus from pups at P0 were prepared as described previously^{55 (link)}. In brief, neurons were dissociated by papain buffer (Sigma Aldrich), and were then placed in poly-D-lysine slides (BD) or plates in Basal Eagle Medium (Sigma-Aldrich). Arabinosylcytosine (Sigma-Aldrich) was used to inhibit glial cell growth. Forskolin (FSK), PKI (PKA inhibitor) and LRRK2 kinase inhibitor LRRK2-IN-1 (Tocris Bioscience, Bristol, UK) as well as all the other compounds were added directly to the medium of cortical neurons from stock solution and unless otherwise stated these compounds were from Calbiochem.

Parisiadou L., Yu J., Sgobio C., Xie C., Liu G., Sun L., Gu X.L., Lin X., Crowley N.A., Lovinger D, & Cai H. (2014). LRRK2 regulates synaptogenesis and dopamine receptor activation through modulation of PKA activity. Nature neuroscience, 17(3), 367-376.

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Quantifying Cell-Associated Glycosaminoglycans

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After treatment for 72 h, the agarose cell specimens were dissolved and centrifuged at 300 g for 10 mins at 4 °C, which provided a pellet containing cells with their cell-associated matrix. To determine the GAG amounts, the pellets were digested for 12 hrs at 55 °C in papain buffer (200 μg/mL papain in 50 mmol/L EDTA, 5 mmol/L L-cysteine, pH 3.0) (Sigma Chemical, USA). GAG was quantified using the dimethylmethylene blue assay.33 (link) The metachromatic reaction of GAG with dimethylmethylene blue was monitored using a spectrophotometer, and the A₅₄₀/A₅₉₅ ratio was used to determine the amount of GAG using chondroitin sulfate C (Sigma Chemical, USA) as a standard.

Zuo S., Zou W., Wu R.M., Yang J., Fan J.N., Zhao X.K, & Li H.Y. (2019). Icariin Alleviates IL-1β-Induced Matrix Degradation By Activating The Nrf2/ARE Pathway In Human Chondrocytes. Drug Design, Development and Therapy, 13, 3949-3961.

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Isolation of Postnatal Striatal Neurons

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Primary neuronal cultures from the striatum of postnatal day 0 (P0) pups were prepared as described previously [24 (link)]. In brief, neurons were dissociated by papain buffer (Sigma), and were then placed in the poly-D-lysine coated slides (BD) or plates in Basal Eagle Medium (Sigma). Arabinosylcytosine (Sigma) was used to inhibit glial cell growth. Tetrodotoxin (TTX, Sigma), Bicuculline (Sigma) and LRRK2 kinase inhibitor MLi-2 (Tocris Bioscience, Bristol, UK) were added directly to the medium of striatal neurons from the stock solutions.

Chen X., Xie C., Tian W., Sun L., Wang Z., Hawes S., Chang L., Kung J., Ding J., Chen S., Le W, & Cai H. (2020). Parkinson’s disease-related Leucine-rich repeat kinase 2 modulates nuclear morphology and genomic stability in striatal projection neurons during aging. Molecular Neurodegeneration, 15, 12.

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Chondrogenic GAG Quantification Protocol

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At day 21 of chondrogenesis, cultures were washed in phosphate buffered saline and frozen at −80°C before their digestion in 400 μg/ml Papain buffer (Sigma‐Aldrich) supplemented with 0.2M Sodium Phosphate + 5 mM Ethylenediaminetetraacetic acid + 5 mM l‐Cysteine (all Sigma‐Aldrich) at 65°C for 18 hours. GAGs were quantified from Papain‐digested lysates using a Blyscan GAG assay kit (Biocolor, UK) in which GAGs were dyed with 1,9‐dimethyl‐methylene blue and subsequently dissociated with Propan‐1‐ol solution before quantification on an absorbance spectrophotometer at 640 nm. Values were normalized to levels of dsDNA, which were quantified using the PicoGreen assay.

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