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3.
Medchemcomm ; 10(2): 263-267, 2019 Feb 01.
Article in English | MEDLINE | ID: mdl-30881613

ABSTRACT

A fragment library of electrophilic small heterocycles was characterized through cysteine-reactivity and aqueous stability tests that suggested their potential as covalent warheads. The analysis of theoretical and experimental descriptors revealed correlations between the electronic properties of the heterocyclic cores and their reactivity against GSH that are helpful in identifying suitable fragments for cysteines with specific nucleophilicity. The most important advantage of these fragments is that they show only minimal structural differences from non-electrophilic counterparts. Therefore, they could be used effectively in the design of targeted covalent inhibitors with minimal influence on key non-covalent interactions.

4.
Clin Exp Immunol ; 192(3): 302-314, 2018 06.
Article in English | MEDLINE | ID: mdl-29431868

ABSTRACT

The tyrosine kinase zeta chain-associated protein of 70 kDa (ZAP-70) plays a key role in T cell development and signalling. In the absence of ZAP-70, T cell development is arrested in the CD4+ CD8+ double-positive stage, thus ZAP-70 homozygous knockout (ZAP-70-/- ) mice have no mature T cells in their peripheral lymphoid organs and blood, causing severe immunodeficiency. We investigated the early kinetics and long-term effects of wild-type thymocyte transfer on T cell repopulation in ZAP-70-/- mice. We used a single intraperitoneal (i.p.) injection to deliver donor thymocytes to the recipients. Here, we show that after i.p. injection donor thymocytes leave the peritoneum through milky spots in the omentum and home to the thymus, where donor-originated CD4- CD8- double-negative thymocytes most probably restore T cell development and the disrupted thymic architecture. Subsequently, newly developed, donor-originated, single-positive αß T cells appear in peripheral lymphoid organs, where they form organized T cell zones. The established chimerism was found to be stable, as donor-originated cells were present in transferred ZAP-70-/- mice as late as 8 months after i.p. injection. We demonstrate that a simple i.p. injection of ZAP-70+/+ thymocytes is a feasible method for the long-term reconstitution of T cell development in ZAP-70-deficient mice.


Subject(s)
Adoptive Transfer/methods , Immunologic Deficiency Syndromes/therapy , Severe Combined Immunodeficiency/immunology , T-Lymphocytes/cytology , Thymocytes/transplantation , ZAP-70 Protein-Tyrosine Kinase/deficiency , Animals , Immunologic Deficiency Syndromes/genetics , Immunologic Deficiency Syndromes/immunology , Lymphocyte Count , Mice , Mice, Knockout , Severe Combined Immunodeficiency/genetics , T-Lymphocytes/immunology , ZAP-70 Protein-Tyrosine Kinase/genetics , ZAP-70 Protein-Tyrosine Kinase/immunology
5.
J Eur Acad Dermatol Venereol ; 31(9): 1453-1461, 2017 Sep.
Article in English | MEDLINE | ID: mdl-28467693

ABSTRACT

BACKGROUND: The hormone sensitivity of melanoma and the role of 'classical' oestrogen receptor (ER) α and ß in tumour progression have been intensively studied with rather contradictory results. The presence of 'non-classical' G protein-coupled oestrogen receptor (GPER) has not been investigated on human melanoma tissues. OBJECTIVE: To analyse the expression of GPER, ERα and ERß in pregnancy-associated (PAM) and in non-pregnancy-associated (NPAM) melanomas in correlation with traditional prognostic markers and disease-free survival (DFS). METHODS: Receptor protein levels were tested using immunohistochemistry in 81 formalin-fixed paraffin-embedded melanoma tissues. PAMs (n = 38) were compared with age- and Breslow thickness-matched cases (n = 43) including non-pregnant women (NPAM-W) (n = 22) and men (NPAM-M) (n = 21). The association between receptor expression and DFS was analysed by uni- and multivariate Cox proportional hazards regression. RESULTS: G protein-coupled oestrogen receptor was detected both in PAMs and NPAMs. In 39 of the 41 (95.1%) GPER-positive melanomas, GPER and ERß were co-expressed. GPER/ERß-positive melanomas were significantly more common in PAM compared to NPAM (P = 0.0001) with no significant difference between genders (P = 0.4383). In PAMs, the distribution of GPER and ERß was similar (78.4% vs. 81.6%; P = 0.8504), while in NPAM, ERß was the representative ER (60.5% vs. 27.9%; P = 0.0010) without gender difference (59.1% vs. 61.9%). GPER-/ERß-positive melanomas were associated with lower Breslow thickness, lower mitotic rate and higher presence of peritumoral lymphocyte infiltration (PLI) compared to GPER-/ERß-negative cases (P = 0.0156, P = 0.0036 and P = 0.0001) predicting a better DFS (HR = 0.785, 95% CI 0.582-1.058). Despite the significantly higher frequency of GPER and ERß expression in PAM, no significant difference was found in DFS between PAM and NPAM. All but one case failed to show ERα expression. CONCLUSIONS: The presence of GPER and its simultaneous expression with ERß can serve as a new prognostic indicator in a significant subpopulation of melanoma patients.


Subject(s)
Melanoma/metabolism , Pregnancy Complications, Neoplastic/metabolism , Receptors, Estrogen/metabolism , Receptors, G-Protein-Coupled/metabolism , Skin Neoplasms/metabolism , Adult , Disease-Free Survival , Female , Humans , Immunohistochemistry , Melanoma/complications , Pregnancy , Skin Neoplasms/complications
6.
Bull Environ Contam Toxicol ; 95(4): 452-8, 2015 Oct.
Article in English | MEDLINE | ID: mdl-26215460

ABSTRACT

To evaluate copper uptake and its toxicity on bioenergy grass giant reed (Arundo donax L.), experiments were carried out using two epigenetic clonal lines - American (BL) and Hungarian (20SZ) ecotypes - grown on elevated Cu concentrations up to 26.8 mg L(-1). Neither ecotype showed any noticeable foliar symptoms of Cu toxicity at concentrations tested up to 10 mg L(-1). Dry mass of plants of both ecotypes significantly increased at the highest Cu treatment compared to control. Although the BL ecotype had greater capacity to uptake Cu than 20SZ, the dry mass and shoot length of BL was higher than that of 20SZ. Values of bioconcentration and transportation factors were higher in the BL than in the 20SZ ecotype. Almost 45 % of total Cu content within the whole plant was found in the plant root of both ecotypes. This demonstrated both ecotypes can be utilized for Cu phytoremediation alongside with significant biomass production.


Subject(s)
Copper/toxicity , Poaceae/drug effects , Biodegradation, Environmental , Biomass , Copper/metabolism , Plant Roots/drug effects , Plant Roots/metabolism , Poaceae/metabolism
7.
Environ Sci Pollut Res Int ; 21(12): 7773-80, 2014 Jun.
Article in English | MEDLINE | ID: mdl-24638838

ABSTRACT

An in vitro experiment was carried out to evaluate the phytoremediation potentials of two somatic embryo-derived ecotypes of Arundo donax-BL (American ecotype) and 20SZ (Hungarian ecotype)-of copper from synthetic wastewater. The two ecotypes were grown under sterile conditions in tubes containing a nutrient solution supplied with increasing doses of Cu (0, 1, 2, 3, 5, 10, and 26.8 mg L(-1)) for 6 weeks. The translocation and bioaccumulation factors and removal rate were estimated. In general, increasing Cu concentration in nutrient solution slightly decreased root, stem and leaf biomass without toxicity symptoms up to 26.8 mg L(-1). Moreover, both ecotypes showed high Cu removal efficiency from aqueous solution. However, Cu removal rate ranged between 96.6 to 98.8% for BL ecotype and 97 to 100% for 20SZ ecotype. Data illustrated that both BL and 20SZ ecotypes may be employed to treat Cu-contaminated water bodies up to 26.8 mg L(-1).


Subject(s)
Biodegradation, Environmental , Copper/metabolism , Poaceae/metabolism , Wastewater/chemistry , Water Pollutants/metabolism , Biomass , Ecotype , In Vitro Techniques , Plant Roots/metabolism , Random Allocation , Seedlings/metabolism , Water Pollutants/chemistry
8.
Pathol Oncol Res ; 19(2): 141-8, 2013 Apr.
Article in English | MEDLINE | ID: mdl-23274761

ABSTRACT

Transforming growth factor ß (TGF-ß) superfamily consists of numerous cytokins that regulate various cellular processes. TGF-ß, the prototype of the family, signals through its cell surface serine/threonin kinase receptors and besides its role in cell differentiation, migration, adhesion etc. it is also able to induce epithelial-mesenchymal (EMT) transition via both Smad- pathway and MAPK- pathway. Among the different types of epithelial-mesenchymal transition, type II that is described to be associated with wound healing, tissue regeneration, organ fibrosis and is induced upon inflammatory stimuli. It can be triggered by secretion of growth factors such as TGF-ß, EGF. Different endocytic routes are used for the internalization of TGF-ß ligand and its receptors and these pathways can control the activity of downstream events. Internalization via clathrin-coated vesicles promotes the signaling while the caveola-mediated endocytosis plays important role in the termination of the events, although the steps of the latter event are less clear. The early endosome is considered a clue compartment in promoting the signaling. Recently published data suggest that the early endosome plays crucial role in the termination of the TGFß signaling as well. It is not only maintain a special environment for the effective signaling but can direct the internalized cargos towards degradative pathways (multivesicular bodies, lysosomes).


Subject(s)
Transforming Growth Factor beta/metabolism , Endocytosis , Signal Transduction
9.
Commun Agric Appl Biol Sci ; 70(4): 693-6, 2005.
Article in English | MEDLINE | ID: mdl-16628905

ABSTRACT

Main method to control the American Corn Rootworm is crop rotation (Camprag et. al., 1994) but we don't know how to determine the possible number of larvae under fall so we cannot use autumn cereals to change the row of cultivated plants. The pest spends almost 10 months in soil in egg and larval state (Chiang, 1973). There are two methods for scouting Diabrotica eggs and larval instars from soil over the winter. One of the two most important methods is holding soil samples on fixed temperature (Fromm et al., 1999). This method takes more than one and a half month but its result is highly reliable. The conventional egg-washing technique takes fewer days to count the number of Diabrotica eggs in soil but it has lower effectiveness than the other one because the eggs in a sample cannot be counted correctly. Our results show that the effectiveness of egg washing with high concentrated salty water (NaCl) is high and the method is quick enough to help planning the crop rotation even under the autumn period (Takács et al., 2004).


Subject(s)
Coleoptera/growth & development , Soil/parasitology , Agriculture/methods , Animals , Female , Larva/growth & development , Male , Population Density , Temperature , Zea mays/parasitology
10.
Commun Agric Appl Biol Sci ; 69(3): 111-7, 2004.
Article in English | MEDLINE | ID: mdl-15759402

ABSTRACT

Larvae of WCR are feeding on the roots of corn while plants fall down. The egg hatching is continuous and soil insecticides are not effective to kill larvae. Unfortunately the recent control methods while we incorporate disinfectors Into the soil under seeding are not able to give enough effect on larvae of WCR under the whole period of larval development. We use to saw corn in the middle of April but eggs hatching start in the middle of May. The effectiveness of insecticides takes about one month so they are not able to protect plants from larvae are feeding on roots (Luckman et al., 1974 and Luckmann et al., 1975). They cause yield losses or in case of plant fall we can not harvest the corn. We have tested a material in greenhouse screening and field trips that is able to absorb insecticides and bind them into its body. This material is able to emit the agents continuously under the vegetation and we can protect our plants against the damages of WCR larvae. Our results shows that the material is able to elongate the effectiveness of the pesticides over 60 days and able to push the number of larvae under the economical threshold.


Subject(s)
Coleoptera/growth & development , Insect Control/methods , Insecticides , Larva/drug effects , Animals , Plant Roots/parasitology , Zea mays/parasitology
11.
Commun Agric Appl Biol Sci ; 69(3): 305-10, 2004.
Article in English | MEDLINE | ID: mdl-15759428

ABSTRACT

The cotton bollworm (Helicoverpa armigera Hbn.) is a poliphagous pest. Caterpillars feed on flowers, crops and seeds. In 2001 the meaningful catching-period was in August (Szeoke, 2001). In 2003 we detected the swarming already in June. We observed many caterpillars on its nutritive crops. It caused significant economic damage in this year. 1ST EXAMINATION: We collected larvae and reared pupae out of it in a pot. We took it into the soil. The swarming of the moths from the pots was in June. The mortality was high, more than 90%. 2ND EXAMINATION: We made cold tests with pupae. We examined 5 x 10 pupae in three treatments. In the first treatment we reduced the temperature to -2 degrees C for 4 weeks. 92% of the pupae survived this cold. In the second treatment we reduced the temperature to -2 degrees C for 3 weeks and to -7 degrees C for 1 week. 86% of the pupae survived this procedure. In the third treatment we reduced the temperature to -2 degreesbC for 3 weeks and -15 degrees C for 1 week. 100% of the pupae were perished. 3RD EXAMINATION: In the first treatment we raised caterpillars on 13 hours lighting and 24 degrees C. The swarming was from 20th April to 4th May 2004. In the second treatment we reared the worms on 20 hours lighting and 18 degrees C. The main swarming was on 3rd January 2004. So we could say that the cotton bollworm has diapause. The more effective factor of the diapause is the length of the lighting.


Subject(s)
Gossypium/parasitology , Animals , Ecosystem , Hungary , Larva , Lepidoptera/growth & development , Population Density , Pupa , Seasons
12.
Cell Immunol ; 212(2): 126-37, 2001 Sep 15.
Article in English | MEDLINE | ID: mdl-11748929

ABSTRACT

In this paper we report the development of the sinus network of mouse spleen during the first postnatal month as studied with a set of new rat monoclonal antibodies (mAbs) against mouse splenic endothelial cell subpopulations. One of the new mAbs (IBL-7/1) also stained B-cell lineage cells in the spleen shortly after the birth as confirmed by three-color flow cytometry. This B-cell staining in the primordial follicles vanished by the fourth postnatal week, so that the expression of IBL-7/1 antigen was restricted to the marginal sinus endothelium and some red pulp sinuses and a minor B-cell subset in the spleen, presumably distinct from the follicular B-cell compartment. The other mAb (IBL-9/2) selectively labeled the sinusoids of the deeper part of the red pulp, without any reactivity against hemopoietic cells. The IBL-9/2-reactive cells in newborns appeared as isolated elements throughout spleen, and during the segregation of white and red pulps they formed an extensive network in the red pulp outside the marginal zone. Double-labeling immunofluorescence revealed that most of these sinusoids also stained weakly with IBL-7/1 mAb, whereas the strongly IBL-7/1-positive vessels of this region were IBL-9/2 negative. Neither of these mAbs reacted with the central artery. The comparative phenotypic analysis of the various vascular segments indicates that the splenic sinusoids of the marginal zone and red pulp, respectively, are lined with a heterogeneous array of endothelium. For the precise identification, isolation, and characterization of the possible homing function of these endothelium subsets these region-specific mAbs may be of potential value.


Subject(s)
B-Lymphocyte Subsets/immunology , Endothelium, Vascular/cytology , Spleen/blood supply , Splenic Artery/cytology , Age Factors , Animals , Animals, Newborn , Antibodies, Monoclonal/immunology , Antibody Specificity , Bone Marrow/immunology , Cell Lineage , Endothelium, Vascular/immunology , Erythroid Precursor Cells/immunology , Female , Flow Cytometry , Immunophenotyping , Mice , Mice, Inbred BALB C , Microcirculation , Rats , Rats, Wistar , Spleen/growth & development , Spleen/ultrastructure , Splenic Artery/growth & development , Splenic Artery/immunology
13.
Cell Immunol ; 214(1): 45-53, 2001 Nov 25.
Article in English | MEDLINE | ID: mdl-11902828

ABSTRACT

Follicular dendritic cells (FDCs) represent a unique cell population of antigen trapping cells restricted to follicles within the secondary lymphoid tissues. FDCs appear to be involved in the formation of primary follicles during the ontogeny of lymphoid tissue. We sought to determine the kinetics and tissue distribution of cells in the spleen of newborn mice expressing various differentiation antigens restricted to FDCs using immunohistochemistry with monoclonal antibodies (mAb) against FDCs and in vivo immune complex binding and retention. The earliest FDC-specific marker displayed was the antigenic determinant recognized by the FDC-M1 mAb, which was detectable by Day 3 prior to follicle formation on cells located around the peripheral part of the developing white pulp. The appearance of CD21/35 (complement receptor Type 2 and 1, CR1.2) was observed at the end of the first week, revealing a focal pattern in B-cell-rich areas. In addition, at that time there were some FDC-M1-positive cells in the nonfollicular part of the periarteriolar region. The administration of anti-horseradish peroxidase antibody followed by soluble antigen HRP into 7-day-old newborn mice resulted in the trapping and retention of immune complexes onto FDCs even in the absence of Fcgamma receptors. The appearance of another FDC-specific marker, FDC-M2, was observed during the second week after birth and was restricted on the cells located in the same area as CR1.2 cells. The Fcgamma receptor Type II appeared on FDCs after the second postnatal week. The above sequence of phenotypic maturation could also be observed in newborns after lethal irradiation at Day 3. This indicates that not only mature FDCs but also their precursors are highly radioresistant, and their phenotypic maturation follows a programmed path that requires only a small number of mature B cells.


Subject(s)
Dendritic Cells/immunology , Spleen/growth & development , Spleen/immunology , Animals , Animals, Newborn , Antigen-Antibody Complex/analysis , Antigens, Differentiation/analysis , B-Lymphocytes/immunology , Cell Differentiation , Immunophenotyping , Kinetics , Mice , Mice, Inbred BALB C , Phenotype , Radiation Tolerance , Receptors, Complement 3b/metabolism , Receptors, Complement 3d/metabolism , Receptors, IgG/metabolism , Spleen/cytology , Stem Cells/physiology , Tissue Distribution
14.
Hybridoma ; 18(2): 177-82, 1999 Apr.
Article in English | MEDLINE | ID: mdl-10380017

ABSTRACT

The homing of lymphocytes into various peripheral lymphoid organs involves a complex set of interactions between the circulating lymphoid cells and the local endothelium. While the initial binding and the adhesion processes of lymphocytes leading to their homing to the lymph nodes have thoroughly been studied, relatively little is known about the lymphoid-endothelial interactions taking place in the spleen. Our aim was to isolate rat monoclonal antibodies (MAbs) against the endothelial cells of the mouse spleen. Using splenic stroma derived from irradiated mice as antigen, two new rat MAbs were isolated. The MAb designated as IBL-7/1 bound to the sinus-lining (littoral) cells in the red pulp, marginal zone, and to the T- and B-cell compartments of the white pulp, respectively. However, it did not react with the central arteriole in the periarteriolar lymphoid sheath (PALS). In contrast to this pattern, the IBL-7/22 MAb recognized a shared antigen expressed by the sinusoidal and arterial endothelium. In addition to the endothelial reactivity, the IBL-7/22 MAb also stained the reticular components of the PALS and red pulp, but not that of the follicles. In vivo labelling with fluorescein (FITC)-conjugated IBL-7/1 MAb followed by confocal microscopic analysis revealed that the antigen recognized was expressed on the luminal surface of the sinusoids. The treatment of mice with IBL-7/1 MAb did not result in the altered distribution of T and B cells. These two new MAbs may be valuable tools for the phenotypic analysis of splenic endothelium, and can be used for the identification of various endothelial cell subpopulations of the mouse spleen.


Subject(s)
Antibodies, Monoclonal/immunology , Antibody Specificity , Endothelium, Vascular/cytology , Endothelium, Vascular/immunology , Spleen/blood supply , Animals , Antigens , Arteries/immunology , B-Lymphocytes/immunology , Fluorescent Antibody Technique , Hybridomas , Mice , Mice, Inbred BALB C , Phenotype , Rats , Rats, Wistar , T-Lymphocytes/immunology
15.
Genomics ; 62(3): 406-16, 1999 Dec 15.
Article in English | MEDLINE | ID: mdl-10644438

ABSTRACT

Protein tyrosine phosphatases (PTPs) mediate the dephosphorylation of phosphotyrosine. PTPs are known to be involved in many signal transduction pathways leading to cell growth, differentiation, and oncogenic transformation. We have cloned a new family of novel protein tyrosine phosphatase-like genes, the Ptpl (protein tyrosine phosphatase-like; proline instead of catalytic arginine) gene family. This gene family is composed of at least three members, and we describe here the developmental expression pattern and chromosomal location for one of these genes, Ptpla. In situ hybridization studies revealed that Ptpla expression was first detected at embryonic day 8.5 in muscle progenitors and later in differentiated muscle types: in the developing heart, throughout the liver and lungs, and in a number of neural crest derivatives including the dorsal root and trigeminal ganglia. Postnatally Ptpla was expressed in a number of adult tissues including cardiac and skeletal muscle, liver, testis, and kidney. The early expression pattern of this gene and its persistent expression in adult tissues suggest that it may have an important role in the development, differentiation, and maintenance of a number of different tissue types. The human homologue of Ptpla (PTPLA) was cloned and shown to map to 10p13-p14.


Subject(s)
Chromosome Mapping , Cloning, Molecular , Gene Expression Regulation, Developmental , Protein Tyrosine Phosphatases/biosynthesis , Protein Tyrosine Phosphatases/genetics , Amino Acid Sequence , Animals , Base Sequence , Caenorhabditis elegans , Chick Embryo , Chromosomes, Human, Pair 10/genetics , DiGeorge Syndrome/genetics , Heart/embryology , Humans , In Situ Hybridization, Fluorescence , Liver/embryology , Liver/enzymology , Mice , Molecular Sequence Data , Multigene Family , Muscle, Skeletal/embryology , Muscle, Skeletal/enzymology , Myocardium/enzymology , Organ Specificity , Sequence Homology, Amino Acid , Sheep
16.
J Immunol Methods ; 218(1-2): 117-21, 1998 Sep 01.
Article in English | MEDLINE | ID: mdl-9819128

ABSTRACT

In lethally irradiated mice, mixed lymphohaemopoietic chimerism can be established after their reconstitution with adult or embryonic rat haemopoietic stem cells. In this report we describe a simple fluorescent staining protocol for the determination of origin and type of various leukocyte cell subpopulations (rat or mouse, and rat T, B and myeloid cells, respectively) using whole blood samples from such animals. These data were comparable to those obtained in peripheral lymphoid tissues. Exploiting the slightly heterogeneous reactivity of the mouse monoclonal anti-rat CD45 antibody (MRC OX-1) revealed by the application of a PE-labelled monoclonal anti-mouse IgG1 secondary reagent together with a FITC-labelled rat anti-mouse CD45 mAb (IBL-5/25), we could determine the donor-derived T, B and myeloid cells from one sample in a two-step procedure. Further advantages to this easy staining procedure are that PBLs as test cells are readily available and the typing can be repeated, thus offering the opportunity for continuous monitoring of the degree of chimerism.


Subject(s)
Flow Cytometry/methods , Hematopoietic Stem Cell Transplantation , Leukocytes/cytology , Radiation Chimera , Animals , B-Lymphocyte Subsets/cytology , Hematopoietic Stem Cells/cytology , Liver/cytology , Mice , Mice, Inbred BALB C , Rats , T-Lymphocyte Subsets/cytology
17.
Dev Immunol ; 6(3-4): 179-85, 1998.
Article in English | MEDLINE | ID: mdl-9814591

ABSTRACT

The development of B cells is accompanied by their ability to specifically enter the peripheral lymphoid tissues. Recently, we described a novel rat monoclonal antibody (IBL-2; IgG2b/kappa) reacting with a 26/29-kD heterodimeric structure of the cell surface. This mAb has been found to recognize differentially the peripheral B cells of mice depending on their tissue origin. The majority of splenic B cells as well as the mature B cells in the bone marrow were stained with this mAb, whereas the B lymphocytes isolated from LN or Peyer's patches displayed only negligible reactivity. We extended these observations by analyzing the relationship between the expression of IBL-2 antigen and L-selection on the surface of B-cell precursors in the bone marrow by multiparameter flow cytometry. Within the B220 positive compartment, a significant difference of L-selectin expression could be observed between the various IBL-2-reactive subsets. Furthermore, we investigated whether evidences for the establishment of tissue-associated phenotypic heterogeneity similar to that found in normal mice could be found upon the adoptive transfer of normal unselected splenic lymphocytes into SCID recipients (Spl-SCID). It has been found that a large part of the splenic B cells preserved their IBL-2 reactivity, whereas the LN B cells had lost the IBL-2 antigen in Spl-SCID. These data indicate that the phenotypic difference within the SCID mice may be the result of the migration of B lymphocytes from the spleen toward the lymph nodes, and the altered expression of the IBL-2 antigen correlates with this process.


Subject(s)
Antigens, Surface/analysis , B-Lymphocytes/immunology , Lymph Nodes/immunology , Spleen/immunology , Adoptive Transfer , Animals , Antibodies, Monoclonal/immunology , B-Lymphocytes/metabolism , Bone Marrow Cells/immunology , Cell Differentiation , Female , Flow Cytometry , Fluorescent Antibody Technique , Hematopoietic Stem Cells/immunology , Hematopoietic Stem Cells/metabolism , L-Selectin/analysis , Mice , Mice, Inbred BALB C , Mice, SCID , Phenotype , Rats
18.
Immunology ; 86(4): 560-7, 1995 Dec.
Article in English | MEDLINE | ID: mdl-8567022

ABSTRACT

After their primary differentiation and selection in the bone marrow, the cells of B lineage are distributed to the peripheral lymphoid system. Here we report that, with the use of a novel rat monoclonal antibody (IBL-2), a tissue-related phenotypic difference could be observed in the peripheral B-cell compartment in mouse. The antigen recognized by this antibody is a 25,000/29,000 MW heterodimeric cell surface molecule which is resistant to phosphatidylinositol-phospholipase C treatment, but is sensitive to proteases. The antigen was found to be expressed by the majority of B cells from the spleen, whereas the B cells from other peripheral sources (lymph nodes and Peyer's patches) proved to be negative. The staining pattern of splenic B cells was heterogeneous, containing a substantial dim population (IBL-2lo), and a smaller, intensely stained fraction (IBL-2hi) within the positive subset. Unlike the B cells, the T cells were negative in every peripheral lymphoid tissue analysed. In addition, the ratios between the various IBL-2-reactive B cells (positive to negative and, within the positive population, the IBL-2lo to IBL-2hi, respectively) in the spleen were quite similar to that of B cells in the bone marrow. Furthermore, the levels of L-selectin expressed by the various IBL-2-reactive subpopulations were found to be heterogeneous both in the bone marrow and in the spleen. The bone marrow cells could be resolved into double negative, L-selectin +/-/IBL-2lo, L-selectin--IBL-2lo, and L-selectin-/IBL-2hi populations, respectively. In the spleen, an additional fraction with L-selectin+/IBL-2- phenotype could be detected. In both tissues, the overwhelming majority of IBL-2hi cells were found at the MEL-14- compartment. We conclude that either these findings may reflect a heterogeneous development state within the peripheral B-cell pool, with a substantial fraction of splenic B cells being less differentiated than those in other peripheral lymphoid tissues, or alternatively, the differential reactivity of murine B cells with the IBL-2 monoclonal antibody is due to their tissue location.


Subject(s)
B-Lymphocyte Subsets/immunology , Spleen/immunology , Animals , Antibodies, Monoclonal , Bone Marrow/immunology , Female , Flow Cytometry , Immunoblotting , Immunophenotyping/methods , L-Selectin/analysis , Lymphoid Tissue/immunology , Male , Mice , Mice, Inbred BALB C , Splenectomy
20.
J Immunol Methods ; 169(1): 35-40, 1994 Feb 28.
Article in English | MEDLINE | ID: mdl-7510754

ABSTRACT

A monoclonal anti-FITC antibody (F4/1) was produced and demonstrated to be specific for both the free and protein-conjugated (either soluble or cell-bound) form of fluorescein, or carboxyfluorescein. When mouse thymocytes were labelled with a novel fluorescein derivative 5(6)-carboxyfluorescein succinimidyl ester (CFl-NSE), the incorporation of fluorescein was predominantly membrane-bound as demonstrated immunohistochemically. The coupling of CFl-NSE to cells displays a random distribution pattern as shown by immunoblotting of cell extracts prepared by detergent solubilization of CFl-NSE-labelled thymocytes. In addition, the Thy-1.2 antigen immunoprecipitated from a CFl-NSE-labelled thymocyte lysate with a rat monoclonal antibody (Mab) could be detected using the anti-FITC Mab. The molecular weight of the immunoprecipitated material could be estimated immediately by reference to the FITC-labelled molecular weight markers electrophoresed simultaneously.


Subject(s)
Antibodies, Monoclonal , Antigens, Surface/analysis , Fluorescein-5-isothiocyanate , Haptens , Immunohistochemistry/methods , Thymus Gland/cytology , Animals , Cell Line , Electrophoresis, Polyacrylamide Gel , Fluorescein , Fluoresceins , Hybrid Cells , Immunoblotting , Mice , Mice, Inbred BALB C , Precipitin Tests , Rats , Staining and Labeling
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