Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

RPMI 8866 lymphoblastoid cells, known to express surface Fc epsilon R, were tested for their ability to regulate the in vitro synthesis of human IgE. Cell-free supernatants (CFS) of RPMI 8866 cells enhanced in a dose-dependent fashion the spontaneous IgE synthesis by B cells of allergic individuals. For maximum activity the CFS had to be added during the first 3 days of culture. CFS did not significantly alter the spontaneous synthesis of IgM or IgG, but they suppressed IgA synthesis both in B cell cultures and in pokeweed mitogen-stimulated peripheral blood mononuclear cells cultures. Cyclosporin A did not suppress either the spontaneous Ig production by B cells nor the IgE-potentiating activity of CFS. The enhancing activity of CFS was related to its content in IgE binding factors (IgE-BFs); these factors were detected by their ability to inhibit the rosetting of RPMI 8866 cells with IgE-coated erythrocytes (E-IgE). Both the IgE-BFs and the IgE-potentiating activity of the supernatants of RPMI 8866 cell cultures could be removed by absorption with IgE-Sepharose, from which they could subsequently be eluted with glycine-HCl buffer. IgE-BFs were identified as glycoproteins on the basis of their sensitivity to trypsin and to neuraminidase. By filtration of the RPMI 8866 cell supernatants through a Sephadex G75 column, IgE-binding activity was found to be associated with two fractions with molecular sizes in the range of 10,000-15,000 and 30,000-40,000. The IgA-suppressing activity of the RPMI 8866 culture filtrates could be absorbed with sIgA-Sepharose from which it was subsequently recovered by elution with glycine-HCl buffer. Most unexpectedly, sIgA-Sepharose also removed IgE-BFs and IgE-potentiating activity from the RPMI 8866 supernatants; both could be recovered by subsequent elution from sIgA-Sepharose with gycline-HCl buffer. These data are provisionally interpreted as indicating that the IgE-BFs secreted by RPMI 8866 cells had affinity for both IgE and sIgA and that they exerted a reciprocal effect on the in vitro synthesis of IgE and IgA.
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PMID:In vitro synthesis of IgE by human lymphocytes. II. Enhancement of the spontaneous IgE synthesis by IgE-binding factors secreted by RPMI 8866 lymphoblastoid B cells. 623 79

Eight cross-linking fixatives were tested for preservation of extracellular or intracellular IgG, IgA, IgM, IgD, kappa and lambda light chains, J chain and secretory component. Most of the selected fixatives have been used in recent immunohistochemical studies of lymphoproliferative processes and comprised routine formalin, glutaraldehyde(1%)-formalin, Baker's formalin-calcium, formalin-sublimate, acetic acid(2%)-formalin-saline, Bouin's fluid, Susa fixative, and carbodiimide. The results obtained in artificial test substrates with defined amounts of IgG or IgA and in biological substrates (colon mucosa, tonsils, and different types of lymphomas) were compared by immunofluorescence with the antigenic preservation afforded by fixation in cold 96% ethanol (with or without inclusion of a pre-fixation 48 h washing period). An antigen concentration at least an eight-fold higher was necessary for detection with most other fixatives. Bouin's and Susa fixatives were peculiar in that they required antigen concentration 150 times higher for detection of IgG but only 3-8 times higher for IgA. Light chains were relatively well preserved by all fixatives except glutaraldehyde. For all cross-linking fixatives, the extent of antigenic masking depended on the concentration of environmental proteins, and the efficiency of unmasking with pronase or trypsin, therefore, varied with the location in the tissue. The J chain was particularly vulnerable to degradation during proteolytic treatment. The extensive masking of extracellular immunoglobulin in formalin-fixed tissue afforded a relatively good signal-to-noise ratio for immunoglobulin-producing cells when kappa and lambda chains were traced. Thus, differentiation between polyclonal and monoclonal B-cell processes on the basis of cytoplasmic labelling was often better in undigested sections. However, the light-chain type of membrane immunoglobulin could usually not be determined in directly fixed tissue. Ethanol fixation preceded by washing in saline afforded such determination and also preserved certain T-cell and HLA-DR antigens as well as diffuse alpha-naphthylbutyrate esterase. Reactive and malignant macrophages could further be traced by their cytoplasmic expression of L1 antigen, both in formalin- and ethanol-fixed material.
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PMID:Evaluation of tissue preparation methods and paired immunofluorescence staining for immunocytochemistry of lymphomas. 635 Feb 34

Different immunohistological methods were compared in 30 renal biopsy cases. The direct immunoperoxidase method gave identical results with immunofluorescence in 88% concerning IgG and C3, while IgA, IgM and fibrinogen have been identically detected by the PAP method in 82%. The non-identical results of the direct method consisted, with one exception, of negative readings, while the non-identical PAP readings were positive with two exceptions. Thus the trypsin-immunoperoxidase method proved to be well applicable on paraffin embedded formalin fixed material. In 10 cases the HRP-labelled direct method was applied also to fresh, frozen sections. It gave almost entirely identical results to immunofluorescence.
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PMID:Application of the immunoperoxidase technique to formalin fixed, paraffin embedded kidney biopsies. 635 3

Twenty-two cases of feline glomerulonephritis were investigated for the presence of immune complexes within the glomerulus using the peroxidase-antiperoxidase (PAP) method. This method was used with formalin-fixed paraffin-wax embedded tissues which were pretreated with trypsin and with frozen sections of kidney tissue. Of a total of 25 kidney specimens examined (two cats had repeated biopsies) the composition of the deposits was 23/25 IgG, 17/25 C3, 11/25 IgM and 2/25 IgA. Serial studies of two cats showed a progression of the disease from initial nephrotic syndrome to chronic renal failure. With the more severe form of the disease there was a tendency for the deposition of complement and more than one class of immunoglobulin within the glomeruli.
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PMID:An immunohistological study of feline glomerulonephritis using the peroxidase-antiperoxidase method. 638 92

A technique has been devised to isolate the thin papillary part of the dermis from punch biopsies. Papillary dermis has been treated with various proteolytic enzymes in order to release or solubilize the granular IgA deposits from the papillary dermis of patients with dermatitis herpetiformis. Incubation of the thin skin preparations with pepsin caused a disappearance of the specific fluorescence with antibodies to human IgA. After peptic digestion small amounts of IgA could be detected in the supernatants. There was some evidence that this amount was larger for preparations from patients with dermatitis herpetiformis than from controls. Corresponding procedures with trypsin, collagenase, or elastase had no detectable effect on the IgA deposits. The experiments with elastase seemed to give support for previous reports on association between the granular IgA deposits and the microfibrils of elastic fibers.
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PMID:Dermatitis herpetiformis: preparation of papillary dermis and the effect of proteolytic enzymes on the IgA deposits. 639 32

The immunohistochemical detection of J chain in histological sections of tonsils was compared to that of immunoglobulin IgG, IgM, and IgA. Pretreatment of tissue sections with detergents (saponin and Triton X-100) and with proteolytic enzyme (trypsin) at different experimental conditions has shown (i) 0.1% trypsin digestion for 1 hr at room temperature increased dramatically both the number of J-chain-positive cells detected, and the intensity of the color reaction, while the detergents had no noticeable effect; (ii) under the same conditions, trypsin also improved the detection of immunoglobulins, but quantitatively and qualitatively far less effectively than for J chain. The effect of trypsin occurred after 30 min. These results indicate that in tissue sections, the antigenic determinants of J chain are "masked" in the molecule of immunoglobulin in such a manner that the use of trypsin hydrolysis may be important for its successful detection at the light microscopic level. In the case of intact immunoglobulins, this is less important.
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PMID:The immunohistochemical detection of J chain in lymphoid cells in tissue sections: the necessity of trypsin digestion. 640 64

Schistosoma haematobium eggs were recovered in urine from schoolchildren in Mwachinga Village, Kwale District, Kenya. The surface of the eggs showed a pattern similar to that observed in a circumoval precipitin (COP) test. Observed precipitates were removed by treatment of the eggs with pepsin, but were not affected by trypsin. Similar precipitates occurred after incubating the pepsin-treated eggs with urine supernatants from the same children. Attempts were made to identify the components that might contribute to observed bleb (precipitate) formation. The fluorescent antibody test revealed both IgG and IgM in the precipitate, while urine examination by immunodiffusion revealed IgG, IgM, IgA and C3. These components probably participated in the bleb formation. This possibility seemed more likely because of the presence of anti-egg antibodies in the immunoglobulins identified in the urine, and agrees well with the expected activity of pepsin on these immunoglobulins. These results, therefore, suggest that S. haematobium eggs recovered in urine contain precipitates formed mainly by immunoglobulins; it is necessary to treat such eggs with pepsin before carrying out a COP test. The relevance of these findings to the immunodiagnosis of Schistosomiasis haematobium in man is discussed.
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PMID:Precipitates found around Schistosoma haematobium eggs from human urine prior to circumoval precipitin test. 644 51

Antinucleolar antibody (ANoAb) was tested for in sera from 25 patients with systemic lupus erythematosus (SLE), 61 with progressive systemic sclerosis (PSS), 22 with chronic active hepatitis (CAH) and 28 healthy persons, using immunofluorescence reactivity with acetone-fixed monolayers of cultured human fibroblasts, and a procedure to reveal ANoAb when other antinuclear antibodies were concurrently present. ANoAb was found on direct testing in sera from 6 patients with SLE, 15 with PSS and 7 with CAH, but not in any of 28 sera from healthy persons; homogeneously reactive antinuclear antibody was also present in the serum of these 6 cases of SLE, in 6 of the 15 with PSS and in 3 of the 7 with CAH and, in SLE specifically, pre-treatment of fibroblast monolayers with DNase "unmasked" the presence of ANoAb in a further 7 sera which had shown only homogeneous nuclear staining in fibroblasts. ANoAb belonged to the IgG, IgM and IgA class in sera from cases of SLE and PSS, and to only the IgG and IgM class in sera from cases of CAH. ANoAb titres were highest in patients with PSS. ANoAb were sensitive to RNase in 5 cases, to RNase and DNase in 6, and were sensitive to combinations of RNase, DNase, NaC1, and trypsin in the remaining cases. We conclude that (i) fibroblast monolayers are a suitable substrate for the demonstration of ANoAb, (ii) homogeneous staining of cell nuclei may mask ANoAb, so that the incidence of ANoAb becomes higher in SLE than in PSS, (iii) low-titre ANoAb in CAH not visualized in frozen tissue sections may be detected on fibroblast monolayers, and (iv) nucleolar antigens probably include RNA, RNA bound to DNA, and RNA bound to proteins.
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PMID:Antinucleolar autoantibodies demonstrated by monolayers of human fibroblasts in sera from patients with systemic lupus erythematosus, progressive systemic sclerosis and chronic active hepatitis. 674 43

The unlabeled antibody peroxidase-antiperoxidase (PAP) technique was used to demonstrate IgA in paraffin sections of human tonsil fixed in eight commonly known fixatives. In all but one of the fixatives, treatment with trypsin prior to immunostaining resulted either in digestion of sections, or no change in positive staining. In tissues fixed in neutral buffered formalin-saline, positive staining was absent unless the sections had been treated with trypsin, and it has been shown that digestion requirements of different areas throughout a section may vary according to the effectiveness of fixation.
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PMID:Influence of fixatives on the immunoreactivity of paraffin sections. 675 4

This study explores the use of immunofluorescence on routine formalin-fixed paraffin-embedded tissue to distinguish monoclonal from polyclonal lymphoplasmacytic proliferations. Sixteen tissues containing plasma cell or lymphocyte and plasma cell proliferations were studied. Five-micron sections were deparaffinized in xylene, rehydrated, and treated with 0.1% trypsin for two hours. After washing with phosphate-buffered saline, separate sections were stained with fluorescein-conjugated antibody to IgG, IgA, IgM, IgD, and IgE, and a single section was stained with both fluorescein-conjugated anti-kappa and rhodamine-conjugated antilambda. The latter section was useful to distinguish nonspecific adsorption of the fluorochromes. Where possible, results were correlated with immunoelectrophoretic studies of serum and urine. Eleven specimens with monoclonal and four specimens with polyclonal lymphoplasmacytic proliferations were readily identified, including a case of giant lymph node hyperplasia with a monoclonal IgDK plasma cell component (confirmed by specific absorption studies). Identification of monoclonality by immunofluorescence preceded immunoelectrophoretic identification in one case. One other case gave equivocal results by fluorescence. Further, the method worked on formalin-fixed decalcified tissues, although a somewhat heavier background staining was noted. This method offers a simple, reliable technique to establish or confirm the diagnosis of monoclonal lymphoplasmacytic lesions.
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PMID:Demonstration of monoclonal lymphoplasmacytic proliferations by immunofluorescence on routine formalin-fixed, paraffin-embedded tissue. 680 84


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