Targetted localisation and imaging of a murine lymphoma using 131I-labelled monoclonal antibody.

In vivo tumor targetting with radiolabelled monoclonal antibodies is a promising approach for the diagnosis and therapy of tumors. A specific monoclonal antibody (mAb), DLAB was generated to the Dalton's lymphoma associated antigen (DLAA) from Haemophilus paragallinarum-induced spontaneous fusion. In order to study the tumor localisation and biodistribution properties of the monoclonal antibody, scintigraphic studies were performed using the radiolabelled DLAB. 131-labelled DLAB was administered intravenously into Swiss mice bearing Dalton's lymphoma and external scintiscanning was performed at different time intervals. Clear tumor images were obtained which revealed selective and specific uptake of radiolabel and the results were compared with biodistribution data. The radioiodinated monoclonal antibody showed fast tumor uptake which increased significantly to 14.6% injected dose (ID)/g at 12 hr post-injection. Enhanced blood clearance of radioactivity resulted in higher tumor/blood ratio of 5.96 at 48 hr. 131I-labelled DLAB resulted in selective and enhanced uptake of the radioactivity by the tumor compared to the non-specific antibody and the results suggest the potential use of spontaneous fusion for producing specific monoclonal antibodies for tumor detection and therapy.

Factors affecting chemistry of reduction-mediated 99mTc-labelling of monoclonal antibodies and polyclonal human immunoglobulins.

In developing a new method for preparing a radiopharmaceutical for clinical investigation, a thorough understanding of reaction stoichiometry is crucial in optimizing the labelling chemistry. Factors determining labelling efficiency of the 2-mercaptoethanol (2-ME)-mediated 99mTc-labelling of antibody molecules were elucidated using anti-tumor monoclonal antibodies of different IgG subclasses (i.e. IOR-CEA(IgG1), M170(IgG1), 3F8(IgG3) and EMD (IgG2a)) and polyclonal human immunoglobulins (Sandoglobulin). Antibodies which were sensitive to 2-ME reduction (i.e. required 500-1000 molar excess of 2-ME) could tag 99mTc with high efficiency since they possessed abundant reactive sites (i.e. sulfydryl groups) for 99mTc binding. Reduction sensitivity of antibodies was unlikely to be affected by IgG subclass and could be rated as follows: Sandoglobulin > IOR-CEA > 3F8 > M170 > EMD. Concentrations of the reduced antibodies for effective labelling appeared to be related to the reduction sensitivity, i.e. 0.2, 0.4 and 0.6 mg/ml were required for labelling of IOR-CEA, 3F8 and M170 respectively. In addition, susceptibility to 2-ME reduction seemed to reflect the rate of antibody labelling. For 2-ME resistant molecules, i.e. M170 and EMD, successful labelling could be achieved by using a slow 99mTc reducing agent such as SnCl2 instead of SnF2 which reacted more rapidly. Since 2-ME generates reactive sulfhydryl groups that are distal to antigen binding sites, the immunoreactivity of the modified antibody was not affected by the effect of reduction.

Valor diagnóstico de los anticuerpos anti-Ro (SS-A) en lupus eritematoso sistémico ANA positivo / Diagnostic value of anti-Ro(SS-A) antibodies in ANA-positive systemic lupus erythematosus

Objetivo: Definir la participación de los anticuerpos anti-Ro(SS-A) en las lesiones del lupus eritematoso sistémico (LES). Material y método: Se compararon características demográficas, clínicas y serológicas en 33 pacientes con LES positivos y negativos para los anticuerpos anti-Ro(SS-A). Los anticuerpos antinucleares totales (ANA) fueron determinados por inmunofluorescencia indirecta sobre cortes de hígado de rata; las especificidades antinucleares anti-Ro(SS-A), anti-Sm/RNP, anti-Scl-70, y anti-Jo-1 fueron evaluados por ensayos inmunoenzimáticos, y los anti-DNA de doble cadena (dc) por radioinmunoanálisis. También se determinó la presencia del factor reumatoide (FR, aglutinación de látex), y de los inmunocomplejos circulantes por los métodos de precipitación con polietilenglicol (ICC-PEG) y el inmunoenzimático en base a la unión de C1q (ICC-C1q). Resultados: Todos los pacientes resultaron positivos para los ANA y el 30.3 por ciento(10/33) fueron positivos para los anticuerpos anti-Ro(SS-A). La distribución de los anticuerpos anti-Ro(SS-A) no dependió de la edad, sexo y raza de los pacientes con LES. El daño renal histológico, las lesiones cutáneas, las citopenias hematológicas, la artritis y los síntomas neurológicos no mostraron asociación con la presencia de los anticuerpos anti-Ro(SS-A). Los anti-Ro(SS-A) tampoco resultaron indicativos de la actividad de la enfermedad lúpica. En el plano serólogico, se encontró correlación positiva entre los anticuerpos anti-Ro(SS-A) y las cifras de anticuerpos anti-DNAdc (t = -3.65 p< 0.01), y la presencia de ICC-PEG (x2 = 3.94 p< 0.05). Conclusiones: La relación encontrada entre los anticuerpos anti-Ro(SS-A) y los anti-DNAdc subraya el papel diagnóstico de los anti-Ro(SS-A) para el LES, aunque su presencia no se acompaña de caracteristicas clínicas distintivas del LES en pacientes ANA positivos.

La gammagrafía en la evaluación de pacientes pediátricos con fiebre de origen desconocido / Scintigraphic explorations in children with fever unknown origin

La localización de focos infecciosos en pacientes con fiebre de origen desconocido (FOD) es frecuentemente difícil y muchas veces insatisfactoria. Las exploraciones gammagráficas diagnósticas con leucocitos marcados en pacientes pediátricos con FOD tienen un gran potencial clínico para el diagnóstico de focos infecciosos sospechados o insospechados como la causa de la FOD y deben ser consideradas como las exploraciones diagnósticas de elección en ausencia de signos que sugieran la localización anatómica del proceso infeccioso así como cuando las imágenes obtenidas mediante la tomografía axial computada o el ultrasonido no demuestren anormalidades y cuando, por la naturaleza o evolución de las lesiones inflamatorias/infecciosas, éstas no tengan bordes definidos. Una exploración gammagráfica con leucocitos marcados que no demuestra la existencia de alguna anormalidad prácticamente excluye la existencia de focos inflamatorios activos. Las exploraciones gammagráficas con leucocitos marcados con In-111 y Tc-99m-HMPO, así como la radioinmunolocalización de focos infecciosos con el anticuerpo monoclonal antigranulocitos Tc-99m-BW250/183 en pacientes con FOD pueden llegar a ser más económicas que otras exploraciones radiológicas, pudiendo realizarse sin ningún problema en pacientes postquirúrgicos. Así mismo, el diagnóstico puede realizarse tempranamente dado que no es necesaria la consolidación del foco infeccioso para que sea radiológicamente detectable. Los procesos malignos metastásicos óseos y la espondilitis pueden ser también diagnosticados tempranamente por los hallazgos relativamente fotopénicos característicos encontrados en la médula ósea

Reduction-mediated 99mTc-labeling of antitumor monoclonal antibodies: effect of increasing specific activity on antibody binding kinetics.

Reduction-mediated 99mTc-labeling of antibodies has gained widespread acceptance in preparation of tumor imaging agents. Increased specific activity to enhance detection signals has raised the question of whether such an attempt would cause change in antibody binding kinetics. To answer this question, two antitumor monoclonal antibodies, i.e. IOR-CEA (IgG1) and EMD (IgG2a) were labeled with 99mTc to yield specific activities ranging from 549-4414 MBq/mg. Regression analysis of the binding data revealed that the binding kinetics of IOR-CEA were shifted from monovalent to bivalent binding upon increasing the specific activities. This phenomenon of affinity enhancement was confirmed by the dissociation study where we found soluble CEA had greater difficulty in extracting the cell-bound IOR-CEA labeled at higher specific activity. The bivalent bindings was further supported by the finding that IOR-CEA with higher specific activities delivered less than expected radioactivity to tumor targets despite their immunoreactivities being well preserved. For EMD, the kinetics seemed to be shifted from bivalent to monovalent interaction. At higher specific activities, adverse changes in immunoreactivity were recognized. Breakage of EMD into 99mTc-Fab fragments was likely to occur and was supported by the observation that EMD delivered more than expected radioactivities to target cells upon increasing specific activity. Precaution should be taken when one deals with high specific activity labeling since this might alter the antibody binding kinetics either favorably or unfavorably.

Avaliaçäo clínica de dois localizadores apicais eletrônicos de terceira geraçäo / Clínical evaluation of two third-generation electronic apical finders

A proposta desta investigaçäo foi comparar in vivo o comprimento real de dentes portadores de polpa viva com as medida fornecidas pelos localizadores apicais eletrônicos. Dezoito dentes unirradiculares tiveram as suas medidas oferidas pelos aparelhos, observando a interferência de líquido de Dakin, soluçäo de Milton, água oxigenada 10 volumes e associaçäo tergentol-furacin, usados como substâncias irrigadoras. Os dentes foram extraídos e os resultados apontaram, para o Grupo I - Apit, 38,8 por cento de medidas coincidentes com o comprimento real dos espécimes e, para o Grupo II - Endosonic, 27,7 por cento. A porcentagem das medidas encontradas além do forame apical foi de 38,9 por cento para Grupo I e, para o Grupo II, 50,0 por cento. Já a observaçäo da posiçäo aquém forame produziu resultados idênticos para ambos os grupos (22,3 por cento). Dentro das condiçöes experimentais, foi possível concluir que as substâncias irrigadoras utilizadas näo proporcionaram alteraçöes nas medidas obtidas

Heterogeneity of macrophage populations: antibodies detecting various populations of macrophages

Two groups of antibodies against mouse resident alveolar and peritoneal macrophages have been raised individually in rabbits by immunization with whole cell suspensions of lavaged alveolar and peritoneal macrophages. The whole antisera reacted positively with neutrophils and lymphocytes. However, upon extensive absorptions with these cells the activity became restricted to macrophages. The immunoglobulins were purified up to 8 fold by repeated precipitations with saturated ammonium sulphate. The antibodies were labeled with fluorescein isothiocyanate and further used. The specificity of antibodies was confirmed by direct and indirect immunofluorescence and complement-dependent cytotoxicity. In the cytolysis of target cells in the presence of complement, the antibodies could lyse more than 80% of cells at dilutions as low as 1:512