RESUMO
Activity of placental alkaline phosphatase (PAP) was studied in blood serum of 53 healthy persons and of 72 oncologic patients, using solid-phase immunoenzymatic analysis with polyclonal antibodies towards PAP. The enzyme was detected both in blood serum of healthy persons and of oncologic patients. The blood serum under study was preheated at 65 degrees in order to inactivate the intestinal phosphatase--the only isoenzyme cross-reacting with antibodies to thermostable PAP. This controlled heating treatment decreased distinctly the possibility of pseudo-positive reactions. The limiting values of the PAP activity were about 0.15 un/L in blood sera of healthy persons. Higher values were considered as an evidence of pathological state. After screening analysis of blood sera from patients with various forms of malignant tumors the PAP activity above 0.15 un/L was observed in 20% of the patients; the enzymatic activity exceeded these values in 54% of patients with ovary carcinoma. The data obtained suggest that the procedure developed as an adequate means for estimation of thermostable PAP isoenzymes as well as that the rate of PAP activity might serve as marker of malignancy in ovary and testis carcinomas independently on level of total activity of alkaline phosphatase.
Assuntos
Fosfatase Alcalina/sangue , Biomarcadores Tumorais/análise , Neoplasias Ovarianas/diagnóstico , Placenta/enzimologia , Neoplasias Testiculares/diagnóstico , Adulto , Feminino , Humanos , Técnicas Imunoenzimáticas , Masculino , Pessoa de Meia-IdadeRESUMO
Four different chromatographic methods of IgG isolation from rabbit antisera to placental alkaline phosphatase (HPAP) have been compared. The antibodies were obtained by ion-exchange chromatography and affinity chromatography on protein-A-sepharose, on the sepharose with immobilized antigen. IgG samples were characterized by the content of specific antibodies to HPAP and checked in enzyme immunoassay (EIA). IgG purified on immobilized antigen were found to be the optimal both from the point of view of the specific antibodies content and EIA sensitivity, but satisfactory results could be also obtained with ion-exchange and protein-A-chromatography purified IgG. The last two isolation methods are simpler and provide 3-10 ng/ml sensitivity of HPAP detection, which is lower, as compared with the test employing affinity antibodies (1 ng/ml), but allows the detection of HPAP in serum samples.