dispase  (Worthington Biochemical)

Journal: Acta physiologica (Oxford, England)

Article Title: Isolation of mitochondrial subpopulations from skeletal muscle: optimizing recovery and preserving integrity.

doi: 10.1111/apha.13182

Figure Lengend Snippet: Two distinct cell fragments found within a myofibrillar pellet after homogenization and centrifugation of dispase pellet (Scale line = 2μm) in panel (a) and (b). The sarcolemma and interfibrillar (IFM) mitochondria are identified.

Article Snippet: Reagents: Dispase (neutral protease) was obtained from Worthington Biochemical Corporation (Lakewood, NJ, USA).

Techniques: Homogenization, Centrifugation

Journal: Acta physiologica (Oxford, England)

Article Title: Isolation of mitochondrial subpopulations from skeletal muscle: optimizing recovery and preserving integrity.

doi: 10.1111/apha.13182

Figure Lengend Snippet: Two distinct fragments of a cell found within the skeletal muscle pellet treated with dispase protease (Scale line = 2μm) in panel (a) and (b). The sarcolemma, subsarcolemmal (SSM) and interfibrillar (IFM) mitochondria are identified.

Article Snippet: Reagents: Dispase (neutral protease) was obtained from Worthington Biochemical Corporation (Lakewood, NJ, USA).

Techniques:

Journal: Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease

Article Title: SMAD3 Deficiency Promotes Inflammatory Aortic Aneurysms in Angiotensin II-Infused Mice Via Activation of iNOS

doi: 10.1161/JAHA.113.000269

Figure Lengend Snippet: Defective contractile function of S3KO aorta. A through C, PE‐induced contraction of WT and S3KO aortic rings treated with placebo (A), 200 μmol/L l‐NAME (B), or 200 μmol/L aminoguanidine (C) (n=4 per group; duplicate experiments were performed). Contraction was measured at PE concentrations ranging from 10 −10 to 10 −3 mol/L, and concentration–response curves were constructed. Note that contraction is expressed as the percentage of maximal contraction induced by incubation of aortic rings in 80 mmol/L high potassium (Hi‐K + ) buffer. For placebo and l‐NAME treatment, endothelium‐intact aortic segments were used, whereas endothelium‐denuded aortas were used in aminoguanidine treatment to eliminate the vasorelaxant effect of eNOS‐derived NO. D, pEC 50 of WT and S3KO aortas in response to PE, indicating tissue sensitivity toward the drug. Note that pEC 50 is the negative logarithm of EC 50 , which refers to the concentration of a drug that induces a half‐maximal response. E, Maximal contraction force generated when incubated with 80 mmol/L Hi‐K + buffer. F, Aortic mRNA level of AngII type 1a (AT1a), type 1b (AT1b), and type 2 (AT2) receptors in WT and S3KO mice (n=3 per group; triplicate experiments were performed). Gene expression was normalized to RPL27. G through I, AngII‐induced contraction in ex vivo aortic segments treated with placebo (G), 200 μmol/L l‐NAME (H), or 200 μmol/L aminoguanidine (I) (n=5 per group; duplicate experiments were performed). Note that AngII elicited a rapid and transient contraction of infrarenal aortic segments at a concentration of 1 μg/mL, whereas minimal (

Article Snippet: Briefly, the whole aorta was dissected out from its origin at the left ventricle to the iliac bifurcation, denuded of periaortic fat, cut into small pieces (≈2‐mm segments), and incubated in 5 mL of Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal bovine serum (#16000036; Life Technologies) and 1.36 mg/mL of type II collagenase (#LS004174; Worthington Biochemical Corporation) in agitation at 115 rpm for 2 hours at 37°C.

Techniques: Concentration Assay, Construct, Incubation, Derivative Assay, Generated, Mouse Assay, Expressing, Ex Vivo

Journal: Life sciences

Article Title: Cardiac O-GlcNAcylation blunts autophagic signaling in the diabetic heart

doi: 10.1016/j.lfs.2012.06.011

Figure Lengend Snippet: The role of the hexosamine biosynthesis pathway (HBP) in blunting autophagic signaling. Representative western blots showing Beclin-1 and LC3 protein in cardiomyocytes from non-diabetic (Con) and type 2 diabetic ( db/db ) under basal conditions and in response

Article Snippet: Briefly, hearts were rapidly excised from heparinized and anesthetized mice, perfused retrogradely with Ca2+ -free perfusion buffer consisting of (in mM) 0.6 KH2 PO4 , 0.6 Na2 HPO4 , 1.2 MgSO4 • 7H2 O, 0.032 phenol red, 12 NaHCO3 , 10 KHCO3 , 10 HEPES, 30 taurine, 10 2,3-butanedione monoxime, and 5.5 glucose, pH 7.46, followed by buffer containing 12.5 μM CaCl2 and 0.4 mg/ml collagenase type 2 (Worthington).

Techniques: Western Blot

Journal: Life sciences

Article Title: Cardiac O-GlcNAcylation blunts autophagic signaling in the diabetic heart

doi: 10.1016/j.lfs.2012.06.011

Figure Lengend Snippet: The temporal progression of body weight (A) and survival (B) following either sham or trans-aortic constriction surgery to induce a pressure overload (PO) in non-diabetic (control), high fat-fed, or high fat-fed and STZ treated type 2 diabetic rats. †

Article Snippet: Briefly, hearts were rapidly excised from heparinized and anesthetized mice, perfused retrogradely with Ca2+ -free perfusion buffer consisting of (in mM) 0.6 KH2 PO4 , 0.6 Na2 HPO4 , 1.2 MgSO4 • 7H2 O, 0.032 phenol red, 12 NaHCO3 , 10 KHCO3 , 10 HEPES, 30 taurine, 10 2,3-butanedione monoxime, and 5.5 glucose, pH 7.46, followed by buffer containing 12.5 μM CaCl2 and 0.4 mg/ml collagenase type 2 (Worthington).

Techniques:

Journal:

Article Title: Exploring protein interfaces with a general photochemical reagent

doi: 10.1110/ps.051960406

Figure Lengend Snippet: Separation by size-exclusion chromatography of tryptic peptides derived from HEWL labeled with methylene carbene. After the clean-up step described above (see Fig. ), HEWL samples were reduced, carbamidomethylated, and digested with TPCK-trypsin

Article Snippet: Complete enzymatic digestion of these carbamidomethylated samples with TPCK trypsin was achieved in NH4 CO3 H (0.1 M at pH 8.0), after 12–24 h at 37°C using a 2% (w/w) enzyme:substrate ratio.

Techniques: Size-exclusion Chromatography, Derivative Assay, Labeling