Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.4 (trypsin)
 42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The procedure for isolation and purification of Pasteurella multocida serovariant D toxin has been described. It includes the three steps of protein precipitation from cultural filtrates by 70% ammonium sulfate, chromatography of the concentrated material on Ultragel AcA44 gel-filtration on Sephracryl S-200. The proposed technique permits one the 155-fold purification of the preparation with 32.6% yield estimated by biological activity. The obtained purified preparation is homogeneous in polyacrylamide gel electrophoresis. The immunological methods also confirm the homogeneity of the preparation. The minimal dermonecrotic dose for guinea pigs of the purified 120 kDa toxin is 78 ng and LD50 for mice is 280 ng. Pasteurella multocida toxin is found to be a thermolabile protein sensitive to trypsin, glutaraldehyde and formaldehyde treatments.
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PMID:[Isolation and certain properties of the dermonecrotic toxin from Pasteurella multocida]. 178 3

We have devised a simple procedure for immunostaining of sections that have previously undergone autoradiographic visualization of mRNAs by in situ hybridization. Classical hybridocytochemistry techniques were performed first on cryostat sections of formaldehyde-fixed tissue. Standard methods were used for slide coating by emulsion dipping and for revelation, fixation, and coverslipping steps. The key to this method is the emulsion removal, or permeabilization, by a short trypsin incubation (0.2% for 20-30 sec) which facilitates the good access of antibodies used in a subsequent immunocytochemical technique to section epitopes. Usual immunofluorescence and immunoperoxidase procedures were successfully performed after this treatment. The immunoreactivity of several neuropeptides was well preserved after this procedure. In addition to its usefulness in our studies, this general method should be applicable to many other situations in which autoradiographic and immunocytochemical detections must be coupled.
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PMID:A simple method for coupling in situ hybridization and immunocytochemistry: application to the study of peptidergic neurons. 203 43

Results of the comparative study of trypsin- and chymotrypsin-like serine proteases from pyloric caeca of salmon fishes and trypsin and chymotrypsin of bulls are presented in the paper. The hydrolytic activity of salmon proteases with respect to methyl ethers of N-benzoyl-L-leucine is 2.4 times higher than that of bull chymotrypsin, but with respect to methyl esters of N-benzoyl-L-tyrosine and N-benzoyl-L-arginine the activity of salmon proteases is 6.5 and 80 times lower than that of bull chymotrypsin and trypsin. Salmon proteases in contrast to bull trypsin and chymotrypsin hydrolyze but slightly N-glutaryl-L-phenylalanine para-nitroanilide. It shown that fish proteases are not absolutely specific to synthetic substrates, which is a result of their less pronounced (than in case of bull trypsin and chymotrypsin) differences in structures of binding centres. The study of the salmon protease interaction with some immobilized ligands has confirmed the higher affinity of enzymes to reagents with two space-separated aromatic rings in their composition. It is supposed that salmon proteases interact with such reagents through two sites: hydrophobic "pockets" and probably additional binding site of the active centre. The salmon protease preparation demonstrates higher resistance to inactivating action of formaldehyde within the range of concentrations 2-16% than bull chymotrypsin does.
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PMID:[Comparative study of the properties of serine proteases of lower and higher vertebrates]. 208 90

The mechanism whereby cytolytic lymphocytes protect themselves from killing mediated by their own cytotoxic protein, perforin, was studied. By using a competition assay, we demonstrated that the resistance of cells to perforin-mediated cytolysis is inversely correlated with their ability to absorb perforin, with tumor cells and noncytotoxic lymphocytes that are susceptible to perforin-mediated lysis being able to absorb perforin from the supernatant much better than CTL. The evidence implies that there is molecule on cytolytic lymphocytes that interferes with perforin-binding activity, resulting in the inability of perforin to lyse these cells. The molecule is most likely a surface protein or complex of proteins because its activity decreases after CTL treatment with the proteolytic enzymes trypsin and papain, and the activity can be recovered by incubation of the treated CTL cells at 37 degrees C for 6 h. The recovery can be blocked by emetine, cycloheximide, and actinomycin D, inhibitors of protein and RNA/DNA synthesis. The protein contains carbohydrate groups that play an important role in the function of the protein, as indicated by the fact that inhibition of glycosylation by tunicamycin and cleavage of sialic acid from the protein with neuraminidase result in a significant increase of perforin binding to CTL. Cross-linkage of CTL membrane proteins with glutaraldehyde and formaldehyde and blockage of the functional domains of the protein with an antiserum against CTL also inhibit the activity of this protein. Temperature-dependence studies that allow for a dissociation of the binding and pore-forming stages of perforin-mediated hemolysis suggest that the protective protein interferes at the perforin-binding stage.
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PMID:Resistance of cytolytic lymphocytes to perforin-mediated killing. Inhibition of perforin binding activity by surface membrane proteins. 210 15

Human deciduous teeth undergoing physiologic root resorption were extracted and fixed with a mixture of formaldehyde and glutaraldehyde and processed for scanning (SEM) and transmission (TEM) electron microscopy, and for acid (ACPase) and alkaline phosphatase (ALPase) cytochemistry. The resorbant organ, rich in odontoclasts, cementoblasts, fibroblasts, and macrophages, formed prominent resorption lacunae in root dentin. SEM observations of resorption lacunae treated with trypsin solution showed islands of newly-formed cementum matrix in part of the resorbing dentin surfaces. Such cementum consisted of bundles of densely-arranged collagen fibrils and, in part, contained forming cementocytic lacunae and canaliculi. Active cementoblasts adjacent to odontoclasts on resorbing dentin surfaces showed cuboidal outlines and were characterized by the presence of numerous cisterns of rough endoplasmic reticulum, well-developed Golgi complexes, secretion granules, and many mitochondria. They sometimes formed a thin layer of cementoid and/or cementum matrix upon the resorbing dentin surface. These cementoblasts had ACPase-positive lysosomes in the cell bodies and exhibited intense ALPase activity along the plasma membranes of whole cell surfaces. These results suggest that, during root resorption, 1) active cementoblasts are present adjacent to active odontoclasts and 2) these cementoblasts are involved in remodeling the resorbing dentin surfaces.
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PMID:Possible role of cementoblasts in the resorbant organ of human deciduous teeth during root resorption. 214 74

A number of fixation and decalcification procedures were evaluated to determine their suitability for immunohistochemistry on trephine samples of bone marrow after paraffin embedding. In particular, the immunoreactivity of antigens characteristic for various hematopoietic cell lines (immunoglobulin heavy and light chains for plasmacytoid cells; elastase for neutrophil myeloid cells; lysozyme, alpha-1-antitrypsin and alpha-1-antichymotrypsin for hystiocytic cells; leukocyte common antigen for lymphocytes; hemoglobin and glycophorin A for erythroid cells; Factor VIII-related antigen for thrombocytoid cells) as well as some antigens specific for epithelial tumors (CEA, 115D8, and keratin) were investigated. Fixation in a mercuric chloride-formaldehyde mixture followed by decalcification in acetic acid-formaldehyde-saline proved to be the best procedure for antigen preservation and retention of morphologic detail. Moreover, there is no need of trypsinization when using this procedure. The only exception was Factor VIII-related antigen in megakaryocytes, which was best demonstrated in trypsin-digested sections of formalin-fixed and acetic acid-decalcified biopsies.
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PMID:Influence of fixation and decalcification on the immunohistochemical staining of cell-specific markers in paraffin-embedded human bone biopsies. 241 61

Paraffin sections of formaldehyde-fixed renal biopsies were labeled for complement C3 by a polyclonal rabbit antibody to human complement C3, by the peroxidase-antiperoxidase complex (PAP) and the avidin-biotin peroxidase complex (ABC) techniques, respectively. All tissues had C3 deposits according to direct immunofluorescence on fresh frozen sections. Staining for muramidase was introduced as an intrinsic control for the degree of tissue proteolysis after the necessary trypsin digestion prior to the immunoenzyme labeling. The results indicated that even minute deposits of C3 could be detected in paraffin sections by the ABC method, which was more sensitive than the PAP technique; the ABC method allowed a maximal dilution of 1:2,400 of the primary antibody as compared to 1:800 for the PAP technique.
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PMID:Conditions for the immunohistochemical demonstration of complement factor C3 in formaldehyde-fixed and paraffin-embedded renal tissues. 242 Jul 64

The influence of antibody absorption procedures and proteolytic pre-treatment of formaldehyde-fixed placental tissue on the localization of pregnancy-associated plasma protein A by immunoperoxidase technique was examined. Apparently monospecific IgG fraction of the anti-plasma protein applied directly on fixed tissue resulted in staining of connective tissue and a thin apical rim of the syncytiotrophoblast. Further absorption of the antibody with foetal connective tissue abolished this staining reaction. Pre-treatment of the fixed placental tissue with trypsin prior to application of the antibody, which had been absorbed with connective tissue, resulted in staining within the cytoplasm of the syncytiotrophoblast exclusively. Identical staining was seen when this IgG preparation was used directly on frozen placental tissue. The results point to the importance of the specificity of the antibody preparations and of proteolytic unmasking of epitopes when fixed tissues are used for localization studies of pregnancy-associated plasma protein A by immunoperoxidase technique.
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PMID:Immunohistochemical aspects of immunological cross-reaction and masking of epitopes for localization studies on pregnancy-associated plasma protein A. 242 24

Sections of formaldehyde-fixed paraffin-embedded cortical and hippocampal brain tissue from five cases with senile dementia of Alzheimer type (SDAT) and five cases with Pick's disease (PD) were immunostained with the monoclonal antibodies (mabs) 147, RT 97, BF 10 and 8D8 with and without pretreatment with alkaline phosphatase (AP) or trypsin (Tr). The mabs 147, RT 97 and BF 10 had previously been demonstrated to bind exclusively to phosphorylated epitopes of neurofilament proteins, while mab 8D8 is shown in this report to bind mainly, but not exclusively, to phosphorylated neurofilament epitopes. The mabs RT 97, BF 10 and 8D8, but not 147 stain most, if not all, Pick bodies (PB) and Alzheimer neurofibrillary tangles (NFT). When sections are pretreated with AP or Tr the immunostaining with mab BF 10 is very resistent in both PB and NFT. This resistance of PB and NFT is in contrast to the reduced staining of axons and of swollen cells in PD by the same enzymatic pretreatment. Immunostaining with mab RT 97 of PB and NFT is reduced moderately by AP and considerably by Tr. Only when stained with mab 8D8 is there a discrepancy between PB and NFT in their reaction to the pretreatment with AP: NFT staining with mab 8D8 is not affected, while that of PB is abolished. Thus, in spite of their different ultrastructure, PB and NFT are very similar immunocytochemically and in the accessibility of their phosphorylated epitopes to enzymatic treatment.
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PMID:Alzheimer dementia and Pick's disease: neurofibrillary tangles and Pick bodies are associated with identical phosphorylated neurofilament epitopes. 244 59

We describe an immunohistochemical method using a monoclonal antibody to localize estrogen receptors (ER) in formalin-fixed, paraffin-embedded tissue. The avidin-biotin-peroxidase complex method was used, preceded by trypsin treatment to expose antigenic sites. In 111 breast cancer specimens studied simultaneously by a dextran-coated charcoal (DCC) assay and the paraffin section method, agreement on receptor status was found in 101 (91%) specimens. Quantitative staining features showed a high degree of correlation with the results of the steroid binding assay (r = 0.81). Studies on the influence of fixation on ER localization done in rabbit uteri showed that fixatives mainly composed of coagulating reagents (Carnoy's, Zenker's, Bouin's, Lilly's AAF, Helly's, ethanol) precluded ER staining, whereas cross-linking fixatives (formaldehyde, glutaraldehyde) preserved antigenic sites, although the immunoreactivity of the receptor was somewhat decreased. Studies on the effect of enzyme preincubation showed this to increase antigenic expression of ER in formaldehyde-fixed breast tumors and in formaldehyde-, glutaraldehyde-, and Zamboni-fixed rabbit uteri.
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PMID:Immunohistochemical demonstration of estrogen receptors (ER) in formalin-fixed, paraffin-embedded human breast cancer tissue by use of a monoclonal antibody to ER. 246 14


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