Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 166
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Clin Nephrol ; 71(4): 387-96, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19356371

RESUMO

AIM: End-stage renal disease (ESRD) is often complicated by chronic inflammation and malnutrition. We tested whether serum tartrate-resistant acid phosphatase (TRACP) isoform 5a relates to other markers of inflammation in ESRD. MATERIAL: Predialysis serum was collected from 99 ESRD patients (51 male, 48 female) aged 55 +/- 15 years and a control group of 36 healthy subjects (8 male, 28 female) aged 43.2 +/- 10.5 years. METHODS: Serum TRACP 5a activity and protein, TRACP 5b activity and C-reactive protein (CRP) were estimated by in-house immunoassays. Commercial kits were used for serum bone-specific alkaline phosphatase, Ntelopeptides of Type I collagen, interleukin-6 (IL-6) and fetuin-A. Intact parathyroid hormone was determined by chemiluminescent assay. Albumin, cholesterol, triglycerides, ferritin and hemoglobin were compared to the hospital reference ranges. Bone mineral density (BMD) was measured at the heel in 69 patients and all control subjects and expressed as g/cm2 and age-corrected T-score. RESULTS: Mean (median) levels of all serum markers were significantly elevated in ESRD except fetuin-A, which was significantly reduced. Mean BMD (g/cm2) was not different than control, but mean T-score was significantly reduced. TRACP 5a protein correlated with CRP, triglycerides and ferritin, but not with IL-6 or any other nutritional or bone markers or BMD. TRACP 5b activity correlated with all bone markers and BMD, but not with inflammation or nutritional markers. CONCLUSION: Our findings suggest that TRACP 5a may be a useful marker to estimate the degree of inflammation in ESRD patients on chronic hemodialysis.


Assuntos
Fosfatase Ácida/sangue , Isoenzimas/sangue , Falência Renal Crônica/sangue , Adulto , Albuminas/metabolismo , Fosfatase Alcalina/sangue , Biomarcadores/sangue , Densidade Óssea , Proteína C-Reativa/metabolismo , Estudos de Casos e Controles , Colágeno Tipo I/sangue , Feminino , Humanos , Inflamação/sangue , Interleucina-6/sangue , Falência Renal Crônica/terapia , Modelos Lineares , Masculino , Pessoa de Meia-Idade , Hormônio Paratireóideo/sangue , Isoformas de Proteínas/sangue , Diálise Renal , Estatísticas não Paramétricas , Fosfatase Ácida Resistente a Tartarato , alfa-Fetoproteínas/metabolismo
2.
J Bone Miner Res ; 16(4): 788-93, 2001 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-11316008

RESUMO

Tartrate-resistant acid phosphatase (TRAP) isoform 5b is a potential serum marker for osteoclastic activity. Biochemical assays for serum TRAP activity with para-nitrophenylphosphate (pNPP) have low specificity for bone because of hydrolysis by unrelated nontype 5 TRAPs of blood cells and by related isoform 5a. Our purpose was to increase the specificity of TRAP assay for osteoclastic activity by using naphthol-ASBI phosphate (N-ASBI-P) as a substrate for serum type 5 TRAP activity and heparin as an inhibitor of isoform 5a. TRAP activity in individual and pooled sera of normal subjects and patients with endstage renal disease (ESRD) and rheumatologic diseases was quantitated using pNPP and N-ASBI-P as substrate at pH 5.5 and 6.1. For some experiments, heparin (23U/ml) was added as a specific inhibitor of isoform 5a activity. Isoforms 5a and 5b were separated from serum pools by cation exchange chromatography and identified by nondenaturing polyacrylamide gel electrophoresis (PAGE). N-ASBI-P was selectively hydrolyzed by TRAP isoform 5b. TRAP assays with pNPP and N-ASBI-P correlated only in ESRD sera, which contained primarily isoform 5b. The two assays did not correlate in normal or rheumatic sera with significant amounts of 5a. Heparin inhibited isoform 5a activity approximately 50% but had little effect on isoform 5b activity. Biochemical assay of serum TRAP activity can be made specific for isoform 5b by using N-ASBI-P and heparin. This method can be adapted to simple microplate biochemical or immunochemical assays. This simplified method for assessment of osteoclastic TRAP 5b activity warrants a detailed investigation in diseases of bone metabolism.


Assuntos
Fosfatase Ácida/metabolismo , Remodelação Óssea , Ensaios Enzimáticos Clínicos/métodos , Isoenzimas/metabolismo , Compostos Organofosforados/metabolismo , Osteólise/diagnóstico , Fosfatase Ácida/antagonistas & inibidores , Fosfatase Ácida/isolamento & purificação , Compostos de Anilina/metabolismo , Artrite Reumatoide/enzimologia , Biomarcadores , Cromatografia por Troca Iônica , Eletroforese em Gel de Poliacrilamida , Inibidores Enzimáticos/farmacologia , Heparina/farmacologia , Humanos , Concentração de Íons de Hidrogênio , Hidrólise , Isoenzimas/antagonistas & inibidores , Isoenzimas/isolamento & purificação , Falência Renal Crônica/enzimologia , Osteoclastos/metabolismo , Osteólise/sangue , Osteólise/enzimologia , Sensibilidade e Especificidade , Especificidade por Substrato , Fosfatase Ácida Resistente a Tartarato
3.
Clin Chem ; 47(1): 74-80, 2001 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-11148180

RESUMO

BACKGROUND: Tartrate-resistant acid phosphatase (AcP) 5b is a marker of osteoclastic activity and bone resorption. Immunoassays for serum TRAcP may lack sensitivity and specificity because of the presence of non-bone isoform 5a. The purpose of this study was to isolate the serum isoforms, quantify their disease-related expressions, and test an improved immunoassay for TRAcP 5b. METHODS: We separated TRAcP isoforms chromatographically from pooled sera of healthy, rheumatoid arthritis (RA) and endstage renal disease (ESRD) subjects. TRAcP isoforms were identified by electrophoresis and quantified by biochemical and immunochemical assays. Serum TRAcP activity in healthy, RA, and ESRD cohorts was assessed at pH 5.5 and 6.1, and compared with bone alkaline phosphatase (BAP) and N-telopeptides of type I collagen (NTx). RESULTS: TRAcP isoforms 5a and 5b were present in all sera; 5b was identical to osteoclastic TRAcP. In serum from healthy subjects, 5a accounted for 87% of the enzyme protein but only 55% of the activity. In RA, both isoforms were increased two- to threefold in protein, but their specific activities were subnormal. In ESRD, only 5b was abnormal, being increased fivefold in protein and threefold in activity. In RA sera, TRAcP activity did not correlate with either BAP or NTx. In ESRD sera, TRAcP activity correlated with BAP and NTx only when measured at pH 6.1. CONCLUSIONS: All sera contained both TRAcP isoforms 5a and 5b, but only 5b was present in bone. TRAcP isoform expression was variable in different diseases. Measurement of TRAcP activity at pH 6.1 improves the specificity of immunoassay for isoform 5b.


Assuntos
Fosfatase Ácida/metabolismo , Isoenzimas/metabolismo , Osteoclastos/enzimologia , Artrite Reumatoide/enzimologia , Biomarcadores/sangue , Humanos , Imunoensaio , Falência Renal Crônica/enzimologia , Valores de Referência , Sensibilidade e Especificidade , Fosfatase Ácida Resistente a Tartarato
4.
Clin Chim Acta ; 301(1-2): 147-58, 2000 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-11020469

RESUMO

The objective of this study was to identify the isoform, type-5a or type-5b, responsible for increased tartrate-resistant acid phosphatase (TRAP) activity in endstage renal disease (ESRD) and TRAP protein in rheumatoid arthritis (RA). We studied 24 sera each from healthy, ESRD and RA subjects. Type-5 TRAP activity and protein were quantitated by immunoassays. Isoform expression was determined by computerized imaging of non-denaturing polyacrylamide gels (PAGE) stained for TRAP activity. Other biochemical markers included: intact parathyroid hormone (iPTH), total and bone-specific alkaline phosphatase (TAP, BAP), N-telopeptides of type-I collagen (NTx), and free pyridinoline (Pyd). Isoform 5a was normal in both ESRD and RA. Isoform 5b was elevated in ESRD only. Serum TRAP activity correlated with both isoforms 5a and 5b in RA, but only with 5b in ESRD. TRAP protein assays did not correlate with PAGE assays for 5a or 5b. TRAP activity, but not protein, correlated with BAP and NTx in RA sera. Both TRAP activity and protein correlated with iPTH, TAP and Pyd in ESRD sera. Increased TRAP activity in ESRD was due to increased osteoclastic isoform 5b and related to bone turnover. Increased TRAP protein in RA was suspected, but not proven, to be isoform 5a and not related to bone turnover. Heterogeneity of serum TRAP and preferential expression of isoforms has clinical significance in different diseases including ESRD and RA.


Assuntos
Fosfatase Ácida/sangue , Artrite Reumatoide/sangue , Isoenzimas/sangue , Falência Renal Crônica/sangue , Osso e Ossos/metabolismo , Eletroforese em Gel de Poliacrilamida , Humanos , Fosfatase Ácida Resistente a Tartarato
5.
Clin Chem ; 45(12): 2150-7, 1999 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-10585347

RESUMO

BACKGROUND: Tartrate-resistant acid phosphatase (TRAP; EC 3.1.3.2) is a product of osteoclasts and a biochemical marker of bone resorption rate. However, erythrocytes and platelets contribute to total TRAP activity in serum, reducing the specificity of direct biochemical assays in serum. Osteoclast TRAP is also known as type-5 TRAP and is antigenically unique. Immunoassays are sought to improve the specificity and sensitivity of TRAP as a bone marker. METHODS: We developed two colorimetric microplate assays for type-5 TRAP: an enzyme capture immunoassay to measure antibody-bound enzymatic activity, and a two-site immunoassay to measure bound enzyme protein. Both use the same monoclonal antibody (14G6) to capture type-5 TRAP, which permits determination of specific activity of serum TRAP in health and disease. RESULTS: Both TRAP assays were linear from one-tenth to fivefold the mean value in 18 healthy subjects. In these subjects, the mean (SD) TRAP activity was 3.2 (0.54) U/L for the enzyme capture assay and 37 (13) microg/L for the two-site assay. Mean TRAP activity was not significantly increased in 64 patients with endstage renal disease requiring hemodialysis (HD) or 99 unselected patients with rheumatic diseases. By contrast, TRAP protein was increased in both the HD and rheumatic disease groups. The specific activity of TRAP in the 17 of 64 HD sera that had increased TRAP activity (0.088 U/microg) was similar to that in healthy subjects (0.091 U/microg). By contrast, the specific activity of TRAP in the 31 of 99 rheumatic sera with increased TRAP protein (0.035 U/microg) was significantly decreased. CONCLUSIONS: Wide sample distributions for TRAP activity in HD patients and TRAP protein in rheumatic disease patients suggest the presence of subpopulations of HD patients with increased TRAP activity and of rheumatic patients with increased TRAP protein. Each assay for TRAP activity and protein may have its own biological significance and clinical applications in specific groups of patients.


Assuntos
Fosfatase Ácida/análise , Imunoensaio/métodos , Isoenzimas/análise , Fosfatase Ácida/sangue , Fosfatase Ácida/imunologia , Adulto , Especificidade de Anticorpos , Reabsorção Óssea/sangue , Feminino , Peroxidase do Rábano Silvestre , Humanos , Isoenzimas/sangue , Isoenzimas/imunologia , Falência Renal Crônica/sangue , Masculino , Diálise Renal , Doenças Reumáticas/sangue , Sensibilidade e Especificidade , Fosfatase Ácida Resistente a Tartarato
6.
J Ky Med Assoc ; 97(7): 299-302, 1999 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-10412257

RESUMO

Bone marrow necrosis is most frequently diagnosed at postmortem examination. Antemortem diagnosis is still uncommon. We illustrate four cases where initial bedside attempts at needle aspiration and biopsy of primary and metastatic tumor tissue from the sternum were complicated by inadequate specimen retrieval secondary to marrow necrosis and/or tissue destruction by tumor. In these cases, CT guidance was useful in the precise localization of the bulk of the tissue mass and consequently the successful retrieval of adequate diagnostic specimens. We demonstrate CT guidance as an excellent and convenient alternative in circumstances where adequate marrow aspirations and biopsies are difficult and complicated.


Assuntos
Neoplasias da Medula Óssea/diagnóstico , Neoplasias da Medula Óssea/secundário , Medula Óssea/patologia , Tomografia Computadorizada por Raios X/métodos , Adenocarcinoma/secundário , Biópsia por Agulha/métodos , Medula Óssea/diagnóstico por imagem , Carcinoma de Células de Transição/secundário , Humanos , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/patologia , Necrose , Neoplasias da Próstata/patologia , Sensibilidade e Especificidade , Neoplasias da Bexiga Urinária/patologia
7.
J Ky Med Assoc ; 97(2): 61-5, 1999 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-10073058

RESUMO

Acute promyelocytic leukemia (APL) is a distinct subtype of acute myeloid leukemia characterized by hypergranular leukemic cells, bleeding diathesis and t(15; 17) translocation. The t(15; 17) translocation leads to the production of the PML-RAR alpha fusion protein which plays a vital role in the pathogenesis of APL by arresting normal differentiation of myeloid precursors. However, in the presence of high concentrations of all-trans-retinoic acid (ATRA), the PML-RAR alpha fusion protein serves to stimulate cell differentiation. The diagnosis of APL and the detection of residual disease are based on the t(15; 17) translocation. Treatment with a combination of ATRA and anthracycline-AraC chemotherapy has shown a higher rate of complete remission in APL. We report the case of a 71-year-old male with the rare microgranular variant of APL to illustrate these findings. The patient was treated with a combination of ATRA and Daunorubicin-AraC chemotherapy and achieved complete remission. He developed retinoic acid syndrome as a complication of therapy with ATRA. The methods for diagnosis, the molecular mechanisms in the oncogenesis of APL, rationale of treatment of APL with ATRA, complications of therapy and the new concepts in the treatment of ATRA-resistant APL are discussed.


Assuntos
Leucemia Promielocítica Aguda/diagnóstico , Idoso , Antibióticos Antineoplásicos/administração & dosagem , Antimetabólitos Antineoplásicos/administração & dosagem , Antineoplásicos/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Cromossomos Humanos Par 15/genética , Cromossomos Humanos Par 17/genética , Citarabina/administração & dosagem , Daunorrubicina/administração & dosagem , Humanos , Leucemia Promielocítica Aguda/tratamento farmacológico , Leucemia Promielocítica Aguda/genética , Masculino , Translocação Genética/genética , Tretinoína/administração & dosagem
9.
J Ky Med Assoc ; 96(1): 10-2, 1998 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-9470310

RESUMO

We present herein a patient with multiple medical illnesses, marked weight loss, and chronic anemia. Cytologic studies of his bone marrow revealed hypocellularity and gelatinous transformation (GTBM). The latter is a disorder of the hematopoietic system commonly occurring in chronically ill patients and is associated with various hematologic abnormalities. The etiology of GTBM is uncertain. Patients with this disorder may have associated medical problems but malnutrition seems to play a role. Review of the literature implies a significant associated morbidity and mortality, and a possible reversal of gelatinous changes with intense nutritional support. An otherwise dismal prognosis may be avoided by prompt diagnosis of this disorder and intensive nutritional support.


Assuntos
Doenças da Medula Óssea/patologia , Medula Óssea/patologia , Idoso , Anemia/complicações , Doenças da Medula Óssea/etiologia , Doença Crônica , Humanos , Masculino , Distúrbios Nutricionais/complicações
11.
Biotech Histochem ; 73(6): 316-24, 1998 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-9888357

RESUMO

Tartrate-resistant acid phosphatase (TRAP) is expressed abundantly by osteoclasts and is required for bone resorption. This enzyme is emerging as an important biomarker in bone pathology, both for histochemical identification of osteoclasts and as a serum marker of osteoclast activity and increased bone turnover. Rat and mouse models are becoming popular systems for studying osteoclast development, bone physiology and morphogenesis, and bone diseases such as osteoporosis. We have developed two unique antibodies to human TRAP purified from hairy cell leukemia spleen. Both antibodies (9C5 and 14G6) are suitable for immunohistochemistry of osteoclasts and macrophages. Only one (14G6) is capable of immunoprecipitating active TRAP from human cell lysates. Antibody 9C5 reacts with a denatured epitope of TRAP while antibody 14G6 probably reacts with a native, conformational determinant. The high degree of homology among TRAPs of various species predicts that these antibodies should be suitable for work in experimental animals as well as humans. Immunohistochemical staining, electrophoretic analyses, immunoprecipitation and immunoblotting assays of human rat and mouse TRAP were carried out to test the validity of these antibodies as cell markers in rodents. Both antibodies were suitable for immunohistochemistry in all species. Antibody 9C5 was suitable for immunoblotting of denatured TRAP of all species tested. Antibody 14G6 reacted with the native TRAP of humans only and failed to immunoprecipitate mouse or rat TRAP activity. Although TRAP is a phylogenetically conserved protein, subtle, species-specific determinants exist. Care should be exercised when anti-TRAP antibodies are used for immunoassay in experimental animals.


Assuntos
Fosfatase Ácida/imunologia , Anticorpos Monoclonais/imunologia , Isoenzimas/imunologia , Animais , Biomarcadores , Mapeamento de Epitopos , Humanos , Imuno-Histoquímica , Leucemia de Células Pilosas/enzimologia , Camundongos , Ratos , Especificidade da Espécie , Fosfatase Ácida Resistente a Tartarato , Células Tumorais Cultivadas
12.
South Med J ; 90(10): 1053-5, 1997 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-9347822

RESUMO

Because of the emergence of warfarin resistance in rodents, second-generation anticoagulants named "superwarfarins" were developed and marketed in over-the-counter rodenticide products. The availability of these compounds has resulted in accidental or intentional human ingestions, which cause severe bleeding. The methods for diagnosis and treatment of patients using superwarfarins are different from those for patients taking the regular warfarins. We report a case of intentional superwarfarin ingestion that caused petechiae and hematuria. Although the patient denied taking anticoagulant, the persistence of vitamin K-dependent factor deficiency led us to investigate the serum for anticoagulant rodenticides. We found high levels of brodifacoum, a superwarfarin compound. This case emphasizes the need for suspicion of superwarfarin poisoning in patients who show unexplained bleeding due to deficiency of vitamin K-dependent factors and resistance to treatment.


Assuntos
4-Hidroxicumarinas/intoxicação , Hematúria/induzido quimicamente , Rodenticidas/intoxicação , Varfarina/intoxicação , Adulto , Testes de Coagulação Sanguínea , Humanos , Masculino , Intoxicação/psicologia , Deficiência de Vitamina K/induzido quimicamente
13.
Am J Clin Pathol ; 108(3): 308-15, 1997 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-9291460

RESUMO

The demonstration of tartrate-resistant acid phosphatase (TRAP) activity has long been a cornerstone in the diagnosis of hairy cell leukemia (HCL). Recently a monoclonal antibody to this enzyme has been developed that can be used in an immunoperoxidase method on paraffin-embedded tissues. By using a peroxidase-labeled streptavidin biotin method, paraffin sections of B5 and formalin-fixed tissue from 86 cases of HCL (41 bone marrow, 36 spleen, 9 liver) were stained with the antibody to TRAP and compared against staining for CD20 (L26) and DBA.44 (DAKO, Carpinteria, Calif). In addition, 193 specimens (127 bone marrow, 42 lymph node, 19 spleen, 5 other) from a variety of neoplastic and nonneoplastic hematologic conditions were stained using the monoclonal antibody to TRAP. For comparison, these cases were also stained with DBA.44. In the cases of HCL, 80 of 86 specimens were immunoreactive for TRAP. While the antibody to TRAP generally stained less than 50% of the hairy cells, CD20 and DBA.44 stained 90% and 50% to 60% of hairy cells, respectively. Two of three cases of marginal zone lymphoma showed weak immunoreactivity to the TRAP antibody. Two specimens from a patient with Gaucher's disease and 8 of 13 cases of mastocytosis also showed positivity to the TRAP antibody in the macrophages and mast cells, respectively. In contrast, staining for DBA.44 was positive in 3 of 9 cases of B-cell large cell lymphoma, 1 of 4 cases of mantle cell lymphoma, and in the paraimmunoblasts of 1 of 7 cases of small lymphocytic lymphoma. Only HCL was TRAP and DBA.44 positive. This antibody to TRAP is a useful addition to the diagnosis of HCL but should be used in conjunction with CD20 and DBA.44. The use of this antibody to determine minimal residual disease after chemotherapy was not addressed.


Assuntos
Fosfatase Ácida/análise , Biomarcadores Tumorais/análise , Isoenzimas/análise , Leucemia de Células Pilosas/enzimologia , Transtornos Linfoproliferativos/enzimologia , Fosfatase Ácida/imunologia , Anticorpos Monoclonais/análise , Anticorpos Monoclonais/imunologia , Biomarcadores Tumorais/imunologia , Medula Óssea/enzimologia , Medula Óssea/patologia , Neoplasias da Medula Óssea/diagnóstico , Neoplasias da Medula Óssea/enzimologia , Neoplasias da Medula Óssea/patologia , Diagnóstico Diferencial , Doença de Gaucher/diagnóstico , Doença de Gaucher/enzimologia , Doença de Gaucher/patologia , Humanos , Imuno-Histoquímica/métodos , Isoenzimas/imunologia , Leucemia de Células Pilosas/diagnóstico , Leucemia de Células Pilosas/patologia , Leucemia Linfocítica Crônica de Células B/diagnóstico , Leucemia Linfocítica Crônica de Células B/enzimologia , Leucemia Linfocítica Crônica de Células B/patologia , Fígado/enzimologia , Fígado/patologia , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/enzimologia , Neoplasias Hepáticas/patologia , Linfonodos/enzimologia , Linfonodos/patologia , Linfoma de Células B/diagnóstico , Linfoma de Células B/enzimologia , Linfoma de Células B/patologia , Transtornos Linfoproliferativos/diagnóstico , Transtornos Linfoproliferativos/patologia , Macrófagos/enzimologia , Macrófagos/patologia , Mastócitos/enzimologia , Mastócitos/patologia , Inclusão em Parafina , Patologia Clínica/métodos , Baço/enzimologia , Baço/patologia , Neoplasias Esplênicas/diagnóstico , Neoplasias Esplênicas/enzimologia , Neoplasias Esplênicas/patologia , Fosfatase Ácida Resistente a Tartarato
14.
Hybridoma ; 16(2): 175-82, 1997 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-9145320

RESUMO

A major product of osteoclasts, tartrate-resistant acid phosphatase (TRAP) is an essential but insufficient enzyme for bone resorption. TRAP is an excellent cell marker for osteoclasts and macrophages and is being investigated as a serum marker for osteoclast activity in diseases of bone destruction. For decades, TRAP has also been used as a marker for hairy cell leukemia. Immunoassays for TRAP are sought to increase the sensitivity and specificity of the TRAP test for bone and hairy cells. Our laboratory recently developed a monoclonal antibody to TRAP (9C5) useful for immunohistochemical identification of TRAP-positive cells in paraffin sections. Herein, we characterize 9C5 in greater detail and report production of another anti-TRAP monoclonal antibody antibody (14G6) reactive with native, active enzyme antigen. Enzyme immunoassay, immunoprecipitation, western blot, and immunohistochemical analyses revealed the contrasting properties of 9C5 and 14G6. Antibody 9C5 reacts with a heat-denatured epitope and is suitable for denaturing western blot analysis and for immunohistochemistry. Antibody 14G6 reacts with a conformational determinant destroyed by heat and is suitable for immunoprecipitation of active TRAP, although 20% to 30% of activity is inhibited in the immune complexes. Having characterized several properties of these anti-TRAP antibodies, 9C5 and 14G6 may be useful for development of TRAP-specific immunoassays in bone pathology and hematology. They will certainly be of use for the study of biosynthesis, regulation, expression, and function of TRAP.


Assuntos
Fosfatase Ácida/imunologia , Especificidade de Anticorpos , Imuno-Histoquímica/métodos , Isoenzimas/imunologia , Anticorpos Monoclonais , Biomarcadores , Western Blotting , Reabsorção Óssea/enzimologia , Reabsorção Óssea/imunologia , Epitopos , Humanos , Hibridomas , Técnicas Imunoenzimáticas , Leucemia de Células Pilosas/enzimologia , Leucemia de Células Pilosas/imunologia , Macrófagos/imunologia , Monócitos/imunologia , Fosfatase Ácida Resistente a Tartarato
15.
J Histochem Cytochem ; 44(3): 235-44, 1996 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-8648083

RESUMO

We have developed a monoclonal antibody (9C5) for immunohistochemical localization of tartrate-resistant acid phosphatase (TRAcP). This antibody reacts with a denatured epitope of TRAcP and requires enhancement methods to promote antigenicity in paraffin-embedded tissues. We used this antibody to systematically examine proteolytic digestion and heat denaturation conditions for epitope enhancement in both paraffin sections and fixed smears. The goal was to increase the sensitivity of the immunohistochemical stain for TRAcP. Optimal conditions for proteolytic digestion were established. Denaturation in a conventional boiling water bath was compared to microwave irradiation in several commonly used solutions. Immunohistochemistry was compared directly to TRAcP cytochemistry in fixed smears from hairy cell leukemia specimens to gauge the level of sensitivity of our improved method. Attempts were made to "retrieve" the 9C5 epitope from overfixed tissues and aged smears. Maximal immunoreactivity of TRAcP was achieved by microwave irradiation in a citrate or Tris buffer of pH 6.0-8.0 without the need for a subsequent protease digestion step. With this method of epitope enhancement, immunohistochemistry with antibody 9C5 was as sensitive as direct cytochemical staining of TRAcP activity. However, once a tissue specimen had been overfixed or a smear stored for a year or more, the 9C5 epitope was no longer retrievable. The key element in epitope enhancement for 9C5 immunohistochemistry is heat denaturation of the target epitope. Immunohistochemistry of TRAcP in paraffin sections would be a great asset to the study of specialized forms of the monocyte/macrophage lineage and to the process of macrophage activation. It would also provide another means for more precise evaluation of residual disease in bone marrow of patients treated for hairy cell leukemia.


Assuntos
Fosfatase Ácida/análise , Biomarcadores Tumorais/análise , Isoenzimas/análise , Leucemia de Células Pilosas/enzimologia , Fosfatase Ácida/imunologia , Epitopos , Humanos , Imuno-Histoquímica/métodos , Isoenzimas/imunologia , Sensibilidade e Especificidade , Fosfatase Ácida Resistente a Tartarato
16.
Histochem J ; 28(3): 195-200, 1996 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-8735286

RESUMO

Tartrate-resistant acid phosphatase is an inducible marker of cell differentiation and activation expressed by specialized cells of macrophage lineage and some activated lymphocytes. Clinically, this phosphatase is a diagnostic marker for hairy cell leukaemia and osteoclast activity. The cDNA for this enzyme has been cloned from a placental expression library, yet the cell(s) expressing the enzyme protein has not been determined with certainty. Our laboratories have developed a monoclonal antibody, 9C5, suitable for immunohistochemical localization of tartrate-resistant acid phosphatase in paraffin sections. The purpose of this study was to use antibody 9C5 to identify cells expressing tartrate-resistant acid phosphatase in sections of paraffin-embedded, normal, full-term placenta and to determine if those cells expressed other macrophage markers including CD68 (PG-M1 antibody), LN5, lysozyme, alpha 1-antitrypsin and alpha 1-antichymotrypsin. Histochemical localization of activity in frozen sections was compared with immunohistochemical localization in paraffin sections of the same tissue specimens. The activity and antigenicity of this enzyme were detected in decidual cells, syncytiotrophoblast, and some macrophages distributed throughout maternal and embryonic tissues, but not in neutrophils. Unlike other tissues previously examined, placenta contains significant numbers of the phosphate-positive cells that are not of macrophage origin.


Assuntos
Fosfatase Ácida/análise , Isoenzimas/análise , Placenta/enzimologia , Anticorpos Monoclonais , Antígenos/análise , Antígenos CD/análise , Antígenos de Diferenciação Mielomonocítica/análise , Biomarcadores/análise , Diferenciação Celular/fisiologia , Feminino , Histocitoquímica , Humanos , Imuno-Histoquímica , Macrófagos/química , Macrófagos/citologia , Muramidase/análise , Placenta/citologia , Fosfatase Ácida Resistente a Tartarato , alfa 1-Antiquimotripsina/análise , alfa 1-Antitripsina/análise
17.
Am J Clin Pathol ; 104(4): 397-402, 1995 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-7572788

RESUMO

Immunohistochemical studies were done on formalin-fixed, paraffin-embedded tissues to evaluate the specificity of a newly developed monoclonal antibody (9C5) against tartrate-resistant acid phosphatase. Sections from 195 specimens were examined, which included 33 types of tissues/organs. These tissues included normal, inflammatory, and neoplastic processes. Neoplastic tissues from 14 patients with hairy cell leukemia served as positive controls. Epitope enhancement was accomplished either by microwave irradiation in citrate buffer or by boiling in water followed by trypsin digestion. Tissues were reacted with monoclonal antibody 9C5 and stained with either the avidin-biotin peroxidase method or the alkaline phosphatase anti-alkaline phosphatase method. The hairy cells of all cases of hairy cell leukemia reacted positively with 9C5. Other positively stained cells included osteoclasts, activated macrophages and giant cells. Immunohistochemical studies with 9C5, when interpreted within the context of the specificity of this antibody, are useful for the diagnosis and assessment of treatment results for hairy cell leukemia. Monoclonal antibody 9C5 also may be useful as a marker for osteoclasts and the activated macrophages and for the diagnosis of disorders involved by these cells.


Assuntos
Fosfatase Ácida/metabolismo , Isoenzimas/metabolismo , Anticorpos Monoclonais/imunologia , Especificidade de Anticorpos , Biomarcadores Tumorais , Hematopoese , Humanos , Imuno-Histoquímica/métodos , Inflamação/enzimologia , Inflamação/patologia , Células de Kupffer/metabolismo , Leucemia de Células Pilosas/enzimologia , Leucemia de Células Pilosas/patologia , Macrófagos/metabolismo , Neoplasias/enzimologia , Neoplasias/patologia , Valores de Referência , Fosfatase Ácida Resistente a Tartarato
18.
Blood ; 85(10): 2839-44, 1995 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-7742545

RESUMO

Tartrate-resistant acid phosphatase (TRAcP) has been an indispensible marker for hairy cell leukemia (HCL) for over two decades. However, the traditional TRAcP cytochemical stain cannot be performed effectively on sections of paraffin-embedded tissues that are important resources for histopathologic evaluation in diagnosis and treatment of HCL. Wide variation in expression of TRAcP activity by hairy cells (HCs) within and among patients is an interesting biologic phenomenon that has not been explained and can cause some diagnostic uncertainty as well. To solve the problem of staining TRAcP in paraffin sections and to begin to address the questions of variable TRAcP expression in HCL, we developed a monoclonal antibody to TRAcP, 9C5, for immunohistochemical identification of HCs. In smears of blood and bone marrow, immunocytochemistry of TRAcP using 9C5 was as specific but slightly less sensitive than direct cytochemical staining of enzymatic activity. In paraffin sections of spleen and bone marrow from HCL patients, immunohistochemistry with 9C5 stained the HCs with high sensitivity and specificity and clearly showed the characteristic diffuse infiltration by HCs. Other cells noted to stain strongly with 9C5 were occasional macrophages in bone marrow smears and osteoclasts and occasional tissue macrophages in paraffin sections. These are cells known to express abundant TRAcP activity. Immunohistochemistry with anti-TRAcP monoclonal antibody 9C5 may have utility as an added option in the diagnosis of HCL, as a means to evaluate residual disease in HCL patients undergoing new treatments, and as a way to address questions regarding variable expression of TRAcP activity by HCs within and among patients with HCL. Also, 9C5 has potential as a reagent for the immunoassay of bone-derived serum TRAcP in patients with certain bone diseases and cancers with bone metastasis.


Assuntos
Fosfatase Ácida/análise , Isoenzimas/análise , Leucemia de Células Pilosas/diagnóstico , Fosfatase Ácida/imunologia , Anticorpos Monoclonais/imunologia , Biomarcadores Tumorais , Humanos , Imuno-Histoquímica/métodos , Leucemia de Células Pilosas/enzimologia , Fosfatase Ácida Resistente a Tartarato
19.
Hematol Oncol Clin North Am ; 8(4): 665-81, 1994 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-7961285

RESUMO

With new understanding in the biology of hematologic diseases and the rapid progress in their treatment, it is essential to diagnose and classify the various hematologic diseases with great precision. In recent years, the cell type-specific markers have been used successfully in cell identification for more precise diagnosis and classification of neoplastic diseases. The cytochemical method for cell identification is simple and is most useful for disorders of granulocytes, monocytes/histiocytes, hairy cells, and mast cells. The immunochemical methods are particularly useful in identifying the cell types, the clonality, and the maturation of cells in the lymphoid disorders. They are also helpful in the diagnosis of disorders of the erythroid and megakaryocytic cells, as well as nonhematopoietic neoplasms with marrow involvement.


Assuntos
Doenças Hematológicas/diagnóstico , Histocitoquímica , Imuno-Histoquímica , Doença Aguda , Doença Crônica , Humanos , Leucemia/diagnóstico , Síndromes Mielodisplásicas/diagnóstico , Transtornos Mieloproliferativos/diagnóstico
20.
Diagn Cytopathol ; 9(4): 448-52, 1993 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-8261853

RESUMO

Fine-needle aspiration (FNA) of the lymph node was done in five patients with histiocytic necrotizing lymphadenitis (Kikuchi's disease). In four patients, the aspirates were found to have many small and large atypical lymphocytes, some reactive, phagocytic histiocytes, and intense extracellular debris. Neutrophils, plasma cells, or multinucleated giant cells were not seen. These cytologic findings were considered diagnostic for Kikuchi's disease. In one patient, the aspirate did not show significant histiocytosis or tissue necrosis and was considered nondiagnostic. In patients with both typical clinical features and characteristic cytologic findings in the lymph node aspirates, FNA of the lymph node alone will suffice for diagnosis. In those patients with typical clinical features but nondiagnostic findings in the FNA aspirates, the diagnosis of Kikuchi's disease may have to be established either on repeated nodal FNA or on lymph node biopsy.


Assuntos
Linfonodos/patologia , Linfadenite/patologia , Adolescente , Adulto , Biópsia por Agulha , Feminino , Humanos , Necrose
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...